1 Supplementary Figures and Tables

2

3 for manuscript:

4 Time course analysis of the brain transcriptome during transitions between brood care and 5 reproduction in the clonal raider ant

6 Romain Libbrecht*1,2, Peter R. Oxley*1,3, Daniel J. C. Kronauer1

7 *co-first authors

8 1 Laboratory of Social Evolution and Behavior, The Rockefeller University, 1230 York Avenue, New York, 9 NY 10065, USA

10 2 Institute of Organismic and Molecular Evolution, Johannes Gutenberg University, Johannes-von-Müller- 11 Weg 6, 55128 Mainz, Germany

12 3 Samuel J. Wood Library, Weill Cornell Medicine, 1300 York Avenue, New York, NY 10065, USA

13 Correspondence: [email protected]

14 [email protected]

15

A B C 1.5 RC12 Number of with more 0.0 than 2-fold change in expression

RC06 RC12 RB12 BR96 RB06 BC06 RB24 BR06 RC06 12- through 96-hour time points RB48 RB06 BR96 BC06 RB96 BR12 BR48 BC12 564 BC12 BR06 BR24 BR12 BR48 RB96 BR24 RB12 RB24 18 RB48 16 34 6-hour BR 0 1 4 6-hour RB 16 RC12 BR96 BC06 BR06 RC06 RB06 RB96 BC12 BR12 BR48 BR24 RB12 RB24 RB48 17 Supplementary Figure 1

18 Outlier analysis. PCA and distance map of genes showing greater than 2-fold change in 19 expression. A) PCA plot of brood-swap and control samples. Clustering was based on the mean 20 expression of each group, for 967 genes with more than 2-fold change in expression 21 between samples. Percentages on each axis indicate the proportion of variance explained by 22 the indicated principal component. The color of each sample indicates the expected similarity to 23 the control samples; dark blue indicates reproductive phase, dark green indicates brood care 24 phase. Sample names are as per Figure 1. B) Heatmap showing Euclidean distance between all 25 samples, based on all genes with more than 2-fold change in expression, and clustered 26 according to average distances between samples. Blue and green color bar above heatmap 27 indicates similarity to control samples, as in A. Sample names are as per Figure 1. C) Venn 28 diagram showing outcome of eliminating the 6-hour time points. Numbers in the small circles 29 indicate genes with > 2-fold change in expression between 6-hour control and treatment 30 samples in the reproduction to brood care transition (blue) and brood care to reproduction 31 transition (green) (a priori true positives). Red numbers indicate genes that show > 2-fold 32 change in expression after removal of the 6-hour time points. Thus, elimination of the 6-hour 33 samples does not substantially reduce the number of DEGs identified with large expression 34 changes.

35

Brood care to reproduction Reproduction to brood care

● ●

100000 ● − ● ● 100000

− ● ● ● ●● ● ● ●●● ● ● ● ●●●● ● ● ● ● ● ●●●● ● ● ● ● ● ● ● ● ● ●●●●●● ● ● ● ● ● ● ● ● ●●● ●● ● ● ● ● ● ● ● ● ● ● ● ● 150000 ● − 150000 − 200000 − 200000 − BIC BIC 250000 − 250000 BIC scoreBIC BIC scoreBIC − ● 300000 − 300000 − 350000 − 350000

● ● ●●●●●●●●●●●●●●●●●●●●●●●● − ● ● ● ● ● ● ● ● ● ● ● ● ● ● ●

0 20 40 60 80 100 120 0 20 40 60 80 100

Number Gof clusters Number ofG clusters 36 37 Supplementary Figure 2

38 Evaluation of BIC scores for selection of optimal number of clusters. Genes were clustered into 39 all even numbered cluster sizes between 2-120 (brood care to reproduction) or 2-110 40 (reproduction to brood care). The optimal cluster size was determined to be the cluster with the 41 lowest BIC score after stabilization to the plateau seen on the right of each graph. Arrows show 42 the cluster selected for each transition.

43

100%

80%

Edges linking clusters 60% of genes regulated in:

Same direction

40% Opposite direction

20%

0%

Proportion of edges linking clusters between transitions Complete Conserved network network

44 45 Supplementary Figure 3

46 If the same genes regulated both transitions, we would expect network edges that link clusters 47 of genes regulated in opposite direction between transitions to represent a higher proportion of 48 edges in the conserved network (with only non-random connections) compared to the complete 49 network (which includes random connections). Contrary to this expectation, we found a lower 50 proportion in the complete network (χ2 = 22.4, p < 0.00001).

51

BR

RB

GO:0003824 [MF] 'catalytic activity' GO:0004252 [MF] '-type endopeptidase activity' GO:0005216 [MF] 'ion channel activity' GO:0005615 [CL] 'extracellular space' GO:0005319 [MF] 'lipid transporter activity' GO:0005789 [CL] 'endoplasmic reticulum membrane' GO:0005506 [MF] 'iron ion binding' GO:0005834 [CL] 'heterotrimeric G- complex' GO:0006030 [BP] 'chitin metabolic process' GO:0008237 [MF] 'metallopeptidase activity' GO:0005576 [CL] GO:0006508 [BP] '' GO:0015986 [BP] 'ATP synthesis coupled proton 'extracellular region' GO:0006811 [BP] 'ion transport' transport' GO:0052689 [MF] GO:0006869 [BP] 'lipid transport' GO:0035176 [BP] 'social behavior' 'carboxylic ester hydro- GO:0007160 [BP] 'cell-matrix adhesion' GO:0035556 [BP] 'intracellular signal transduction' lase activity' GO:0008061 [MF] 'chitin binding' GO:0043565 [MF] 'sequence-specific DNA binding' GO:0009166 [BP] 'nucleotide catabolic process' GO:0046034 [BP] 'ATP metabolic process' GO:0016705 [MF] 'oxidoreductase activity, acting on paired donors, with incorporation GO:0046933 [MF] 'proton-transporting ATP synthase or reduction of molecular oxygen' activity, rotational mechanism' GO:0016787 [MF] 'hydrolase activity' GO:0055114 [BP] 'oxidation-reduction process' GO:0020037 [MF] 'heme binding' GO:0043169 [MF] 'cation binding'

52 53 Supplementary Figure 4

54 GO terms significantly enriched in clusters enriched for DEGs. Only two of these GO terms were 55 common to both transitions (RB: transition from reproduction to brood care; BR: transition from 56 brood care to reproduction). The diameter of the circles is proportional to the number of 57 enriched GO terms.

58

1.5

0.0

RB48

BR12

BR48

BR24

RB96

BC12

RB24

RB12

RC12

BR96

59 RB48 BR12 BR48 BR24 RB96 BC12 RB24 RB12 RC12 BR96 60 Supplementary Figure 5

61 Genes associated with forkhead also segregate with position in the colony cycle. Heatmap 62 showing Euclidean distances between all samples for the 438 genes that contained at least one 63 TFBS for forkhead with a minimum score of 95%. The dendrogram was constructed using the 64 average distances between samples. The blue and green color bar above the heatmap 65 indicates average ovary activation score, as in Figure 2A. Sample names are as per Figure 1.

