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Supporting Information Figure S1. The functionality of the tagged Arp6 and Swr1 was confirmed by monitoring cell growth and sensitivity to hydeoxyurea (HU). Five-fold serial dilutions of each strain were plated on YPD with or without 50 mM HU and incubated at 30°C or 37°C for 3 days. Figure S2. Localization of Arp6 and Swr1 on chromosome 3. The binding of Arp6-FLAG (top), Swr1-FLAG (middle), and Arp6-FLAG in swr1 cells (bottom) are compared. The position of Tel 3L, Tel 3R, CEN3, and the RP gene are shown under the panels. Figure S3. Localization of Arp6 and Swr1 on chromosome 4. The binding of Arp6-FLAG (top), Swr1-FLAG (middle), and Arp6-FLAG in swr1 cells (bottom) in the whole chromosome region are compared. The position of Tel 4L, Tel 4R, CEN4, SWR1, and RP genes are shown under the panels. Figure S4. Localization of Arp6 and Swr1 on the region including the SWR1 gene of chromosome 4. The binding of Arp6- FLAG (top), Swr1-FLAG (middle), and Arp6-FLAG in swr1 cells (bottom) are compared. The position and orientation of the SWR1 gene is shown. Figure S5. Localization of Arp6 and Swr1 on chromosome 5. The binding of Arp6-FLAG (top), Swr1-FLAG (middle), and Arp6-FLAG in swr1 cells (bottom) are compared. The position of Tel 5L, Tel 5R, CEN5, and the RP genes are shown under the panels. Figure S6. Preferential localization of Arp6 and Swr1 in the 5′ end of genes. Vertical bars represent the binding ratio of proteins in each locus. The binding of Arp6-Flag (Top), Swr1-Flag (middle), and Arp6-Flag in swr1 cells (bottom) in the region 228K-244K of Chr 6R were compared. The orientation of transcription of the genes of Watson strand and Crick strand in the region was shown by arrows in the map over the panels. Regions of divergent promoters are indicated with gray shadow. Figure S7. Correlation of the localizations of Arp6 and Swr1. The Arp6-binding log2 ratios of Arp6-binding loci (change p- value <0.025) in wild-type (A) and in swr1 cells (B) are represented as scatterplots versus the Swr1 binding log2 ratio in each Arp6 binding locus of wild-type cells. The yellow lines represent the hypothetical pattern of the data if Arp6 and Swr1 bind equally on the chromosomes. Figure S8. ChIP analysis for Htz1 in cells lacking Arp6 or Swr1. Htz1 association to the promoter of GAL1, SWR1, and ribosomal protein (RPL13A and RPS16B) genes was analyzed using ChIP with an anti-Htz1 antibody (abcam, ab4626) and quantified using real-time quantitative PCR in wild-type (WT), arp6, and swr1 cells. The values are indicated as percentage of input DNA obtained by ChIP with anti-Htz1 antibody. The data points represent the mean ± SD for at least three independent experiments. Figure S9. Quantitative analysis of RDS1 (YCR106W) and UBX3 (YDL091C) in arp6- and htz1-deletion mutants. The same amount of total RNA from wild-type, arp6, and htz1 cells was analyzed using real-time quantitative RT–PCR. The ACT1 gene was analyzed as a control. The relative amount of the transcript of the genes to ACT1 is shown. The data points represent the mean ± SD for at least three independent experiments. Figure S10. ChIP analysis for nuclear pore complex with GAL1 gene in arp6 cells. The association of GAL1 gene with NPC was analyzed using ChIP with an antibody against nuclear pore complex proteins (Mab414, abcam, ab24609) in wild-type (WT) and arp6 cells grown on the glucose- or galactose containing media. Immunoprecipitated DNA was quantified using real-time PCR probed for GAL1 gene. The percentage of recovered DNA over input is plotted relative to wild-type cells on glucose as 1. The data points represent the mean ± SD for at least three independent experiments. Table S1. Presence of Arp6 in nonrepetitive 10 kb subtelomere zones. (0.05 MB DOC) Table