Autosomal Dosage Compensation in Drosophila Melanogaster Strains Trisomic for the Left Arm of Chromosome 2 (Enzyme Levels/Gene Modulation/Regulatory Loci) ROBERT H

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Proc. Natd Acad. Sci. USA Vol. 79, pp. 1200-1204, February 1982 Genetics Autosomal dosage compensation in Drosophila melanogaster strains trisomic for the left arm of chromosome 2 (enzyme levels/gene modulation/regulatory loci) ROBERT H. DEVLIN, DAVID G. HOLM, AND THOMAS A. GIIGLIATTI Department of Zoology, University of British Columbia, Vancouver, British Columbia, Canada V6T 1W5 Communicated by M. M. Green, September 30, 1981 ABSTRACT Drosophila melanogaster individuals trisomic for tural genes are located on the left arm of chromosome 2, have an entire chromosome arm can survive to late' stages of pupal de- been measured in trisomies and in normal diploids. Addition- velopment. We have examined the levels of five enzymes whose ally, we have examined the activity oftwo enzymes whose struc- structural genes are located on the left arm ofchromosome 2 both tural genes are located elsewhere in the genome and, therefore, in trisomy 2L and in diploid strains. In trisomies, three distally remain diploid in all strains used. The result of these prelimi- mappingloci showedcompensated levels ofexpression close to that nary studies affords evidence for a form of autosomal dosage observed in the diploid strains. Analysis ofelectrophoretic variants compensation in Drosophila nelanogaster. of a compensated locus revealed that all three alleles are active in trisomies. Thai-two proximally located loci displayed dose-dependent levels of expression. Therefore, at the level of theindi- MATERIALS AND METHODS vidual gene, autosomal compensation appears to bean all-or-nene phenomenon. Furthermore, the compensatory response may be Genetic Strains and Procedures. The following strains of regionally distributed along the chromosome arm. The presence D. melanogaster were used: (i) Oregon R, (ii) C(2L)SHJ +; of both autosomal and sex-linked dosage compensation prompts F(2R)bw/F(2R)bw, (iii) C(2L)VHlit;F(2R)bw/F(2R)bw, (iv) us to speculate that these phenomenon are similar homeostatic C(2L)VD4aiGpdhB Bi/In(2L)Cy,al Cy GpdhA;F(2R)bw/ mechanisms that modulate gene expression both in euploid and F(2R)bw, (v) C(2L)P,b;C(2R)SK1A cn bw, (vi) al GpdhB Bi L/ aneuploid genomes. In(2L+2R)Cy,al Cy GpdhA, (vii) Df(2L)GdhA/In(2LR)SM1,Cy It, (viii) GpdhB0-l-4/In(2LR)SM1 ,Cy it, (ix) b AdhN2 osp pr cn, From individual to individual within a species, the chromosome (x) GpdhA, (xi) GpdhB, (xii) Gpdhc, and (xiii) several strains with complement is remarkably similar with the striking exception translocations between the Y and 2L chromosomes (3), which of the sex chromosomes. In many species, females have twice are listed in Table 2. Characteristics ofthe various mutant alleles as many X chromosomes as males. This imbalance in gene con- and chromosomes have been described previously (2, 5, 6). tent is rectified by dosage compensation, a regulatory mecha- Individuals trisomic for the left arm of chromosome 2 were nism that equalizes gene expression between individuals pos- produced as described (7, 8). Genetic and cytological studies sessing one or two X chromosomes (1). In mammals, equality revealed that whereas monosomic progeny die during embry- of X chromosome-linked gene expression occurs by inactivation ogenesis, a high proportion of the hybrid progeny inheriting ofone X chromosome in females through chromosome conden- three entire left arms of chromosome 2 survive to pharate sation. In contrast, dosage compensation in Drosophila is not adults. In fact, the viability of trisomy 2L- and trisomy X indi- mediated by chromosome condensation; instead, the- transcrip- viduals is comparable (data not shown). tional activity of X chromosome-linked genes is regulated to Five gene-enzyme systems on chromosome 2L, one on the synthesize the same amount ofgene product in both males and X chromosome, and one on chromosome 3L have been analyzed females. Thus, normal males (XY), normal' females (XX), and in this study. Their cytological map positions (2, 9-11) are listed metafemales (XXX) show approximately the same level of X in Table 1. To show that the activity of each enzyme was nor- chromosome-linked gene expression. Dosage compensation of mally proportional to the number ofgene copies, enzyme levels sex-linked genes in Drosophila is one ofthe few well-described were measured in strains with small duplications (three doses) examples of transcriptional modulation in eukaryotes. for each of the five loci studied on chromosome 2L. Segmental Previous studies on autosomal aneuploidy in metazoans have aneuploids were generated by a standard technique (3). not provided evidence for autosomal dosage compensation. For Flies trisomic for chromosome 2L, but carrying either a de- example, the amount of product synthesized by an autosomal ficiency or a null allele for a specific locus were produced by gene appears to be directly proportional to the number ofstruc- crossing C(2L)it;F(2R)bw/F(2R)bw females to males that were tural gene templates (2); individuals heterozygous for either a heterozygous for the inverted chromosome In(2LR)SM1,Cy it deficiency or a duplication of a particular autosomal gene pro- and either GpdhBol4 (a noncomplementing null allele for duce, respectively, 0.5 and 1.5 times the amount ofgene prod- Gpdh) or Df(2L)GdhA (a deficiency for the Gpdh structural uct found in normal diploid individuals. In addition, large al- gene) or to males homozygous for AdhN2 (a noncomplementing terations in the amount of chromosomal material are usually null allele for Adh). Larvae carrying either the null allele or lethal (3, 4). However, the complete absence ofautosomal dos- deficiency were distinguished by the presence of pigment in age compensation has never been critically tested. The purpose the larval Malpighian tubules in contrast to their nonpigmented ofthis investigation was to determine the level of gene expres- (It/l/lt sibs. sion in strains trisomic for the left arm ofchromosome 2 relative to normal diploids. The activity of five enzymes, whose struc- Abbreviations: dopa, dihydroxyphenylalanine; PGK, phosphoglycerate kinase;' aGPDH, a-glycerol-3-phosphate dehydrogenase; cMDH, cy- The publication costs ofthis article were defrayed in part by page charge toplasmic malate dehydrogenase; ADH, alcohol dehydrogenase; DDC, payment. This article must therefore be hereby marked "advertise- dopa decarboxylase; IDH, isocitrate dehydrogenase; 6PGD, 6-phos- ment" in accordance with 18 U. S. C. §1734 solely to indicate this fact. phogluconate dehydrogenase. 1200 Downloaded by guest on October 1, 2021 Genetics: Devlin et aL Proc. Natl. Acad. Sci. USA 79 (1982) 1201