66

67

68 Supplementary Table 1

69 All 596 DEGs ranked according to p-value (smaller to larger).

70

Rank Gene ID Adjusted p-value Annotation

1 LOC105277456 2.86E-27 protein-L-isoaspartate(D-aspartate) O-methyltransferase 2 LOC105277457 2.09E-24 equilibrative nucleoside transporter 1 3 LOC105278524 5.91E-18 ILP2 4 LOC105281428_Q 1.25E-16 queen vitellogenin 5 LOC105275236 1.75E-16 carboxypeptidase M 6 LOC105284539 2.37E-12 sodium- and chloride-dependent transporter 1 7 LOC105279766 2.72E-12 uncharacterized LOC105279766 8 LOC105279851 2.05E-11 yrdC domain-containing protein 9 LOC105275986 8.19E-11 uncharacterized LOC105275985 10 LOC105276495 1.17E-10 general odorant-binding protein 69a 11 LOC105285597 3.44E-10 uncharacterized LOC105285597 12 LOC105281011 1.12E-09 transferrin 13 LOC105285466 2.11E-09 uncharacterized LOC105285466 14 LOC105283669 7.35E-09 spermine synthase 15 LOC105276905 8.06E-09 neural/ectodermal development factor IMP-L2 16 LOC105275762 1.75E-08 protein aubergine 17 LOC105280170 4.95E-08 PI-PLC X domain-containing protein 2 18 LOC105281012 4.95E-08 uncharacterized LOC105281012 19 LOC105285380 6.73E-08 chitooligosaccharidolytic beta-N-acetylglucosaminidase 20 LOC105277400 2.52E-07 aldose 1-epimerase 21 LOC105279907 4.37E-07 uncharacterized LOC105279907 22 LOC105285376 4.71E-07 uncharacterized LOC105285376 23 LOC105280731 5.52E-07 inositol monophosphatase 1 24 LOC105275113 8.88E-07 pancreatic lipase-related protein 2-like 25 LOC105276066 1.03E-06 uncharacterized LOC105276066 26 LOC105275269 1.09E-06 uncharacterized LOC105275269 27 LOC105276731 1.09E-06 uncharacterized LOC105276731 28 LOC105283821 1.09E-06 nucleolar protein 6 29 LOC105278211 1.54E-06 centromere protein I 30 LOC105276018 1.94E-06 peroxisomal hydratase-dehydrogenase-epimerase 31 LOC105277056 1.94E-06 neuroparsin-A 32 LOC105288064 3.80E-06 uncharacterized LOC105288064 33 LOC105281928 5.80E-06 L-asparaginase 1 34 LOC105285607 6.17E-06 probable ribosome production factor 1 35 LOC105286129 6.88E-06 homeobox protein TGIF2LX 36 LOC105280465 9.64E-06 uncharacterized LOC105280465 37 LOC105281340 9.64E-06 sphingomyelin phosphodiesterase 38 LOC105283223 9.64E-06 putative mediator of RNA polymerase II transcription subunit 26 39 LOC105279511 9.67E-06 uncharacterized LOC105279511 40 LOC105281817 1.08E-05 uncharacterized LOC105281817 41 LOC105286755 1.08E-05 peptidyl-prolyl cis-trans isomerase 5 42 LOC105282988 1.18E-05 sodium/potassium-transporting ATPase subunit alpha 43 LOC105277203 1.24E-05 RNA polymerase I-specific transcription initiation factor RRN3 44 LOC105281008 1.45E-05 glutamate receptor ionotropic 45 LOC105286732 1.65E-05 anillin 46 LOC105281609 1.97E-05 small subunit processome component 20 homolog 47 LOC105285940 2.17E-05 THAP domain-containing protein 3 48 LOC105281708 2.20E-05 uncharacterized LOC105281708 49 LOC105281726 2.43E-05 ferrochelatase 50 LOC105280087 2.72E-05 protein regulator of cytokinesis 1 51 LOC105279867 2.74E-05 growth factor receptor-bound protein 14 52 LOC105283177 4.99E-05 glutathione S-transferase 53 LOC105276918 5.07E-05 14-3-3 protein epsilon 54 LOC105285920 5.07E-05 vegetative cell wall protein gp1 55 LOC105276675 5.36E-05 NK-tumor recognition protein 56 LOC105284285 5.36E-05 high affinity cAMP-specific and IBMX-insensitive 3' 57 LOC105280553 5.43E-05 uncharacterized LOC105280553 58 LOC105279540 5.72E-05 N-sulphoglucosamine sulphohydrolase 59 LOC105282075 5.77E-05 structural maintenance of protein 5 60 LOC105282643 7.25E-05 PDZ and LIM domain protein 4 61 LOC105283185 7.50E-05 kinetochore-associated protein 1 62 LOC105287151 8.77E-05 D-beta-hydroxybutyrate dehydrogenase 63 LOC105286142 9.53E-05 cyclin-dependent kinases regulatory subunit 64 LOC105277367 0.000105047 twinkle protein 65 LOC105283914 0.000105319 mitochondrial import receptor subunit TOM70 66 LOC105282416 0.000109464 uncharacterized LOC105282416 67 LOC105277606 0.000118589 titin 68 LOC105285641 0.000129828 EP300-interacting inhibitor of differentiation 3 69 LOC105286010 0.000132779 major facilitator superfamily domain-containing protein 12 70 LOC105287874 0.000145545 NADH-ubiquinone oxidoreductase 49 kDa subunit 71 LOC105276419 0.000151946 uncharacterized protein C18orf63 72 LOC105282036 0.000151946 G2/mitotic-specific cyclin-B 73 LOC105282150 0.000151946 ATP synthase subunit alpha 74 LOC105275344 0.000168939 ejaculatory bulb-specific protein 3 75 LOC105276174 0.000175408 sodium/potassium-transporting ATPase subunit beta-2 76 LOC105278710 0.000176586 cyclin-dependent kinase 1 77 LOC105281832 0.000181294 nucleolar protein 58 78 LOC105279728 0.000206086 uncharacterized LOC105279728 79 LOC105279314 0.000213503 trypsin inhibitor 80 LOC105287013 0.000213503 kinesin-like protein KIF20B 81 LOC105280074 0.000220032 uncharacterized LOC105280074 82 LOC105275711 0.000229201 TATA box-binding protein-associated factor RNA polymerase I subunit B 83 LOC105280641 0.000231988 serine/threonine-protein kinase VRK1 84 LOC105279847 0.000235166 uncharacterized LOC105279847 85 LOC105282680 0.00025994 glucosylceramidase 86 LOC105279763 0.000261952 glutamate receptor ionotropic 87 LOC105279565 0.000268443 transcriptional regulator ATRX 88 LOC105288139 0.000292672 uncharacterized LOC105288139 89 LOC105279557 0.000301454 kinesin-like protein KIF19 90 LOC105283374 0.000301454 poly(A) RNA polymerase gld-2 homolog A 91 LOC105286171 0.000319218 contactin-4 92 LOC105285834 0.000381783 ATP-dependent DNA helicase Q5 93 LOC105274638 0.000388134 kinesin-like protein KIF18A 94 LOC105284189 0.000389058 fidgetin-like protein 1 95 LOC105285614 0.000413081 glycerol-3-phosphate dehydrogenase [NAD(+)] 96 LOC105274740 0.000434313 putative leucine-rich repeat-containing protein DDB_G0290503 97 LOC105277839 0.000434313 ribosomal protein S6 kinase beta-1 98 LOC105281660 0.000442336 interleukin enhancer-binding factor 2 99 LOC105276143 0.000465598 NGFI-A-binding protein homolog 100 LOC105277886 0.000499745 glutamate-gated chloride channel 101 LOC105279186 0.000549461 probable RNA-binding protein 19 102 LOC105280308 0.000549461 Fanconi anemia group D2 protein homolog 103 LOC105280529 0.000549461 uncharacterized LOC105280529 104 LOC105282076 0.000549461 small -rich protein 1 105 LOC105286277 0.000549461 palmitoyltransferase ZDHHC3 106 LOC105275889 0.000572928 uncharacterized LOC105275889 107 LOC105275854 0.000582604 annulin 108 LOC105276290 0.000582604 nuclear pore complex protein Nup88 109 LOC105280511 0.000585378 glucose dehydrogenase [FAD 110 LOC105286952 0.000608702 uncharacterized LOC105286952 111 LOC105285125 0.000611241 endoribonuclease Dicer 112 LOC105282639 0.000620718 lachesin 113 LOC105275876 0.000649572 diuretic hormone class 2 114 LOC105279470 0.000649572 signal peptidase complex subunit 1 115 LOC105277844 0.000663379 cytochrome P450 CYP12A2 116 LOC105281212 0.000702038 protein rhomboid 117 LOC105283509 0.000717333 zinc finger BED domain-containing protein 1-like 118 LOC105276079 0.000720172 uncharacterized LOC105276079 119 LOC105286126 0.000720172 succinate dehydrogenase [ubiquinone] iron-sulfur subunit 120 LOC105283079 0.000755324 midasin 121 LOC105276628 0.000766774 uncharacterized MFS-type transporter C09D4.1 122 LOC105278567 0.000766774 zinc finger BED domain-containing protein 1 123 LOC105283043 0.000775008 uncharacterized LOC105283043 124 LOC105283843 0.000775008 phosphatidylinositol transfer protein alpha isoform 125 LOC105286179 0.000775008 uncharacterized LOC105286179 126 LOC105288016 0.000775008 synaptic vesicular amine transporter 127 LOC105276979 0.000791533 isocitrate dehydrogenase [NAD] subunit beta 128 LOC105286439 0.000826073 uncharacterized LOC105286439 129 LOC105277700 0.000859212 actin 130 LOC105274628 0.000891361 uncharacterized LOC105274628 131 LOC105280285 0.00089511 uncharacterized protein CG1785 132 LOC105288014 0.000939241 HEAT repeat-containing protein 1 133 LOC105281721 0.000973321 uncharacterized LOC105281721 134 LOC105283157 0.000973321 carbonic anhydrase 2 135 LOC105283626 0.000973321 uncharacterized LOC105283626 136 LOC105287187 0.000973321 non-homologous end-joining factor 1 137 LOC105276512 0.00103448 S-methyl-5'-thioadenosine phosphorylase 138 LOC105277421 0.00103448 protein timeless homolog 139 LOC105279939 0.001075557 lymphokine-activated killer T-cell-originated protein kinase 140 LOC105276690 0.001096309 folylpolyglutamate synthase 141 LOC105275469 0.001107455 sarcalumenin 142 LOC105285890 0.001107455 uncharacterized LOC105285890 143 LOC105283266 0.001121027 lymphoid-specific helicase 144 LOC105276925 0.001123963 protein seele 145 LOC105287204 0.001123963 twitchin 146 LOC105276190 0.001155579 protein brambleberry 147 LOC105279559 0.001155579 glycine receptor subunit alpha-2-like 148 LOC105280561 0.001172392 calmodulin 149 LOC105278661 0.001211963 ribonucleases P/MRP protein subunit POP1 150 LOC105278082 0.001262696 ADP-ribosylation factor-like protein 13B 151 LOC105284604 0.001354385 general odorant-binding protein 83a 152 LOC105287093 0.001396307 uncharacterized LOC105287093 153 LOC105284249 0.001459572 ribosomal RNA processing protein 1 homolog 154 LOC105284986 0.001480837 ribonucleoside-diphosphate reductase large subunit 155 LOC105283698 0.001521001 pyridoxine-5'-phosphate oxidase 156 LOC105282844 0.001531753 uncharacterized LOC105282844 157 LOC105276853 0.001584309 uncharacterized LOC105276853 158 LOC105287282 0.001590122 nucleotide exchange factor SIL1 159 LOC105285386 0.00171934 nucleoporin Nup37 160 LOC105275345 0.001775821 ejaculatory bulb-specific protein 3 161 LOC105287530 0.001775821 WD repeat-containing protein 46 162 LOC105279696 0.001802057 uncharacterized LOC105279696 163 LOC105280503 0.001861233 glucose dehydrogenase [FAD 164 LOC105286339 0.001892283 RRP12-like protein 165 LOC105277377 0.001910982 troponin C 166 LOC105279473 0.001962256 uncharacterized LOC105279473 167 LOC105280338 0.001962256 CAD protein 168 LOC105277875 0.00197776 RNA-binding protein 40 169 LOC105281005 0.001994437 tubulin epsilon chain 170 LOC105287193 0.002024743 uncharacterized LOC105287193 171 LOC105285809 0.002063242 putative helicase MOV-10 172 LOC105285704 0.002174642 MATH and LRR domain-containing protein PFE0570w 173 LOC105279590 0.002190403 uncharacterized LOC105279590 174 LOC105279671 0.002197433 U3 small nucleolar RNA-associated protein 14 homolog A 175 LOC105285187 0.002212392 uncharacterized LOC105285187 176 LOC105277039 0.002257698 CAPA peptides 177 LOC105277876 0.002257698 adenylate kinase isoenzyme 1 178 LOC105280242 0.002257698 uncharacterized LOC105280240 179 LOC105283045 0.002257698 uncharacterized LOC105283045 180 LOC105283429 0.002257698 uridine phosphorylase 1 181 LOC105286263 0.002257698 ribonuclease P protein subunit p40 182 LOC105278693 0.002374317 troponin C-like 183 LOC105286776 0.002374317 ribonucleoside-diphosphate reductase subunit M2 B 184 LOC105283233 0.002468175 fibrillin-1 185 LOC105284536 0.002470555 uncharacterized LOC105284536 186 LOC105287615 0.002622097 nidogen-2 187 LOC105276738 0.002627827 KH domain-containing protein akap-1 188 LOC105284373 0.002714911 myosin light chain alkali 189 LOC105287207 0.002888659 kinesin-like protein Klp61F 190 LOC105285677 0.003023458 uncharacterized LOC105285677 191 LOC105285139 0.003043257 serine/threonine-protein kinase Chk2 192 LOC105277197 0.00317964 putative sodium-dependent multivitamin transporter 193 LOC105277432 0.003195212 V-type proton ATPase subunit G 194 LOC105287079 0.003237844 protein CNPPD1 195 LOC105280404 0.003258013 tankyrase-1 196 LOC105286412 0.003258013 uncharacterized LOC105286412 197 LOC105283216 0.003309992 mycosubtilin synthase subunit C 198 LOC105274480 0.003328645 zwei Ig domain protein zig-8 199 LOC105280897 0.003328645 titin 200 LOC105278020 0.003375831 origin recognition complex subunit 2 201 LOC105283438 0.003471681 tyrosine--tRNA ligase 