S2. Microarray analysis in arp6Δ and swr1Δ cells. (1.10 MB XLS) Table S3. Binding of Arp6 and Swr1 on ribosomal protein genes. (0.04 MB DOC) Table S4. Expression of RP genes in arp6Δ and swr1Δ cells. (0.04 MB XLS) Table S5. Genes markedly down-regulated in arp6 cells. (0.09 MB DOC) Table S6. Strains used in this study. (0.06 MB DOC) Text S1. Primer sequences. (0.04 MB DOC) SWR1 Arp6 Swr1 Arp6 in swr1 Yoshida et al., Supplementary Fig. S3B Arp6 Swr1 Arp6 in swr1 Yoshida et al., Supplementary Fig. S5 A Arp6 (log2) 4 3.5 3 2.5 r =0.278, n=2001 2 1.5 1 0.5 0 -2-101234567 Swr1 (log2) B Arp6 in swr1 (log2) 4 3.5 3 2.5 2 r =0.138, n=1463 1.5 1 0.5 0 -2-101234567 Swr1 (log2) Yoshida et al., Supplementary Fig. S6 ChIP analysis for Htz1 in arp6 and swr1 mutants 5 WT arp6 4 swr1 3 2 1 % precipitated by anti-Htz1 / input 0 GAL1 SWR1 RPL13A RPS16B Yoshida et al. Supplementary Fig. S7 Quantitative rtPCR for Arp6/Swr1-bound genes (x10- 3 ) 8 WT arp6 htz1 ACT1 6 4 2 Relative expression to 0 RDS1 UBX3 Yoshida et al., Supplementary Fig. S8 ChIP for GAL1-nuclear pore association p<0.05 2.0 WT arp6 1.5 1.0 % (IP/ input) / WT gluc 0.5 0 glucose galactose Yoshida et al., Supplementary Fig. S9 Supplementary Table S1. Presence of Arp6 in nonrepetitive 10 kb subtelomere zones Chr3L (32)1) Chr4L (4) Chr5L (15) Chr6R (34) Total (145) Chr3R (34) Chr4R (13) Chr5R (13) 12) (3%)3) 1 (25%) 3 (20%) Arp6 binding loci 7 (21%) 16 (11%) 3 (9%) 1 (8%) 0 (0%) SWR1 coincidence with 0 (0%) 0 (0%) 2 (13%) 2 (6%) 7 (5%) Swr1 binding 3 (9%) 0 (0%) 0 (0%) 3 (9%) 0 (0%) 0 (0%) swr1 Arp6 binding loci 9 (26%) 22 (15%) 4 (12%) 5 (38%) 1 (8%) 1) number of subtelomeric probes in the array, 2) number of probes on which the ChIP-chip signal is significantly positive, 3) percentage of binding-positive probes SupplemantaTable S2 arp6 swr1 arp6 log2 ratio swr1 log2 ratio t-test p-value t-test p-value YAL001C -0.48778725 0.055121132 -0.651383868 0.009641293 TFC3 transcription factor tau (TFIIIC) subunit 138 Vps8p is a membrane-associated hydrophilic protein which contains a C-terminal cysteine-rich region that conforms to the H2 variant of the YAL002W -0.50137568 0.02012999 0.465583742 0.134286021 VPS8 RING finger Zn2+ binding motif. YAL003W 0.291356151 0.57828274 0.018063402 0.741564133 EFB1 Translation elongation factor EF-1beta, GDP/GTP exchange factor for Tef1p/Tef2p YAL004W -1.37471567 0.023223502 0.446599197 0.094749311 strong similarity to A.klebsiana glutamate dehydrogenase YAL007C 0.514959137 0.367907397 0.193498974 0.126415188 ERP2 p24 protein involved in membrane trafficking YAL008W -1.07248083 0.003902375 0.04877654 0.757249306 FUN14 hypothetical protein YAL009W -0.83886578 0.039587079 -0.007318718 0.898104961 SPO7 sporulation protein YAL010C 0.350119155 0.079737228 -0.110610526 0.32107038 MDM10 Mitochondrial outer membrane protein involved in mitochondrial morphology and inheritance YAL011W -0.34259393 0.310843521 -0.299325162 0.15316135 possible mitochondrial transit peptide/weak similarity to Mus musculus p53-associated cellular protein YAL012W 1.573242516 0.034945842 0.519382719 0.240287741 CYS3 cystathionine gamma-lyase YAL013W 0.10593589 0.260657892 0.288869805 0.318643395 DEP1 regulation of phospholipid metabolism YAL014C 0.190903734 0.569458913 0.169774313 0.044229011 hypothetical protein YAL015C 0.370541536 0.031276471 -0.245836227 0.004561835 NTG1 DNA glycosylase YAL016W -0.64975335 0.0560383 0.036635636 0.638828131 TPD3 protein phosphatase 2A regulatory subunit A YAL017W -1.25829846 0.004518805 0.126513184 0.38324032 FUN31 Serine/threonine kinase YAL018C -0.51051588 0.164932688 -0.264509297 0.317542105 3 transmembrane domains/weak similarity to hypothetical proteins YOL047c and YOL048c YAL019W 0.320792247 0.444493655 -0.211820784 0.280255331 FUN30 SNF2 protein family YAL021C -0.1074359 0.778044634 -0.008790493 0.798167731 CCR4 95 kDa containng leucine rich tandem repeats YAL022C 0.656687352 0.001364239 0.428924658 0.115624767 FUN26 predicted membrane protein YAL023C 0.778388229 0.117515196 0.53048886 0.106271155 PMT2 dolichyl phosphate-D-mannose:protein O-D-mannosyltransferase YAL024C 0.327596538 0.491315967 0.065792226 0.797949412 LTE1 putative GTP-exchange protein YAL025C 1.742728628 0.005475432 -0.503643316 0.197141618 MAK16 putative nuclear protein YAL026C -0.15225171 0.144316487 0.062776186 0.624950436 DRS2 Membrane-spanning Ca-ATPase (P-type),member of the cation transport (E1-E2) ATPases YAL027W -0.19310171 0.23325453 -0.364863449 0.173323366 hypothetical protein YAL028W -1.05821689 0.028312621 -0.12586505 0.263146998 similarity to hypothetical protein YOR324c YAL029C 1.414871688 0.169872272 0.17340552 0.14232505 MYO4 myosin YAL030W -0.82724505 0.037223546 0.300700811 0.135945922 SNC1 homolog of Snc2p, vesicle-associated membrane protein (synaptobrevin) homolog, forms a complex with Snc2p and Sec9p YAL031C -0.66093199 0.001932142 0.091597547 0.555114187 FUN21 FUN21 YAL032C -0.45984855 0.100362467 0.119635039 0.26845368 PRP45 pre-mRNA splicing factor YAL033W 0.516623095 0.085119298 -0.332318037 0.10772256 POP5 An integral subunit of RNase P and apparent subunit of RNase MRP YAL034C -1.37859175 0.009174568 0.462426647 0.033887391 FUN19 Function unknown now YAL034W-A 0.046056852 0.601311241 -0.173771005 0.501575118 MTW1 determining metaphase spindle length YAL035C-A 0.455896607 0.408337787 0.428415177 0.170295679 YAL035W 0.75438664 0.04599226 -0.221359802 0.054514118 FUN12 97 kDa protein YAL036C 1.239106861 0.048698109 0.063806118 0.17119679 FUN11 similar to Xenopus GTP-binding protein DRG YAL037W -0.57645571 0.009922233 -0.165687374 0.532682496 strong similarity to GTP-binding proteins YAL038W 0.891293531 0.223803982 0.838341102 0.122662277 CDC19 Pyruvate kinase YAL039C 0.367285033 0.377663351 0.193111406 0.48023897 CYC3 cytochrome c heme lyase (CCHL) YAL040C -0.19482503 0.251202381 0.567158599 0.138150953 CLN3 G(sub)1 cyclin YAL041W 0.121678523 0.623119848 0.001064527 0.995446938 CDC24 Guanine nucleotide exchange factor (a.k.a.
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  • Supplementary Tables

    Supplementary Tables

    Supplementary Tables Supplementary Table S1: Preselected miRNAs used in feature selection Univariate Cox proportional hazards regression analysis of the endpoint freedom from recurrence in the training set (DKTK-ROG sample) allowed the pre-selection of 524 miRNAs (P< 0.5), which were used in the feature selection. P-value was derived from log-rank test. miRNA p-value miRNA p-value miRNA p-value miRNA p-value hsa-let-7g-3p 0.0001520 hsa-miR-1304-3p 0.0490161 hsa-miR-7108-5p 0.1263245 hsa-miR-6865-5p 0.2073121 hsa-miR-6825-3p 0.0004257 hsa-miR-4298 0.0506194 hsa-miR-4453 0.1270967 hsa-miR-6893-5p 0.2120664 hsa-miR-668-3p 0.0005188 hsa-miR-484 0.0518625 hsa-miR-200a-5p 0.1276345 hsa-miR-25-3p 0.2123829 hsa-miR-3622b-3p 0.0005885 hsa-miR-6851-3p 0.0531446 hsa-miR-6090 0.1278692 hsa-miR-3189-5p 0.2136060 hsa-miR-6885-3p 0.0006452 hsa-miR-1276 0.0557418 hsa-miR-148b-3p 0.1279811 hsa-miR-6073 0.2139702 hsa-miR-6875-3p 0.0008188 hsa-miR-3173-3p 0.0559962 hsa-miR-4425 0.1288330 hsa-miR-765 0.2141536 hsa-miR-487b-5p 0.0011381 hsa-miR-650 0.0564616 hsa-miR-6798-3p 0.1293342 hsa-miR-338-5p 0.2153079 hsa-miR-210-5p 0.0012316 hsa-miR-6133 0.0571407 hsa-miR-4472 0.1300006 hsa-miR-6806-5p 0.2173515 hsa-miR-1470 0.0012822 hsa-miR-4701-5p 0.0571720 hsa-miR-4465 0.1304841 hsa-miR-98-5p 0.2184947 hsa-miR-6890-3p 0.0016539 hsa-miR-202-3p 0.0575741 hsa-miR-514b-5p 0.1308790 hsa-miR-500a-3p 0.2185577 hsa-miR-6511b-3p 0.0017165 hsa-miR-4733-5p 0.0616138 hsa-miR-378c 0.1317442 hsa-miR-4515 0.2187539 hsa-miR-7109-3p 0.0021381 hsa-miR-595 0.0629350 hsa-miR-3121-3p