202 LOC105278746 0.003508497 periodic tryptophan protein 1 homolog 203 LOC105284104 0.003563795 uncharacterized LOC105284104 204 LOC105285852 0.003563795 nucleoside diphosphate kinase 6 205 LOC105283164 0.003564436 protein FAM60A 206 LOC105284641 0.003564436 uncharacterized LOC105284641 207 LOC105286766 0.003564436 dynein heavy chain 7 208 LOC105282386 0.003627323 tRNA-specific adenosine deaminase 2 209 LOC105281770 0.003726834 phosphoglycerate mutase 1 210 LOC105286356 0.003733126 R3H and coiled-coil domain-containing protein 1 211 LOC105285273 0.003745615 UDP-glucuronosyltransferase 2B1 212 LOC105276762 0.003752298 chaoptin 213 LOC105286471 0.003786951 uncharacterized LOC105286471 214 LOC105286745 0.003786951 serine proteinase stubble 215 LOC105287773 0.003786951 uncharacterized LOC105287773 216 LOC105281565 0.0039943 maternal embryonic leucine zipper kinase 217 LOC105281396 0.00400507 angiotensin-converting enzyme 2 218 LOC105278260 0.004033359 tRNA-splicing ligase RtcB homolog 219 LOC105282051 0.004069005 uncharacterized LOC105282051 220 LOC105284988 0.004069005 DNA primase large subunit 221 LOC105277716 0.004092585 facilitated trehalose transporter Tret1 222 LOC105278265 0.004107218 uncharacterized LOC105278265 223 LOC105282754 0.004152694 protein yellow 224 LOC105284776 0.004152694 uncharacterized LOC105284776 225 LOC105275039 0.004204795 uncharacterized LOC105275039 226 LOC105275568 0.004204795 probable pyruvate dehydrogenase E1 component subunit alpha 227 LOC105285003 0.004266796 uncharacterized LOC105285003 228 LOC105276507 0.004334577 notchless protein homolog 1 229 LOC105275184 0.004373376 SWI/SNF-related matrix-associated actin-dependent regulator of chromatin subfamily A-like protein 1 230 LOC105276445 0.004373376 la protein homolog 231 LOC105279818 0.004373376 G2/mitotic-specific cyclin-B3 232 LOC105281245 0.004373376 uncharacterized LOC105281245 233 LOC105285918 0.004373376 probable kinetochore protein NUF2 234 LOC105286949 0.004373376 RNA-binding protein 4.1 235 LOC105286220 0.004374269 nuclear pore complex protein Nup133 236 LOC105278869 0.00442564 uncharacterized LOC105278869 237 LOC105277651 0.004438611 rap1 GTPase-activating protein 1 238 LOC105286343 0.004658075 putative serine protease K12H4.7 239 LOC105283179 0.004746696 carcinine transporter 240 LOC105281685 0.004888436 venom carboxylesterase-6 241 LOC105280491 0.004955404 inosine-5'-monophosphate dehydrogenase 242 LOC105277108 0.004990146 uncharacterized LOC105277108 243 LOC105277299 0.004990146 ionotropic receptor 25a 244 LOC105278142 0.00501094 protein phosphatase methylesterase 1 245 LOC105282563 0.005136377 general transcription factor IIH subunit 1 246 LOC105283753 0.005136377 matrix metalloproteinase-2 247 LOC105287183 0.005136377 uncharacterized LOC105287183 248 LOC105286143 0.005213679 renin receptor 249 LOC105275836 0.005222883 dexamethasone-induced Ras-related protein 1 250 LOC105279586 0.005250814 uncharacterized LOC105279586 251 LOC105284384 0.005250814 protein stum 252 LOC105279197 0.005273064 cyclin-dependent-like kinase 5 253 LOC105277469 0.005407787 tubulin beta chain 254 LOC105285464 0.0055311 calmodulin-binding transcription activator 2 255 LOC105280121 0.00553732 esterase E4 256 LOC105277435 0.005768748 targeting protein for Xklp2 257 LOC105277439 0.005787527 ADP-ribosylation factor-like protein 6 258 LOC105279165 0.005861519 LIM domain and actin-binding protein 1 259 LOC105287714 0.005943601 protein hunchback 260 LOC105274872 0.006089499 V-type proton ATPase subunit B 261 LOC105275324 0.006089499 angio-associated migratory cell protein 262 LOC105286103 0.006237383 glutamate--cysteine ligase catalytic subunit 263 LOC105284633 0.006238679 gliomedin 264 LOC105279018 0.006440394 ras-related and estrogen-regulated growth inhibitor 265 LOC105276398 0.006458399 beta-hexosaminidase subunit beta 266 LOC105276421 0.006458399 reticulon-4-interacting protein 1 267 LOC105277488 0.006458399 neuropeptide CCHamide-2 receptor-like 268 LOC105282470 0.006458399 centromere protein L 269 LOC105284938 0.006505284 telomerase reverse transcriptase-like 270 LOC105281051 0.006549854 protein FAM173B 271 LOC105279471 0.00663211 uncharacterized LOC105279471 272 LOC105281618 0.006925192 uncharacterized LOC105281618 273 LOC105284524 0.0069971 hemocytin 274 LOC105280836 0.00727081 uncharacterized LOC105280836 275 LOC105274893 0.007357978 uncharacterized LOC105274893 276 LOC105286889 0.007357978 uncharacterized LOC105286889 277 LOC105274660 0.007463435 uncharacterized LOC105274660 278 LOC105274901 0.007463435 SH3 domain-binding glutamic acid-rich protein homolog 279 LOC105280103 0.007479243 inositol-tetrakisphosphate 1-kinase 280 LOC105277350 0.007511244 uncharacterized LOC105277350 281 LOC105284074 0.007705767 phospholipase B1 282 LOC105285671 0.007705767 nucleoporin Ndc1 283 LOC105283621 0.007727391 synaptic vesicle glycoprotein 2C 284 LOC105280985 0.007749383 ATP-dependent RNA helicase SUV3 homolog 285 LOC105276851 0.007925953 rho guanine nucleotide exchange factor 10-like protein 286 LOC105277458 0.007925953 DNA replication licensing factor MCM4 287 LOC105281382 0.007996421 deoxynucleoside kinase 288 LOC105279911 0.00811899 sterol O-acyltransferase 1 289 LOC105287024 0.008305523 uncharacterized LOC105287024 290 LOC105275955 0.008312762 DNA topoisomerase 2-binding protein 1-A 291 LOC105278498 0.008312762 neither inactivation nor afterpotential protein G 292 LOC105278931 0.008312762 venom allergen 3 293 LOC105279816 0.008312762 cAMP-specific 3' 294 LOC105285550 0.008312762 heat shock 70 kDa protein cognate 4 295 LOC105287679 0.008312762 kinetochore protein NDC80 homolog 296 LOC105276789 0.008448158 putative methyltransferase NSUN6 297 LOC105286365 0.008459175 uncharacterized LOC105286365 298 LOC105278774 0.008516871 hydroxypyruvate reductase 299 LOC105279012 0.008712235 uncharacterized LOC105279012 300 LOC105283008 0.008867065 uncharacterized LOC105283008 301 LOC105282359 0.008886148 myosin regulatory light chain 2 302 LOC105276807 0.009003188 mitochondrial-processing peptidase subunit beta 303 LOC105286101 0.009100546 uncharacterized LOC105286101 304 LOC105286139 0.009113335 translation elongation factor 2 305 LOC105282872 0.009170765 acylphosphatase-1 306 LOC105285256 0.009279029 copper chaperone for superoxide dismutase 307 LOC105288172 0.009328315 hepatocyte nuclear factor 6-like 308 LOC105281757 0.00959872 type I inositol 1 309 LOC105282576 0.01011098 U3 small nucleolar RNA-interacting protein 2 310 LOC105285936 0.010130592 glutathione S-transferase 1-1 311 LOC105281321 0.010183213 5-phosphohydroxy-L-lysine phospho-lyase 312 LOC105284488 0.010203996 meiosis arrest female protein 1 homolog 313 LOC105277205 0.010456356 aquaporin-12A 314 LOC105278655 0.010905246 ATP synthase lipid-binding protein 315 LOC105284448 0.010964398 uncharacterized LOC105284448 316 LOC105281030 0.010989901 uncharacterized LOC105281030 317 LOC105284205 0.011084639 nucleolar GTP-binding protein 1 318 LOC105280772 0.011298076 uncharacterized LOC105280772 319 LOC105282573 0.011350785 SH2B adapter protein 1 320 LOC105279271 0.011467688 uncharacterized LOC105279271 321 LOC105283206 0.011714415 THAP domain-containing protein 5-like 322 LOC105276279 0.011796248 dynein regulatory complex protein 1 323 LOC105283005 0.012137093 protein lethal(2)essential for life 324 LOC105277437 0.013010433 protein LTV1 homolog 325 LOC105283737 0.013047134 myosin heavy chain 326 LOC105288114 0.013047134 replication factor C subunit 4 327 LOC105277654 0.013069808 uncharacterized LOC105277654 328 LOC105275475 0.013180822 bifunctional methylenetetrahydrofolate dehydrogenase/cyclohydrolase 329 LOC105283200 0.013180822 embryonic polarity protein dorsal 330 LOC105275339 0.01329855 H/ACA ribonucleoprotein complex subunit 4 331 LOC105277860 0.013408944 monocarboxylate transporter 12 332 LOC105277685 0.013431626 solute carrier family 12 member 4 333 LOC105286216 0.01354877 kinesin-like protein KIF23 334 LOC105284533 0.013556775 RISC-loading complex subunit tarbp2 335 LOC105276678 0.013571358 transferrin 336 LOC105281010 0.013571358 uncharacterized LOC105281010 337 LOC105282771 0.013571358 exosome complex component RRP40 338 LOC105276745 0.013910074 pre-rRNA processing protein FTSJ3 339 LOC105281448 0.013910074 twitchin 340 LOC105274926 0.014081898 RNA-binding protein 34 341 LOC105277818 0.014081898 uncharacterized LOC105277818 342 LOC105275821 0.014102391 dnaJ homolog subfamily C member 22 343 LOC105286289 0.014120423 upstream stimulatory factor 2 344 LOC105282390 0.01455199 single-strand selective monofunctional uracil DNA glycosylase 345 LOC105288106 0.01455199 sarcosine dehydrogenase 346 LOC105281668 0.014858442 E3 ubiquitin-protein ligase RFWD2-like 347 LOC105277939 0.015013487 uncharacterized LOC105277939 348 LOC105283516 0.015070799 dual oxidase 2 349 LOC105283205 0.015279418 ATP-dependent RNA helicase Ddx1 350 LOC105282422 0.016180145 cuticle protein 64 351 LOC105274940 0.016450608 homeobox protein prospero 352 LOC105279513 0.016450608 tRNA dimethylallyltransferase 353 LOC105280398 0.016525208 protein will die slowly 354 LOC105279732 0.016535081 chymotrypsin-like elastase family member 2A 355 LOC105287010 0.016601834 uncharacterized LOC105287010 356 LOC105283803 0.016726746 ATP synthase subunit O 357 LOC105278140 0.016770953 BTB/POZ domain-containing protein KCTD16 358 LOC105280581 0.016868053 coiled-coil domain-containing protein 50 359 LOC105281377 0.016956368 uncharacterized LOC105281377 360 LOC105278656 0.017140571 solute carrier family 25 member 40 361 LOC105278757 0.017140571 uncharacterized LOC105278757 362 LOC105287066 0.017519339 DNA ligase 1 363 LOC105287949 0.017530134 protein LLP homolog 364 LOC105283796 0.017845437 ornithine decarboxylase 2-like 365 LOC105278078 0.017864174 uncharacterized LOC105278078 366 LOC105287636 0.017896942 uncharacterized LOC105287636 367 LOC105278194 0.0180767 prolyl 3-hydroxylase OGFOD1 368 LOC105285921 0.018086746 uncharacterized LOC105285921 369 LOC105282866 0.018105674 myb-binding protein 1A 370 LOC105275582 0.018212091 zinc finger protein 275 371 LOC105280537 0.018304925 uncharacterized LOC105280537 372 LOC105275811 0.01861386 ribosome biogenesis protein BOP1 homolog 373 LOC105280125 0.01861386 uncharacterized LOC105280125 374 LOC105281082 0.01861386 uncharacterized LOC105281082 375 LOC105286381 0.018645702 uncharacterized LOC105286381 376 LOC105277441 0.018874679 mucin-2 377 LOC105283628 0.018918359 uncharacterized LOC105283628 378 LOC105280004 0.019061425 dynein assembly factor 3 379 LOC105284779 0.019110966 uncharacterized LOC105284779 380 LOC105280988 0.019550262 ejaculatory bulb-specific protein 3 381 LOC105284119 0.019585838 ribonuclease H-like 382 LOC105280397 0.019846382 DNA-dependent protein kinase catalytic subunit 383 LOC105282816 0.020174174 uncharacterized LOC105282816 384 LOC105276699 0.020382878 uncharacterized LOC105276699 385 LOC105285953 0.020382878 uncharacterized LOC105285953 386 LOC105275455 0.020403346 ankyrin repeat and LEM domain-containing protein 2 387 LOC105277915 0.020403346 probable ATP-dependent RNA helicase DDX17 388 LOC105280774 0.020403346 rho guanine nucleotide exchange factor 3 389 LOC105283366 0.020403346 uncharacterized LOC105283366 390 LOC105285712 0.020403346 tolloid-like protein 2 391 LOC105286720 0.020403346 nucleolar protein 56 392 LOC105282918 0.020409442 putative fatty acyl-CoA reductase CG5065 393 LOC105287587 0.020806822 uncharacterized LOC105287587 394 LOC105274629 0.020916032 uncharacterized LOC105274629 395 LOC105282320 0.020916032 hemicentin-2 396 LOC105276546 0.020954818 uncharacterized LOC105276546 397 LOC105275970 0.02108041 uncharacterized LOC105275970 398 LOC105284165 0.021328675 protein THEM6 399 LOC105285353 0.021510104 uncharacterized LOC105285353 400 LOC105281103 0.021841118 pancreatic triacylglycerol lipase-like 401 LOC105277131 0.022127761 uncharacterized LOC105277131 402 LOC105287812 0.022255454 uncharacterized LOC105287812 403 LOC105284497 0.02226819 intraflagellar transport protein 46 homolog 404 LOC105285007 0.022298915 uncharacterized LOC105285007 405 LOC105277337 0.022352653 tubulin polyglutamylase TTLL4 406 LOC105283527 0.023808013 uncharacterized LOC105283527 407 LOC105285553 0.023808013 uncharacterized LOC105285553 408 LOC105287684 0.023842556 uncharacterized LOC105287684 409 LOC105287283 0.023980629 nucleoplasmin-like protein 410 LOC105277137 0.024077995 protein lethal(2)essential for life 411 LOC105283368 0.024691043 probable ATP-dependent RNA helicase CG8611 412 LOC105284077 0.024988634 juvenile hormone epoxide hydrolase 1-like 413 LOC105279278 0.025129854 lysosomal thioesterase PPT2 homolog 414 LOC105282179 0.025129854 NADH dehydrogenase [ubiquinone] 1 beta subcomplex subunit 2 415 LOC105286407 0.025129854 titin 416 LOC105284466 0.025211033 NADH dehydrogenase [ubiquinone] 1 alpha subcomplex subunit 10 417 LOC105274997 0.025259326 rRNA methyltransferase 2 418 LOC105284252 0.025766983 uncharacterized LOC105284252 419 LOC105280202 0.026009546 glycine-rich cell wall structural protein 420 LOC105276085 0.026393569 uncharacterized LOC105276085 421 LOC105284185 0.026393569 probable RNA-binding protein EIF1AD 422 LOC105285503 0.026393569 protein tyrosine phosphatase domain-containing protein 1 423 LOC105281711 0.026450157 serine/threonine-protein phosphatase PP1-beta catalytic subunit 424 LOC105282531 0.026901222 mitochondrial pyruvate carrier 2 425 LOC105287273 0.026984143 E3 ubiquitin-protein ligase rnf8-A 426 LOC105276433 0.02717383 box A-binding factor 427 LOC105276873 0.02717383 protein tipE 428 LOC105281449 0.02717383 muscle M-line assembly protein unc-89-like 429 LOC105284115 0.027407734 uncharacterized LOC105284115 430 LOC105276534 0.027511855 uncharacterized LOC105276534 431 LOC105282149 0.027531068 guanine nucleotide-binding protein subunit beta-5 432 LOC105285652 0.027531068 proton-coupled transporter-like protein CG1139 433 LOC105287285 0.027531068 zinc transporter ZIP13 homolog 434 LOC105280369 0.02784133 uncharacterized LOC105280369 435 LOC105281610 0.027939891 origin recognition complex subunit 1 436 LOC105285829 0.028011009 guanine nucleotide-binding protein-like 3 homolog 437 LOC105286156 0.028011009 gamma-aminobutyric acid receptor subunit beta-like 438 LOC105285775 0.028055328 uncharacterized LOC105285775 439 LOC105282104 0.028088617 glutamate decarboxylase 440 LOC105277525 0.028172885 uncharacterized LOC105277525 441 LOC105286769 0.028172885 uncharacterized LOC105286769 442 LOC105278343 0.028273197 serine/threonine-protein phosphatase 6 regulatory ankyrin repeat subunit A 443 LOC105284789 0.028448938 uncharacterized LOC105284789 444 LOC105287552 0.028448938 protein snakeskin 445 LOC105275954 0.028521019 cytochrome P450 6B7-like 446 LOC105281037 0.029105349 microtubule-associated protein futsch 447 LOC105279447 0.029260078 poly(ADP-ribose) glycohydrolase ARH3 448 LOC105284794 0.029260078 uncharacterized LOC105284794 449 LOC105284925 0.029260078 chloride channel protein 2 450 LOC105276504 0.029409338 calcineurin subunit B type 2 451 LOC105278482 0.029581681 E3 ubiquitin-protein ligase TRAIP 452 LOC105278101 0.030503559 CCA tRNA nucleotidyltransferase 1 453 LOC105275507 0.03071899 PDZ and LIM domain protein 3 454 LOC105279588 0.030725831 activating signal cointegrator 1 complex subunit 3 455 LOC105286287 0.031079663 uncharacterized LOC105286287 456 LOC105288201 0.031079663 uncharacterized LOC105288201 457 LOC105284128 0.031224018 sialin 458 LOC105279871 0.031406262 oxysterol-binding protein-related protein 2 459 LOC105278676 0.031605854 uncharacterized LOC105278676 460 LOC105275286 0.031779213 innexin inx3 461 LOC105279699 0.03198123 prisilkin-39-like 462 LOC105278724 0.032342333 fatty acid synthase-like 463 LOC105278934 0.032342333 exonuclease 3'-5' domain-containing protein 2 464 LOC105285644 0.032342333 rootletin 465 LOC105287695 0.032342333 serine/threonine-protein kinase polo 466 LOC105278009 0.032494456 diuretic hormone 44 467 LOC105281176 0.032494456 uncharacterized LOC105281176 468 LOC105287539 0.032494456 protein vein 469 LOC105286140 0.032710688 zinc finger protein 474 470 LOC105288078 0.032728271 WD repeat-containing protein 43 471 LOC105278695 0.033077184 filamin-A 472 LOC105282856 0.033177527 N-acetyltransferase ESCO2 473 LOC105285465 0.033323481 AP-2 complex subunit mu 474 LOC105275769 0.033494647 uncharacterized LOC105275769 475 LOC105274965 0.0335862 cyclin-dependent kinase-like 4 476 LOC105279598 0.0335862 glutamate dehydrogenase 477 LOC105276193 0.034334734 uncharacterized LOC105276193 478 LOC105277743 0.034334734 trimethylguanosine synthase 479 LOC105287116 0.034334734 la-related protein 7 480 LOC105281786 0.034343523 hyaluronan mediated motility receptor 481 LOC105285243 0.034343523 lysozyme 482 LOC105277206 0.034521091 sodium- and chloride-dependent GABA transporter ine 483 LOC105285400 0.034714853 gem-associated protein 5 484 LOC105287758 0.034740117 uncharacterized LOC105287758 485 LOC105286522 0.034969332 alkylglycerol monooxygenase-like 486 LOC105279786 0.035783651 1-acyl-sn-glycerol-3-phosphate acyltransferase gamma 487 LOC105282695 0.035783651 surfeit protein 1 488 LOC105282710 0.035783651 protein penguin 489 LOC105283615 0.035783651 serine protease nudel 490 LOC105284985 0.035783651 putative uncharacterized protein DDB_G0271606 491 LOC105277279 0.035909715 probable citrate synthase 2 492 LOC105282173 0.036134281 homeotic protein ultrabithorax 493 LOC105283611 0.036243182 allatostatin A 494 LOC105277912 0.036320028 translation initiation factor eIF-2B subunit epsilon 495 LOC105281216 0.036320028 F-box only protein 21 496 LOC105285869 0.036539819 ubiquitin-conjugating enzyme E2 H 497 LOC105287550 0.036680683 serine/arginine repetitive matrix protein 1 498 LOC105280031 0.036693551 ankyrin repeat 499 LOC105280990 0.036693551 nanos homolog 3 500 LOC105281197 0.036849698 uncharacterized LOC105281197 501 LOC105277728 0.036980806 uncharacterized LOC105277728 502 LOC105279536 0.036980806 uncharacterized LOC105279536 503 LOC105286284 0.036980806 uncharacterized LOC105286284 504 LOC105274442 0.03720635 selenide 505 LOC105277683 0.03720635 bromodomain-containing protein 7 506 LOC105278758 0.03720635 actin 507 LOC105284134 0.03720635 uncharacterized LOC105284134 508 LOC105279916 0.037236442 major facilitator superfamily domain-containing protein 6-A 509 LOC105280171 0.037236442 uncharacterized LOC105280171 510 LOC105284746 0.037236442 negative elongation factor B 511 LOC105275435 0.03744624 alpha-(1 512 LOC105284181 0.037636133 peptidyl-prolyl cis-trans isomerase D 513 LOC105275321 0.037869617 adenosylhomocysteinase 514 LOC105275517 0.03796665 probable tubulin polyglutamylase TTLL2 515 LOC105286503 0.03796665 decaprenyl-diphosphate synthase subunit 2 516 LOC105287422 0.037977018 probable cation-transporting ATPase 13A3 517 LOC105275301 0.038090542 uncharacterized protein CG3556 518 LOC105276468 0.038090542 uncharacterized LOC105276468 519 LOC105277551 0.038090542 glycine N-methyltransferase 520 LOC105278540 0.038090542 dipeptidase 1 521 LOC105279249 0.038090542 ran GTPase-activating protein 1 522 LOC105281707 0.038090542 uncharacterized LOC105281707 523 LOC105285645 0.038090542 uncharacterized LOC105285645 524 LOC105287548 0.038090542 uncharacterized LOC105287548 525 LOC105287790 0.038090542 uncharacterized LOC105287790 526 LOC105282864 0.038322843 uncharacterized LOC105282864 527 LOC105282376 0.039194352 RNA-binding protein 7 528 LOC105281784 0.039200302 transcription termination factor 2 529 LOC105284141 0.039200302 speckle-type POZ protein B 530 LOC105285823 0.039606123 elongation of very long chain fatty acids protein AAEL008004 531 LOC105284850 0.039943123 AP-3 complex subunit sigma-1 532 LOC105276858 0.040006352 transcription factor SPT20 homolog 533 LOC105279064 0.040006352 uncharacterized LOC105279064 534 LOC105285186 0.040140578 structural maintenance of chromosomes protein 6 535 LOC105287357 0.040200326 uncharacterized LOC105287357 536 LOC105274498 0.040851277 sodium- and chloride-dependent GABA transporter 1 537 LOC105276159 0.040851277 uncharacterized protein MAL13P1.304 538 LOC105287783 0.040851277 sodium channel protein Nach 539 LOC105275608 0.041004647 uncharacterized LOC105275608 540 LOC105276234 0.041004647 phospholipid phosphatase 6 541 LOC105279933 0.041004647 leucine-rich repeat-containing G-protein coupled receptor 4 542 LOC105285453 0.041434562 kinectin 543 LOC105280382 0.042129344 major facilitator superfamily domain-containing protein 6 544 LOC105283491 0.042246189 endoribonuclease Dcr-1 545 LOC105281288 0.042424317 RNA-binding protein squid 546 LOC105282239 0.042484526 uncharacterized LOC105282239 547 LOC105281920 0.042763701 548 LOC105287196 0.042763701 leucine-rich repeat protein 1 549 LOC105277627 0.042854837 nucleolar protein 14 homolog 550 LOC105281729 0.042921014 uncharacterized LOC105281729 551 LOC105277339 0.043338805 vicilin-like seed storage protein At2g18540 552 LOC105277777 0.043338805 pseudouridylate synthase 7 homolog 553 LOC105279952 0.043338805 uncharacterized LOC105279952 554 LOC105274595 0.043528611 chromobox protein homolog 3 555 LOC105278868 0.043615876 uncharacterized LOC105278868 556 LOC105279367 0.0436181 lipid storage droplets surface-binding protein 2 557 LOC105274633 0.043830814 ejaculatory bulb-specific protein 3 558 LOC105279164 0.043830814 uncharacterized LOC105279164 559 LOC105285169 0.043830814 sperm flagellar protein 2 560 LOC105287016 0.043833302 adenylate kinase isoenzyme 5 561 LOC105278924 0.044449161 uncharacterized LOC105278924 562 LOC105275563 0.044542688 uncharacterized LOC105275563 563 LOC105281601 0.044542688 eukaryotic translation initiation factor 4E-1A 564 LOC105287573 0.044542688 E3 ubiquitin-protein ligase RAD18 565 LOC105279259 0.044734305 ATP synthase subunit e 566 LOC105280130 0.044903873 uncharacterized LOC105280130 567 LOC105287080 0.044903873 uncharacterized LOC105287080 568 LOC105278713 0.044938377 LIM domain kinase 1 569 LOC105287888 0.045429367 uncharacterized LOC105287888 570 LOC105275551 0.045614788 uncharacterized LOC105275551 571 LOC105275867 0.045614788 uncharacterized LOC105275867 572 LOC105276573 0.045614788 leucine-rich repeat neuronal protein 2 573 LOC105278750 0.045614788 cell adhesion molecule 2 574 LOC105280293 0.045614788 pro-corazonin 575 LOC105284209 0.045614788 lysophospholipid acyltransferase 7 576 LOC105287733 0.045614788 DNA demethylase ALKBH1 577 LOC105287809 0.045614788 uncharacterized LOC105287809 578 LOC105277277 0.045746641 cytochrome P450 9e2 579 LOC105274835 0.045759955 uncharacterized LOC105274835 580 LOC105281338 0.045759955 zinc finger protein 800 581 LOC105288116 0.045759955 uncharacterized LOC105288116 582 LOC105280261 0.04596819 uncharacterized LOC105280261 583 LOC105277096 0.045989542 lysosomal Pro-X carboxypeptidase 584 LOC105281290 0.046006084 late secretory pathway protein AVL9 homolog 585 LOC105288118 0.046887354 stress-associated endoplasmic reticulum protein 2 586 LOC105284513 0.047266765 putative E3 ubiquitin-protein ligase UBR7 587 LOC105282762 0.047379082 L-lactate dehydrogenase-like 588 LOC105282263 0.048000578 uncharacterized LOC105282263 589 LOC105276691 0.048240962 E3 ubiquitin-protein ligase RNF14 590 LOC105285279 0.048240962 tachykinins 591 LOC105284709 0.048566923 uncharacterized LOC105284709 592 LOC105276523 0.048582277 acetyl-coenzyme A synthetase 593 LOC105287556 0.048940312 longitudinals lacking protein 594 LOC105280565 0.049387746 DNA-directed RNA polymerases I and III subunit RPAC1 595 LOC105281982 0.04942483 cytochrome c oxidase assembly factor 3 596 LOC105287317 0.049864337 actin 71 Supplementary Table 2