  • Potent Inhibition of Human Telomerase by U-73122

    Potent Inhibition of Human Telomerase by U-73122

    Journal of Biomedical Science (2006) 13:667–674 667 DOI 10.1007/s11373-006-9100-z Potent inhibition of human telomerase by U-73122 Yi-Jui Chen, Wei-Yun Sheng, Pei-Rong Huang & Tzu-Chien V. Wang* Department of Molecular and Cellular Biology, Chang Gung University, Kwei-San, Tao-Yuan, 333, Taiwan Received 28 April 2006; accepted 14 June 2006 Ó 2006 National Science Council, Taipei Key words: alkylating agents, cancer therapyzU-73122, N-ethylmaleimide, telomerase inhibitor Summary Telomerase activity is repressed in normal human somatic cells, but is activated in most cancers, suggesting that telomerase may be an important target for cancer therapy. In this study, we report that U-73122, an amphiphilic alkylating agent that is commonly used as an inhibitor for phospholipase C, is also a potent and selective inhibitor of human telomerase. The inhibition of telomerase by U-73122 was attributed primarily to the pyrrole-2,5-dione group, since its structural analog U-73343 did not inhibit telomerase. In confirmation, we observed that telomerase was inhibited by N-ethylmaleimide, but not N-ethylsuccinimide. The IC50 value of U-73122 for the in vitro inhibition of telomerase activity is 0.2 lM, which is comparable to or slightly more sensitive than that for phospholipase C. The inhibitory action of U-73122 on telomerase appears to be rather selective since the presence of externally added proteins did not protect the inhibition and the IC50 values for the other enzymes tested in this study were at least an order of magnitude higher than that for telomerase. Furthermore, we demonstrate that U-73122 can inhibit telomerase in hemato- poietic cancer cells.
  • Atherosclerosis-Susceptible and Atherosclerosis-Resistant Pigeon Aortic Cells Express Different Genes in Vivo

    Atherosclerosis-Susceptible and Atherosclerosis-Resistant Pigeon Aortic Cells Express Different Genes in Vivo

    University of New Hampshire University of New Hampshire Scholars' Repository New Hampshire Agricultural Experiment Station Publications New Hampshire Agricultural Experiment Station 7-1-2013 Atherosclerosis-susceptible and atherosclerosis-resistant pigeon aortic cells express different genes in vivo Janet L. Anderson University of New Hampshire, [email protected] C. M. Ashwell University of New Hampshire - Main Campus S. C. Smith University of New Hampshire - Main Campus R. Shine University of New Hampshire - Main Campus E. C. Smith University of New Hampshire - Main Campus See next page for additional authors Follow this and additional works at: https://scholars.unh.edu/nhaes Part of the Poultry or Avian Science Commons Recommended Citation J. L. Anderson, C. M. Ashwell, S. C. Smith, R. Shine, E. C. Smith and R. L. Taylor, Jr. Atherosclerosis- susceptible and atherosclerosis-resistant pigeon aortic cells express different genes in vivo Poultry Science (2013) 92 (10): 2668-2680 doi:10.3382/ps.2013-03306 This Article is brought to you for free and open access by the New Hampshire Agricultural Experiment Station at University of New Hampshire Scholars' Repository. It has been accepted for inclusion in New Hampshire Agricultural Experiment Station Publications by an authorized administrator of University of New Hampshire Scholars' Repository. For more information, please contact [email protected]. Authors Janet L. Anderson, C. M. Ashwell, S. C. Smith, R. Shine, E. C. Smith, and Robert L. Taylor Jr. This article is available at University of New Hampshire Scholars' Repository: https://scholars.unh.edu/nhaes/207 Atherosclerosis-susceptible and atherosclerosis-resistant pigeon aortic cells express different genes in vivo J.