72 All gene clusters identified, along with their corresponding enrichment for differentially 73 expressed genes, gene ontology terms, and transcription factor binding sites.

74

Column heading Description cluster Cluster id. This is the id that is referenced throughout the paper. change_int The interval in which the greatest change is observed. 0: 0-12 hrs, 1: 12-24hrs, 2: 24-48 hrs, 3: 48-96 hrs. change_mag The direction and magnitude of change in gene expression at the interval indicated above. num_genes the number of genes assigned to the cluster num_degs the number of significantly differentially expressed genes (DEGs) assigned to the cluster % DEGs the percentage of DEGs in the cluster odds_rat the odds ratio from a fisher's exact test, examining enrichment of the cluster for DEGs p_val the p-value calculated from fisher's exact test for cluster enrichment of DEGs q_val the benjamini-hochberg FDR corrected p-value for DEG enrichment (yellow if <0.05) # GO terms the number of significantly enriched gene ontology terms (green if >0) # TFBS the number of significantly enriched transcription factor binding sites (green if >0) GO Term GO terms ID, class and description TFBS TFBS name (Drosophila melanogaster ortholog) and the FDR q-value. Transition from brood care to reproduction Cluster Cluster Enrichment for information on Differentially Expressed pattern of Genes expression num_ change_i change_ num_ % odds # GO Transcription Factor Binding cluster gene p_val q_val # TFBS GO Term ID, type and definition nt mag degs DEGs _rat terms Site and q-value s

OdsH(0.0099) exex(0.0099) CG11294(0.0144) al(0.0153) Dr(0.0157) ro(0.0157) GO:0020037 [MF] 'heme binding' Rx(0.0157) en(0.0173) GO:0016705 [MF] 'oxidoreductase Vsx2(0.0174) Pph13(0.0186) activity, acting on paired donors, with otp(0.0189) CG32532(0.0194) 35 0 -0.57 167 19 11.4% 2.87 0.0001 0.0016 1 1 incorporation or reduction of molecular unpg(0.0194) inv(0.0217) oxygen' GO:0005506 [MF] 'iron ion slou(0.0224) Vsx1(0.0228) binding' repo(0.0237) CG9876(0.0246) ind(0.0258) unc-4(0.0278) Lim1(0.0294) hbn(0.0334) ftz(0.0461) Awh(0.0473) GO:0043169 [MF] 'cation binding' GO:0004252 [MF] 'serine-type endopeptidase activity' GO:0052689 [MF] 'carboxylic ester hydrolase activity' GO:0016787 [MF] 'hydrolase 69 1 -0.52 55 8 14.5% 3.76 0.0025 0.0239 1 1 activity' GO:0003824 [MF] 'catalytic activity' GO:0006508 [BP] 'proteolysis' GO:0007160 [BP] 'cell-matrix adhesion' GO:0009166 [BP] 'nucleotide catabolic process' btn(0.0103) CG18599(0.0108) CG9876(0.0148) lbl(0.0148) gt(0.0152) inv(0.0160) pb(0.0160) Rx(0.0163) CG32105(0.0172) fkh(0.0172) repo(0.0173) PHDP(0.0177) zen(0.0192) zen2(0.0194) GO:0008061 [MF] 'chitin binding' HGTX(0.0194) br(0.0194) GO:0005319 [MF] 'lipid transporter H2.0(0.0202) Vsx2(0.0217) activity' GO:0016705 [MF] ro(0.0223) exex(0.0226) 'oxidoreductase activity, acting on lms(0.0233) onecut(0.0237) paired donors, with incorporation or slou(0.0251) slp1(0.0251) 4 3 1.16 156 31 19.9% 5.67 0 0 1 1 reduction of molecular oxygen' otp(0.0251) Vsx1(0.0258) GO:0005506 [MF] 'iron ion binding' Scr(0.0267) hbn(0.0272) GO:0005576 [CL] 'extracellular region' ap(0.0272) eve(0.0272) GO:0006030 [BP] 'chitin metabolic NK7.1(0.0277) en(0.0278) process' GO:0006869 [BP] 'lipid Pph13(0.0278) Six4(0.0279) transport' Lim1(0.0302) CG11617(0.0308) CG32532(0.0326) unpg(0.0348) C15(0.0359) CG4328-RA(0.0380) Lim3(0.0393) Dbx(0.0411) ems(0.0416) lab(0.0430) Dll(0.0458) Awh(0.0494)

19 3 0.59 245 58 23.7% 7.41 0 0 1 1 fkh(0.0461) opa(0.0000) Trl(0.0001) Mad(0.0002) btd(0.0003) GO:0005216 [MF] 'ion channel activity' usp(0.0013) h(0.0053) 21 3 -0.28 315 25 7.9% 1.93 0.003 0.025 1 1 GO:0006811 [BP] 'ion transport' gcm2(0.0096) dl(var.2)(0.0096) brk(0.0148) Ets21C(0.0402) 6 3 -0.43 231 28 12.1% 3.12 0 0 1 1 7 3 -0.31 35 7 20.0% 5.52 0.0007 0.0074 1 1 36 3 0.28 73 13 17.8% 4.83 0 0.0002 1 1 44 3 0.27 353 41 11.6% 3.01 0 0 1 1 Mad(0.0073) hkb(0.0173) 3 0 0.19 341 5 1.5% 0.32 0.9993 1 1 1 brk(0.0223) Eip74EF(0.0415) zen(0.0105) pb(0.0117) Dfd(0.0194) lab(0.0219) GO:0004984 [MF] 'olfactory receptor CG18599(0.0229) activity' GO:0005549 [MF] 'odorant 12 0 -0.08 319 11 3.4% 0.78 0.8292 1 1 1 Scr(0.0237) vvl(0.0278) binding' GO:0007608 [BP] 'sensory Antp(0.0278) E5(0.0348) perception of smell' lbe(0.0367) btn(0.0461) Lim3(0.0461) zen2(0.0475) odd(0.0162) Gsc(0.0382) 17 0 0.04 481 13 2.7% 0.6 0.9796 1 1 1 GO:0015074 [BP] 'DNA integration' oc(0.0434) h(0.0000) Mad(0.0000) brk(0.0003) opa(0.0068) 26 0 0.55 194 6 3.1% 0.7 0.8564 1 1 1 CTCF(0.0190) btd(0.0194) hkb(0.0308) usp(0.0421) lab(0.0118) C15(0.0157) cad(0.0173) br(var.3)(0.0212) Dbx(0.0237) Cf2(0.0277) GO:0006366 [BP] 'transcription from 27 0 -0.14 202 5 2.5% 0.55 0.9441 1 1 1 exex(0.0278) Ubx(0.0279) RNA polymerase II ' Abd-B(0.0302) abd-A(0.0303) br(0.0341) hb(0.0355) vnd(0.0392) Pph13(0.0475) opa(0.0000) Mad(0.0000) usp(0.0000) Trl(0.0001) 29 0 0.49 83 8 9.6% 2.35 0.0283 0.1433 1 1 brk(0.0002) hkb(0.0003) h(0.0015) btd(0.0042) Mad(0.0002) opa(0.0017) 42 0 0.64 117 0 0.0% 0 1 1 1 1 brk(0.0108) usp(0.0411) br(var.4)(0.0003) Abd- B(0.0004) cad(0.0031) C15(0.0039) CG4328- RA(0.0041) br(var.3)(0.0047) Dr(0.0138) Dll(0.0151) vnd(0.0153) Pph13(0.0157) fkh(0.0157) H2.0(0.0165) br(0.0173) Ubx(0.0194) CG15696-RA(0.0228) GO:0004518 [MF] 'nuclease activity' lbe(0.0237) CG9876(0.0237) 55 0 -0.24 343 25 7.3% 1.75 0.0087 0.059 1 1 GO:0008152 [BP] 'metabolic process' PHDP(0.0238) D(0.0244) Dbx(0.0250) exd(0.0271) NK7.1(0.0299) tup(0.0303) hbn(0.0303) br(var.2)(0.0328) lms(0.0340) Hmx(0.0359) nub(0.0382) E5(0.0387) Lim3(0.0399) Cf2(0.0412) CG32105(0.0414) Rx(0.0416) B-H2(0.0461) abd-A(0.0461) ap(0.0461) hb(0.0480)

slbo(0.0092) Abd-B(0.0157) 58 0 -0.1 340 7 2.1% 0.45 0.9932 1 1 1 H2.0(0.0190) CG4328- RA(0.0206) Dbx(0.0207) eve(0.0041) CG18599(0.0054) vvl(0.0064) lab(0.0064) Scr(0.0064) pb(0.0064) E5(0.0096) HGTX(0.0096) ap(0.0096) Antp(0.0108) zen(0.0116) slou(0.0138) btn(0.0148) fkh(0.0152) vnd(0.0157) H2.0(0.0171) lms(0.0174) ftz(0.0189) ct(0.0189) ind(0.0194) br(0.0194) abd- 60 0 -0.33 279 9 3.2% 0.73 0.8654 1 1 1 A(0.0194) br(var.3)(0.0200) ems(0.0206) Vsx1(0.0207) Abd-B(0.0211) NK7.1(0.0212) br(var.2)(0.0237) lbl(0.0272) C15(0.0279) Awh(0.0299) zen2(0.0340) Dll(0.0380) Pph13(0.0382) nub(0.0402) inv(0.0405) Dfd(0.0415) lbe(0.0415) CG34031(0.0461) otp(0.0461) PHDP(0.0493) Rx(0.0493) Cf2(0.0497)

Mad(0.0001) h(0.0001) 61 0 0.37 290 9 3.1% 0.7 0.8921 1 1 1 usp(0.0010) opa(0.0015) btd(0.0048) brk(0.0151) opa(0.0000) Mad(0.0000) h(0.0001) brk(0.0003) 72 0 0.36 215 5 2.3% 0.52 0.9612 1 1 1 CTCF(0.0157) hkb(0.0172) usp(0.0431) GO:0004812 [MF] 'aminoacyl-tRNA 8 0 -0.26 141 3 2.1% 0.47 0.9493 1 1 1 ligase activity' GO:0006418 [BP] 'tRNA aminoacylation for protein translation' 9 0 -0.18 56 0 0.0% 0 1 1 1 1 24 0 -0.21 283 4 1.4% 0.31 0.9986 1 1 1 34 0 -0.24 135 8 5.9% 1.38 0.238 0.8615 1 1 46 0 -0.25 159 8 5.0% 1.16 0.3932 1 1 1 48 0 0.11 148 2 1.4% 0.3 0.9898 1 1 1 51 0 -0.12 115 1 0.9% 0.19 0.9943 1 1 1 52 0 -0.06 533 6 1.1% 0.24 1 1 1 1 GO:0008236 [MF] 'serine-type peptidase activity' GO:0008138 [MF] 'protein tyrosine/serine/threonine phosphatase activity' GO:0003735 56 0 0.35 64 0 0.0% 0 1 1 1 1 [MF] 'structural constituent of ribosome' GO:0005840 [CL] 'ribosome' GO:0005622 [CL] 'intracellular' GO:0006412 [BP] 'translation' GO:0003735 [MF] 'structural constituent of ribosome' GO:0005840 59 0 0.2 75 2 2.7% 0.6 0.8456 1 1 1 [CL] 'ribosome' GO:0005622 [CL] 'intracellular' GO:0006412 [BP] 'translation' GO:0004190 [MF] 'aspartic-type endopeptidase activity' GO:0016805 [MF] 'dipeptidase activity' GO:0004222 [MF] 'metalloendopeptidase activity' 67 0 0 378 0 0.0% 0 1 1 1 1 GO:0003676 [MF] 'nucleic acid binding' GO:0031012 [CL] 'extracellular matrix' GO:0015074 [BP] 'DNA integration' GO:0006508 [BP] 'proteolysis' 68 0 0.33 56 0 0.0% 0 1 1 1 1 73 0 0.19 173 1 0.6% 0.13 0.9996 1 1 1 GO:0017048 [MF] 'Rho GTPase binding' GO:0043565 [MF] 'sequence- specific DNA binding' GO:0030036 [BP] 'actin cytoskeleton organization' GO:0006355 [BP] 'regulation of 74 0 0.27 67 0 0.0% 0 1 1 1 1 transcription, DNA-templated' GO:0006812 [BP] 'cation transport' GO:0035556 [BP] 'intracellular signal transduction' GO:0016043 [BP] 'cellular component organization' 75 0 0.13 131 2 1.5% 0.34 0.9803 1 1 1 cnc::maf-S(0.0237) 11 1 0.12 261 2 0.8% 0.17 0.9999 1 1 1 GO:0005488 [MF] 'binding' brk(0.0475) GO:0004984 [MF] 'olfactory receptor activity' GO:0005549 [MF] 'odorant binding' GO:0003735 [MF] 'structural HHEX(0.0355) H2.0(0.0356) 18 1 -0.11 315 2 0.6% 0.14 1 1 1 1 constituent of ribosome' GO:0005840 CG4328-RA(0.0405) [CL] 'ribosome' GO:0007608 [BP] 'sensory perception of smell' GO:0006412 [BP] 'translation' GO:0005515 [MF] 'protein binding' GO:0005802 [CL] 'trans-Golgi network' kni(0.0157) brk(0.0233) 30 1 0.15 282 13 4.6% 1.06 0.462 1 1 1 GO:0009113 [BP] 'purine nucleobase CTCF(0.0237) h(0.0418) biosynthetic process' GO:0010508 [BP] 'positive regulation of autophagy' GO:0003735 [MF] 'structural constituent of ribosome' GO:0005840 32 1 -0.24 64 1 1.6% 0.35 0.9431 1 1 1 [CL] 'ribosome' GO:0005622 [CL] fkh(0.0151) 'intracellular' GO:0006412 [BP] 'translation' CTCF(0.0026) h(0.0116) 41 1 0.08 308 8 2.6% 0.58 0.9623 1 1 1 brk(0.0461) GO:0005337 [MF] 'nucleoside 66 1 0.23 107 1 0.9% 0.21 0.9918 1 1 1 Gsc(0.0096) oc(0.0217) transmembrane transporter activity' 16 1 -0.21 109 1 0.9% 0.2 0.9925 1 1 1 GO:0004984 [MF] 'olfactory receptor activity' GO:0005549 [MF] 'odorant 25 1 -0.19 43 1 2.3% 0.52 0.854 1 1 1 binding' GO:0016020 [CL] 'membrane' GO:0007608 [BP] 'sensory perception of smell' GO:0004984 [MF] 'olfactory receptor activity' GO:0005549 [MF] 'odorant 33 1 -0.14 172 3 1.7% 0.39 0.9823 1 1 1 binding' GO:0016020 [CL] 'membrane' GO:0007608 [BP] 'sensory perception of smell' 38 1 0.13 189 2 1.1% 0.23 0.998 1 1 1 gt(0.0303) CG9876(0.0303) Lim1(0.0338) exex(0.0340) lms(0.0347) inv(0.0359) en(0.0393) OdsH(0.0461) 13 2 -0.25 39 1 2.6% 0.58 0.8253 1 1 1 Pph13(0.0461) Awh(0.0461) unpg(0.0461) CG11294(0.0461) Rx(0.0461) repo(0.0473) Vsx2(0.0479) 1 2 -0.14 64 0 0.0% 0 1 1 1 1 10 2 0.33 17 1 5.9% 1.37 0.5323 1 1 1 14 2 -0.11 87 0 0.0% 0 1 1 1 1 40 2 -0.18 40 1 2.5% 0.56 0.833 1 1 1 43 2 0.12 92 4 4.3% 0.99 0.5755 1 1 1 49 2 0.14 40 4 10.0% 2.44 0.0958 0.4282 1 1 53 2 0.12 93 1 1.1% 0.24 0.9845 1 1 1 20 3 -0.11 204 5 2.5% 0.55 0.9471 1 1 1 Eip74EF(0.0251) Mad(0.0000) opa(0.0000) btd(0.0000) brk(0.0000) 39 3 -0.19 391 14 3.6% 0.81 0.8143 1 1 1 gcm2(0.0021) h(0.0027) Trl(0.0056) usp(0.0092) hkb(0.0162) CTCF(0.0271) GO:0005249 [MF] 'voltage-gated potassium channel activity' GO:0015078 [MF] 'hydrogen ion transmembrane transporter activity' GO:0005216 [MF] 'ion channel activity' GO:0033179 [CL] 'proton-transporting Mad(0.0000) opa(0.0000) V-type ATPase, V0 domain' btd(0.0000) brk(0.0000) GO:0008076 [CL] 'voltage-gated 65 3 -0.35 204 17 8.3% 2.02 0.0082 0.059 1 1 Trl(0.0000) hkb(0.0000) potassium channel complex' usp(0.0004) h(0.0006) GO:0005891 [CL] 'voltage-gated CTCF(0.0192) calcium channel complex' GO:0055085 [BP] 'transmembrane transport' GO:0006813 [BP] 'potassium ion transport' GO:0006811 [BP] 'ion transport' GO:0051260 [BP] 'protein homooligomerization' GO:0001104 [MF] 'RNA polymerase II transcription cofactor activity' GO:0004812 [MF] 'aminoacyl-tRNA ligase activity' GO:0004817 [MF] 'cysteine-tRNA ligase activity' GO:0051082 [MF] 'unfolded protein binding' GO:0005524 [MF] 'ATP binding' GO:0004828 [MF] 'serine- tRNA ligase activity' GO:0031072 [MF] 'heat shock protein binding' 71 3 0.13 314 14 4.5% 1.02 0.5082 1 1 1 GO:0000166 [MF] 'nucleotide binding' cad(0.0334) hb(0.0461) GO:0016592 [CL] 'mediator complex' GO:0006281 [BP] 'DNA repair' GO:0009408 [BP] 'response to heat' GO:0006423 [BP] 'cysteinyl-tRNA aminoacylation' GO:0006418 [BP] 'tRNA aminoacylation for protein translation' GO:0006370 [BP] '7- methylguanosine mRNA capping' GO:0006434 [BP] 'seryl-tRNA aminoacylation' 2 3 0.11 178 6 3.4% 0.76 0.7955 1 1 1 5 3 -0.27 42 6 14.3% 3.67 0.0093 0.059 1 1 GO:0031177 [MF] 'phosphopantetheine binding' GO:0016740 [MF] 'transferase activity' GO:0016491 [MF] 'oxidoreductase activity' GO:0016705 [MF] 'oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen' 15 3 0.84 78 6 7.7% 1.83 0.1253 0.5014 1 1 GO:0020037 [MF] 'heme binding' GO:0016788 [MF] 'hydrolase activity, acting on ester bonds' GO:0005506 [MF] 'iron ion binding' GO:0055114 [BP] 'oxidation-reduction process' GO:0008152 [BP] 'metabolic process' GO:0009058 [BP] 'biosynthetic process' GO:0006629 [BP] 'lipid metabolic process' GO:0016884 [MF] 'carbon-nitrogen 22 3 0.37 72 4 5.6% 1.29 0.3862 1 1 1 ligase activity, with as amido- N-donor' 23 3 -0.12 314 7 2.2% 0.49 0.9857 1 1 1 28 3 -0.18 183 10 5.5% 1.27 0.2792 0.9644 1 1 31 3 0.39 141 12 8.5% 2.06 0.0207 0.1122 1 1 37 3 0.19 94 3 3.2% 0.72 0.7847 1 1 1 45 3 0.2 134 11 8.2% 1.98 0.0328 0.1558 1 1 47 3 0.17 29 0 0.0% 0 1 1 1 1 50 3 0.12 204 13 6.4% 1.5 0.1115 0.4708 1 1 GO:0004984 [MF] 'olfactory receptor activity' GO:0005549 [MF] 'odorant 54 3 -0.13 150 7 4.7% 1.07 0.4844 1 1 1 binding' GO:0007608 [BP] 'sensory perception of smell' 57 3 -0.12 354 9 2.5% 0.56 0.9747 1 1 1 62 3 0.26 99 7 7.1% 1.67 0.142 0.5396 1 1 GO:0004984 [MF] 'olfactory receptor activity' GO:0005549 [MF] 'odorant 63 3 -0.2 262 20 7.6% 1.84 0.0111 0.065 1 1 binding' GO:0016020 [CL] 'membrane' GO:0007608 [BP] 'sensory perception of smell' GO:0003735 [MF] 'structural constituent of ribosome' GO:0005840 [CL] 'ribosome' GO:0015935 [CL] 64 3 0.19 117 1 0.9% 0.19 0.9948 1 1 1 'small ribosomal subunit' GO:0005622 [CL] 'intracellular' GO:0042254 [BP] 'ribosome biogenesis' GO:0006412 [BP] 'translation' 70 3 -0.14 72 1 1.4% 0.31 0.9603 1 1 1 76 3 0.06 334 7 2.1% 0.46 0.9919 1 1 1 Transition from reproduction to brood care Cluster Cluster Enrichment for information on Differentially Expressed pattern of Genes expression num_ change_i change_ num_ % odds # GO # Transcription Factor Binding cluster gene p_val q_val GO Term ID, type and definition nt mag degs DEGs _rat terms TFBS Site and q-value s Mad(0.0000) h(0.0001) 93 0 0.25 134 16 11.9% 3.02 0.0002 0.0019 0 6 opa(0.0015) btd(0.0019) brk(0.0025) usp(0.0076) GO:0005789 [CL] 'endoplasmic 56 0 0.21 91 22 24.2% 7.21 0 0 1 0 reticulum membrane' GO:0005576 [CL] 'extracellular region' 65 0 -1.21 76 17 22.4% 6.46 0 0 3 0 GO:0005615 [CL] 'extracellular space' GO:0035176 [BP] 'social behavior' 34 0 0.21 119 17 14.3% 3.73 0 0.0001 0 0 12 0 0.48 82 13 15.9% 4.19 0.0001 0.0004 0 0 8 1 -0.35 134 48 35.8% 13.2 0 0 0 1 fkh(0.0252) GO:0008237 [MF] 'metallopeptidase 47 1 -0.18 206 59 28.6% 9.64 0 0 1 0 activity' 96 1 -0.26 109 17 15.6% 4.13 0 0 0 0 GO:0046933 [MF] 'proton-transporting ATP synthase activity, rotational mechanism' GO:0005834 [CL] 'heterotrimeric G-protein complex' opa(0.0000) btd(0.0001) 90 2 0.16 151 30 19.9% 5.66 0 0 5 6 GO:0046034 [BP] 'ATP metabolic h(0.0026) Mad(0.0029) process' GO:0015986 [BP] 'ATP usp(0.0117) hkb(0.0450) synthesis coupled proton transport' GO:0035556 [BP] 'intracellular signal transduction' GO:0055114 [BP] 'oxidation-reduction Mad(0.0002) usp(0.0345) 70 2 0.22 162 45 27.8% 9.02 0 0 1 4 process' opa(0.0398) Trl(0.0475) GO:0043565 [MF] 'sequence-specific 58 3 0.25 126 15 11.9% 3.01 0.0004 0.0029 1 0 DNA binding' GO:0052689 [MF] 'carboxylic ester 82 3 0.4 133 14 10.5% 2.61 0.002 0.0137 1 0 hydrolase activity' 73 3 0.66 72 23 31.9% 10.7 0 0 0 0 91 3 -0.23 169 41 24.3% 7.45 0 0 0 0 opa(0.0013) Mad(0.0073) btd(0.0117) gcm2(0.0162) 15 0 0.17 187 4 2.1% 0.47 0.966 1 1 8 GO:0005856 [CL] 'cytoskeleton' Trl(0.0162) usp(0.0209) brk(0.0450) schlank(0.0496) GO:0004889 [MF] 'acetylcholine- activated cation-selective channel activity' GO:0004129 [MF] 'cytochrome- 87 0 0.26 180 5 2.8% 0.62 0.8991 1 4 2 c oxidase activity' GO:0045211 [CL] h(0.0073) Mad(0.0142) 'postsynaptic membrane' GO:0043044 [BP] 'ATP-dependent chromatin remodeling' GO:0004190 [MF] 'aspartic-type endopeptidase activity' GO:0003887 [MF] 'DNA-directed DNA polymerase activity' GO:0016805 [MF] 'dipeptidase activity' GO:0008408 [MF] '3'-5' exonuclease activity' GO:0008237 [MF] 'metallopeptidase activity' 5 0 0 385 0 0.0% 0 1 1 10 1 usp(0.0373) GO:0004222 [MF] 'metalloendopeptidase activity' GO:0003676 [MF] 'nucleic acid binding' GO:0031012 [CL] 'extracellular matrix' GO:0015074 [BP] 'DNA integration' GO:0006508 [BP] 'proteolysis' h(0.0252) vis(0.0252) 30 0 0.22 545 2 0.4% 0.08 1 1 0 3 brk(0.0252) 25 0 0.08 235 1 0.4% 0.09 1 1 0 1 kni(0.0252) 9 0 -0.29 78 0 0.0% 0 1 1 0 1 nub(0.0249) GO:0004984 [MF] 'olfactory receptor activity' GO:0003743 [MF] 'translation initiation factor activity' GO:0005549 [MF] 'odorant binding' GO:0004803 [MF] 'transposase activity' 68 0 -0.22 206 0 0.0% 0 1 1 7 0 GO:0007608 [BP] 'sensory perception of smell' GO:0006313 [BP] 'transposition, DNA-mediated' GO:0050909 [BP] 'sensory perception of taste' GO:0008479 [MF] 'queuine tRNA- ribosyltransferase activity' GO:0046873 [MF] 'metal ion 33 0 0.12 255 1 0.4% 0.08 1 1 3 0 transmembrane transporter activity' GO:0008616 [BP] 'queuosine biosynthetic process' GO:0004984 [MF] 'olfactory receptor activity' GO:0005549 [MF] 'odorant 42 0 -0.36 20 0 0.0% 0 1 1 3 0 binding' GO:0007608 [BP] 'sensory perception of smell' GO:0005328 [MF] 'neurotransmitter:sodium symporter activity' GO:0016021 [CL] 'integral 89 0 0.34 51 3 5.9% 1.37 0.3863 1 3 0 component of membrane' GO:0006836 [BP] 'neurotransmitter transport' GO:0043169 [MF] 'cation binding' GO:0004252 [MF] 'serine-type endopeptidase activity' GO:0004132 [MF] 'dCMP deaminase activity' 85 0 -1.16 132 12 9.1% 2.21 0.0129 0.0814 7 0 GO:0016787 [MF] 'hydrolase activity' GO:0003824 [MF] 'catalytic activity' GO:0006220 [BP] 'pyrimidine nucleotide metabolic process' GO:0006508 [BP] 'proteolysis' GO:0016740 [MF] 'transferase activity' GO:0016788 [MF] 'hydrolase activity, acting on ester bonds' GO:0016491 [MF] 'oxidoreductase activity' 37 0 -0.53 119 3 2.5% 0.56 0.8975 1 6 0 GO:0055114 [BP] 'oxidation-reduction process' GO:0008152 [BP] 'metabolic process' GO:0019441 [BP] 'tryptophan catabolic process to kynurenine' GO:0003735 [MF] 'structural constituent of ribosome' GO:0005840 [CL] 'ribosome' GO:0015935 [CL] 26 0 -0.23 157 4 2.5% 0.57 0.9167 1 5 0 'small ribosomal subunit' GO:0042254 [BP] 'ribosome biogenesis' GO:0006412 [BP] 'translation' 61 0 -0.52 29 2 6.9% 1.62 0.3637 1 1 0 GO:0005215 [MF] 'transporter activity' GO:0006850 [BP] 'mitochondrial 80 0 -0.19 95 0 0.0% 0 1 1 1 0 pyruvate transport' 1 0 0.22 66 1 1.5% 0.34 0.948 1 0 0 3 0 -0.39 132 0 0.0% 0 1 1 0 0 4 0 0.25 214 2 0.9% 0.2 0.9993 1 0 0 17 0 0.17 286 2 0.7% 0.15 1 1 0 0 40 0 0.44 82 0 0.0% 0 1 1 0 0 41 0 -0.15 192 3 1.6% 0.34 0.9913 1 0 0 48 0 0.45 214 2 0.9% 0.2 0.9993 1 0 0 51 0 0.16 143 0 0.0% 0 1 1 0 0 52 0 -0.18 246 3 1.2% 0.27 0.9988 1 0 0 63 0 -0.24 113 6 5.3% 1.23 0.3727 1 0 0 74 0 -0.19 149 3 2.0% 0.45 0.9611 1 0 0 78 0 -0.59 183 10 5.5% 1.27 0.2792 1 0 0 84 0 -0.29 159 0 0.0% 0 1 1 0 0 88 0 -0.32 309 10 3.2% 0.73 0.8733 1 0 0 95 0 -0.09 662 3 0.5% 0.1 1 1 0 0 32 0 -0.1 309 13 4.2% 0.96 0.5962 1 0 0 21 0 -0.16 130 0 0.0% 0 1 1 0 0 62 0 -0.33 33 0 0.0% 0 1 1 0 0 GO:0008146 [MF] 'sulfotransferase 19 1 0.25 47 0 0.0% 0 1 1 2 0 activity' GO:0005783 [CL] 'endoplasmic reticulum' GO:0004984 [MF] 'olfactory receptor activity' GO:0005549 [MF] 'odorant 92 1 -0.18 56 1 1.8% 0.4 0.9185 1 4 0 binding' GO:0016020 [CL] 'membrane' GO:0007608 [BP] 'sensory perception of smell' GO:0003995 [MF] 'acyl-CoA dehydrogenase activity' GO:0016627 [MF] 'oxidoreductase activity, acting on the CH-CH group of donors' GO:0050660 [MF] 'flavin adenine 2 1 -0.25 27 0 0.0% 0 1 1 7 0 dinucleotide binding' GO:0006635 [BP] 'fatty acid beta-oxidation' GO:0006631 [BP] 'fatty acid metabolic process' GO:0055114 [BP] 'oxidation-reduction process' GO:0008152 [BP] 'metabolic process' 53 1 -0.19 17 1 5.9% 1.37 0.5323 1 1 0 GO:0003824 [MF] 'catalytic activity' 64 1 -0.11 159 8 5.0% 1.16 0.3932 1 0 0 86 1 -0.33 49 5 10.2% 2.5 0.0616 0.3254 0 0 13 1 0.3 4 0 0.0% 0 1 1 0 0 24 1 -0.18 73 0 0.0% 0 1 1 0 0 29 1 -0.17 130 3 2.3% 0.51 0.9275 1 0 0 83 1 -0.18 46 0 0.0% 0 1 1 0 0 GO:0004553 [MF] 'hydrolase activity, hydrolyzing O-glycosyl compounds' 69 2 0.36 20 1 5.0% 1.15 0.5911 1 2 0 GO:0005975 [BP] 'carbohydrate metabolic process' GO:0019752 [BP] 'carboxylic acid 20 2 0.31 34 1 2.9% 0.66 0.7815 1 1 0 metabolic process' 35 2 0.14 179 1 0.6% 0.12 0.9997 1 1 0 GO:0000166 [MF] 'nucleotide binding' GO:0003777 [MF] 'microtubule motor activity' GO:0005198 [MF] 'structural molecule activity' GO:0006511 [BP] 44 2 0.24 80 2 2.5% 0.56 0.8702 1 4 0 'ubiquitin-dependent protein catabolic process' GO:0007018 [BP] 'microtubule-based movement' GO:0015991 [BP] 'ATP hydrolysis coupled proton transport' GO:0033180 16 2 0.1 209 7 3.3% 0.76 0.8132 1 2 0 [CL] 'proton-transporting V-type ATPase, V1 domain' GO:0016972 [MF] 'thiol oxidase 18 2 -0.09 140 7 5.0% 1.15 0.413 1 1 0 activity' GO:0008061 [MF] 'chitin binding' 23 2 0.46 25 0 0.0% 0 1 1 2 0 GO:0006030 [BP] 'chitin metabolic process' 46 2 0.12 112 8 7.1% 1.69 0.1169 0.5553 0 0 28 2 0.1 168 9 5.4% 1.24 0.3136 1 0 0 31 2 0.17 113 0 0.0% 0 1 1 0 0 36 2 0.07 231 2 0.9% 0.19 0.9996 1 0 0 59 2 -0.17 33 2 6.1% 1.41 0.4261 1 0 0 22 2 0.23 36 0 0.0% 0 1 1 0 0 27 2 -0.08 117 1 0.9% 0.19 0.9948 1 0 0 43 2 -0.21 21 0 0.0% 0 1 1 0 0 49 2 -0.13 120 0 0.0% 0 1 1 0 0 60 2 0.4 16 0 0.0% 0 1 1 0 0 GO:0016641 [MF] 'oxidoreductase activity, acting on the CH-NH2 group of donors, oxygen as acceptor' GO:0004930 [MF] 'G-protein coupled receptor activity' GO:0020037 [MF] 'heme binding' GO:0000062 [MF] 'fatty- acyl-CoA binding' GO:0003824 [MF] 'catalytic activity' GO:0016705 [MF] 77 3 0.36 65 0 0.0% 0 1 1 10 1 . 'oxidoreductase activity, acting on paired donors, with incorporation or reduction of molecular oxygen' GO:0005506 [MF] 'iron ion binding' GO:0055114 [BP] 'oxidation-reduction process' GO:0008152 [BP] 'metabolic process' GO:0008299 [BP] 'isoprenoid biosynthetic process' opa(0.0064) btd(0.0099) 10 3 -0.2 84 8 9.5% 2.32 0.0301 0.1788 0 3 Mad(0.0290) GO:0004984 [MF] 'olfactory receptor activity' GO:0005549 [MF] 'odorant 50 3 0.13 129 0 0.0% 0 1 1 4 0 binding' GO:0016020 [CL] 'membrane' GO:0007608 [BP] 'sensory perception of smell' GO:0004984 [MF] 'olfactory receptor activity' GO:0005549 [MF] 'odorant 45 3 0.09 145 1 0.7% 0.15 0.9985 1 3 0 binding' GO:0007608 [BP] 'sensory perception of smell' GO:0020037 [MF] 'heme binding' GO:0031177 [MF] 'phosphopantetheine binding' GO:0016491 [MF] 'oxidoreductase activity' GO:0016705 [MF] 'oxidoreductase activity, acting on 94 3 0.82 135 10 7.4% 1.76 0.0716 0.3581 8 0 paired donors, with incorporation or reduction of molecular oxygen' GO:0005215 [MF] 'transporter activity' GO:0005506 [MF] 'iron ion binding' GO:0055114 [BP] 'oxidation-reduction process' GO:0008152 [BP] 'metabolic process' GO:0005248 [MF] 'voltage-gated sodium channel activity' GO:0001518 66 3 -0.08 173 2 1.2% 0.25 0.9962 1 3 0 [CL] 'voltage-gated sodium channel complex' GO:0016012 [CL] 'sarcoglycan complex' 11 3 0.29 43 0 0.0% 0 1 1 0 0 14 3 -0.08 175 2 1.1% 0.25 0.9965 1 0 0 39 3 0.06 291 0 0.0% 0 1 1 0 0 54 3 0.22 128 5 3.9% 0.89 0.6633 1 0 0 57 3 0.2 140 2 1.4% 0.31 0.9861 1 0 0 67 3 0.14 282 1 0.4% 0.08 1 1 0 0 72 3 -0.03 563 0 0.0% 0 1 1 0 0 79 3 -0.13 269 11 4.1% 0.93 0.6326 1 0 0 38 3 0.22 56 1 1.8% 0.4 0.9185 1 0 0 55 3 -0.22 204 15 7.4% 1.76 0.0342 0.191 0 0 6 3 -0.47 15 1 6.7% 1.56 0.4886 1 0 0 7 3 -0.17 118 1 0.8% 0.19 0.995 1 0 0 71 3 -0.12 185 3 1.6% 0.36 0.9888 1 0 0 76 3 0.25 47 1 2.1% 0.47 0.878 1 0 0 81 3 -0.27 71 1 1.4% 0.31 0.9584 1 0 0 75 Supplementary Table 3

76 Basic network statistics of gene clusters for time course gene expression between both 77 reproductive and brood care phase transitions.

Network parameters Complete network Conserved network*

Number of nodes 172 143 Connected components 2 11 Network Diameter 4 10 Network Centralization 0.203 0.073 Average number of neighbors 40.744 1.846 Network density 0.238 0.013 Network heterogeneity 0.389 0.863 Isolated nodes 1 0 78 Each gene was assigned to a cluster based on its expression profile. The complete network was 79 constructed using the clusters from both phase transitions. The network edges represent the 80 number of genes shared between two clusters from different transitions. No cluster will share 81 genes with any other cluster from the same transition, and each gene is represented on one and 82 only one edge.

83 * The conserved network is a subset of the complete network, containing all clusters that share 84 more genes than expected by chance (Jaccard Index, 1000 permutations, q-value ≤ 0.05).

85 86

87 Supplementary Table 4

88 Top 40 DEGs (ranked according to log2 fold change in expression for control vs. 12-hour time 89 point and control vs. 96-hour time point for each transition).

90

Top 40 DEGs (highest log2 fold change in expression) Reproduction to brood care Brood care to reproduction 0-96 hours 0-12 hours 0-96 hours 0-12 hours Gene Annotation Gene Annotation Gene Annotation Gene Annotation 1 LOC105286381 uncharacterized LOC105286381 LOC105285921 uncharacterized LOC105285921 LOC105281428_Q queen Vg LOC105287093 uncharacterized LOC105287093 2 LOC105285921 uncharacterized LOC105285921 LOC105286381 uncharacterized LOC105286381 LOC105277525 uncharacterized LOC105277525 LOC105276678 transferrin 3 LOC105285920 vegetative cell wall protein gp1 LOC105276678 transferrin LOC105279933 leucine-rich repeat-containing G-protein coupled receptor 4 LOC105278524 LIRP - ILP2 4 LOC105275344 csp8 LOC105285823 elongation of very long chain fatty acids protein LOC105277435 targeting protein for Xklp2 LOC105278931 venom allergen 3 5 LOC105276678 transferrin LOC105278724 fatty acid synthase LOC105287193 uncharacterized LOC105287193 LOC105277716 facilitated trehalose transporter Tret1 6 LOC105281428_Q queen Vg LOC105275344 csp8 LOC105281920 aminopeptidase N-like LOC105282359 myosin regulatory light chain 2 7 LOC105275345 ejaculatory bulb-specific protein 3 LOC105275345 ejaculatory bulb-specific protein 3 LOC105279314 trypsin inhibitor LOC105283737 myosin heavy chain, muscle 8 LOC105285823 elongation of very long chain fatty acids protein LOC105287552 snakeskin protein LOC105285920 vegetative cell wall protein gp1 LOC105281707 uncharacterized LOC105281707 9 LOC105277525 uncharacterized LOC105277525 LOC105277525 uncharacterized LOC105277525 LOC105284141 speckle-type POZ protein B LOC105287552 snakeskin protein 10 LOC105287552 snakeskin protein LOC105279933 leucine-rich repeat-containing G-protein coupled receptor 4 LOC105282036 G2/mitotic-specific cyclin-B LOC105278758 actin 11 LOC105276573 leucine-rich repeat neuronal protein 2 LOC105276573 leucine-rich repeat neuronal protein 2 LOC105287714 protein hunchback LOC105276573 leucine-rich repeat neuronal protein 2 12 LOC105278724 fatty acid synthase LOC105283737 myosin heavy chain, muscle LOC105281786 hyaluronan mediated motility receptor LOC105281721 uncharacterized LOC105281721 13 LOC105281012 uncharacterized LOC105281012 LOC105276433 box A-binding factor LOC105284119 ribonuclease H-like LOC105281786 hyaluronan mediated motility receptor 14 LOC105281708 uncharacterized LOC105281708 LOC105277700 actin, clone 205 LOC105281784 transcription termination factor 2 LOC105277700 actin, clone 205 15 LOC105288201 uncharacterized LOC105288201 LOC105277377 troponin C LOC105288201 uncharacterized LOC105288201 LOC105277377 troponin C 16 LOC105284141 speckle-type POZ protein B LOC105282359 myosin regulatory light chain 2 LOC105287010 uncharacterized LOC105287010 LOC105275507 PDZ and LIM domain protein 3 17 LOC105282036 G2/mitotic-specific cyclin-B LOC105287093 uncharacterized LOC105287093 LOC105283509 zinc finger BED domain-containing protein 1-like LOC105279933 leucine-rich repeat-containing G-protein coupled receptor 4 18 LOC105284181 peptidyl-prolyl cis-trans isomerase D LOC105287812 uncharacterized LOC105287812 LOC105279586 uncharacterized LOC105279586 LOC105277728 uncharacterized LOC105277728 19 LOC105287714 protein hunchback LOC105281721 uncharacterized LOC105281721 LOC105284633 gliomedin LOC105280772 uncharacterized LOC105280772 20 LOC105277435 targeting protein for Xklp2 LOC105285936 glutathione S-transferase 1-1 LOC105287013 kinesin-like protein KIF20B LOC105279907 uncharacterized LOC105279907 21 LOC105276433 box A-binding factor LOC105278758 actin LOC105283233 fibrillin-1 LOC105279559 glycine receptor subunit alpha-2-like 22 LOC105275236 carboxypeptidase M LOC105281449 muscle M-line assembly protein unc-89-like LOC105283266 lymphoid-specific helicase LOC105283796 ornithine decarboxylase 2-like 23 LOC105287193 uncharacterized LOC105287193 LOC105280202 glycine-rich cell wall structural protein LOC105278524 LIRP - ILP2 LOC105285920 vegetative cell wall protein gp1 24 LOC105281920 aminopeptidase N-like LOC105275821 dnaJ homolog subfamily C member 22 LOC105283796 ornithine decarboxylase 2-like LOC105276699 uncharacterized LOC105276699 25 LOC105283615 serine protease nudel LOC105287204 twitchin LOC105283366 uncharacterized LOC105283366 LOC105282680 glucosylceramidase 26 LOC105279559 glycine receptor subunit alpha-2-like LOC105276699 uncharacterized LOC105276699 LOC105277551 glycine N-methyltransferase LOC105281428_Q queen Vg 27 LOC105287812 uncharacterized LOC105287812 LOC105275435 alpha-(1,3)-fucosyltransferase 6 LOC105287207 kinesin-like protein Klp61F LOC105281103 pancreatic triacylglycerol lipase-like 28 LOC105281784 transcription termination factor 2 LOC105279012 uncharacterized LOC105279012 LOC105287679 kinetochore protein NDC80 homolog LOC105277939 uncharacterized LOC105277939 29 LOC105278931 venom allergen 3 LOC105278924 uncharacterized LOC105278924 LOC105274638 kinesin-like protein KIF18A LOC105275854 annulin 30 LOC105277551 glycine N-methyltransferase LOC105275769 uncharacterized LOC105275769 LOC105284779 uncharacterized LOC105284779 LOC105287317 actin, muscle-like 31 LOC105285597 uncharacterized LOC105285597 LOC105287317 actin, muscle-like LOC105284181 peptidyl-prolyl cis-trans isomerase D LOC105281449 muscle M-line assembly protein unc-89-like 32 LOC105283621 synaptic vesicle glycoprotein 2C LOC105277469 tubulin beta chain LOC105287093 uncharacterized LOC105287093 LOC105284373 myosin light chain alkali 33 LOC105279939 lymphokine-activated killer T-cell-originated protein kinase LOC105275469 sarcalumenin LOC105276433 box A-binding factor LOC105281784 transcription termination factor 2 34 LOC105283366 uncharacterized LOC105283366 LOC105287714 protein hunchback LOC105279939 lymphokine-activated killer T-cell-originated protein kinase LOC105281010 uncharacterized LOC105281010 35 LOC105281245 uncharacterized LOC105281245 LOC105284373 myosin light chain alkali LOC105278724 fatty acid synthase-like LOC105280202 glycine-rich cell wall structural protein 36 LOC105275821 dnaJ homolog subfamily C member 22 LOC105281448 twitchin LOC105287758 uncharacterized LOC105287758 LOC105285169 sperm flagellar protein 2 37 LOC105281786 hyaluronan mediated motility receptor LOC105284497 intraflagellar transport protein 46 homolog LOC105275039 uncharacterized LOC105275039 LOC105277551 glycine N-methyltransferase 38 LOC105283233 fibrillin-1 LOC105281212 protein rhomboid LOC105280537 uncharacterized LOC105280537 LOC105277439 ADP-ribosylation factor-like protein 6 39 LOC105277056 neuroparsin-A LOC105284074 phospholipase B1, membrane-associated LOC105279590 uncharacterized LOC105279590 LOC105275469 sarcalumenin 40 LOC105279064 uncharacterized LOC105279064 LOC105281920 aminopeptidase N-like LOC105281082 uncharacterized LOC105281082 LOC105287204 twitchin 91 Supplementary Table 5

92 Summary of clusters enriched for differentially expressed genes (DEGs) and transcription factor 93 binding sites (TFBSs). TFBS classes identified are as follows: Basic leucine zipper factors 94 (bZIP) – gt; C2H2 zinc finger factors – br, btd, hkb, opa, Trl, opa; Fork head / winged helix 95 factors – fkh, slp1; GCM domain factors – gcm2; Homeo domain factors – al, ap, Awh, btn, C15, 96 CG11294, CG11617, CG18599, CG32105, CG32532, CG4328-RA, CG9876, Dbx, Dll, Dr, ems, 97 en, eve, exex, ftz, H2.0, hbn, HGTX, ind, inv, lab, lbl, Lim1, Lim3, lms, NK7.1, OdsH, onecut, 98 otp, pb, PHDP, Pph13, repo, ro, Rx, Scr, Six4, slou, unc-4, unpg, Vsx1, Vsx2, zen, zen2; 99 Nuclear receptors with C4 zinc fingers – usp; Rel region (RHR) factors – dl; 100 SMAD/NF-1 DNA-binding domain factors – Mad; Zipper-type – h; Tryptophan cluster factors – 101 Ets21C.

102

Time interval with Maximum Number of Number of Cluster # maximal change genes in DEGs in the TFBSs in the cluster change in vector the cluster cluster expression Reproduction to brood care

8 12-24 hrs -0.35 134 48 fkh

93 0-12 hrs 0.25 134 16 Mad, usp, opa, btd, h, brk

90 24-48 hrs 0.16 151 30 Mad, usp,opa, btd, h, hkb

70 24-48 hrs 0.22 162 45 Mad, usp, opa, Trl

Brood care to reproduction

19 48-96 hrs 0.59 245 58 fkh

21 48-96 hrs -0.28 315 25 Mad, usp, opa, btd, Trl, h, brk, gcm2, dl, Ets21C

ap, Awh, br, btn, C15, CG11617, CG18599, CG32105, CG32532, CG4328-RA, CG9876, Dbx, Dll, ems, en, eve, exex, fkh, gt, H2.0, 4 48-96 hrs 1.16 156 31 hbn, HGTX, inv, lab, lbl, Lim1, Lim3, lms, NK7.1, onecut, otp, pb, PHDP, Pph13, repo, ro, Rx, Scr, Six4, slou, slp1, unpg, Vsx1, Vsx2, zen, zen2 al, Awh, CG11294, CG32532, CG9876, Dr, en, exex, ftz, hbn, ind, 35 0-12 hrs -0.57 167 19 inv, Lim1, OdsH, otp, Pph13, repo, ro, Rx, slou, unc-4, unpg, Vsx1, Vsx2 103 Maximum change vector: magnitude and direction of the largest average gene expression change between time intervals.

104