Table 1. Enzyme levels in diploid and trisomic individuals Enzyme and chromosome position Pgk Gpdh cMdh Adh Ddc Idh 6Pgd Genotype 22C-23E 25F-26B 30F-31D 35B2 37B10-37C7 66B-67C 2D3-2D6 Oregon R 0.986 + 0.031 0.212 ± 0.008 3.36 ± 0.056 67.6 ± 2.4 8.61 ± 1.02 110 ± 4.3 36.8 ± 1.2 (10) (14) (10) (14) (16) (10) (10) C(2L)+;F(2R)bw/ 0.831 ± 0.019 0.132 ± 0.008 3.71 ± 0.053 40.9 ± 1.7 10.9 ± 0.784 73.3 ± 2.8 54.0 ± 1.1 F(2R)bw (11) (14) (10) (14) (18) (7) (10) C(2L)+/ 0.973 ± 0.049 0.154 ± 0.005 3.91 ± 0.294 89.0 ± 1.9 16.7 ± 0.714 74.9 ± 1.0 40.2 ± 1.6 +/F(2R)bw(CF Y (10) (10) (4) (10) (20) (10) (10) x OR d) C(2L)+/+/F(2R)bw 0.168 ± 0.019 3.73 ± 0.102 88.1 ± 1.7 81.5 ± 4.0 (ORe x CF d) (3) (11) (3) (3) Noncompensated 1.32 ± 0.016 0.238 ± 0.006 5.39 ± 0.044 74.7 ± 1.80 15.2 ± 0.552 Estimated mean estimate 91.7 ± 1.39 49.7 ± 0.576 Compensated estimate 0.883 ± 0.011 0.159 ± 0.004 3.59 ± 0.029 49.8 ± 1.20 10.1 ± 0.348 Enzyme activities expressed as the mean ± SE in units/mg of protein for Pgk, Gpdh, and cMdh; in units x 103/mg of protein forAdh, Idh, and 6Pgd; and in units x 106/mg of protein for Ddc. Estimates of enzyme activities expected in trisomies for dose-dependent (noncompensated) and compensated gene expression are provided for loci on chromosome 2L, and expected mean values for Idh and 6Pgd are also shown. Numbers in brackets represent the sample sizes. CF = C(2L); F(2R)/F(2R) strain; OR = Oregon R strain.

A compound chromosome heterozygous for two Gpdh elec- 1010 becquerels) of [1-'4C]dopa (New England Nuclear; 3 Ci/ trophoretic variants was constructed by mating C(2L)P,b; mmol; isocitrate dehydrogenase (IDH; EC 1.1.1.42), 0.1 M C(2R)SKlA,cn bw males to al GpdhB Bi L/In(2L+2R)Cy, Tris, pH 8/5 mM Mg2+/0.05 mM NADP/0.5 mM isocitrate; al Cy GpdhA females that had been treated with 2500 rads from and 6-phosphogluconate dehydrogenase (6PGD; EC 1.1.1.44), a 'oCo source. A C(2L)VD4al GpdhB Bl/In(2L)Cy,al Cy GpdhA; 0.1 M Tris, pH 7.5/1.3 mM NADP/20 mM MgC12/3.1 mM 6- C(2R) SKlA,cn bw individual was recovered, and a stock was phosphogluconate. Activities are expressed as units per mg of established. From a cross of females bearing this compound protein in the extract, where one unit is defined as the utili- 2L chromosome with C(2L)P,b;F(2R)bw/F(2R)bw males, zation of1 Amol ofsubstrate per min. Protein concentration was C(2L)VD4al GpdhB Bl/In(2L)Cy,al Cy GpdhA;F(2R)bw/ determined (13), with bovine serum albumin as the standard. F(2R)bw nondisjunctional progeny were recovered. These in- The enzyme levels expected in the absence of dosage com- dividuals, when crossed to flies possessing the Gpdhc allele on pensation were estimated by summing the activity found in the standard second chromosomes, produce trisomies containing compound-free strain plus one half of the activity found in the three different electrophoretic variants. Oregon R strain. This corresponds to the parental contribution All strains were reared at 250C on standard cornmeal sucrose of chromosome 2L to the karyotype of the trisomies. The en- medium supplemented with the mold inhibitor Tegosept zyme levels expected in the presence ofautosomal dosage com- (0.22%) and the antibiotics tetracycline and streptomycin at 10 pensation were estimated as simply two-thirds of the expected mg/liter each. noncompensated value. Enzyme Assays. All determinations of enzyme activity were Electrophoretic Procedures. Electrophoresis of aGPDH in performed on 50- to 150-mg samples of late third instar larvae trisomies possessing three variants was performed in a 12% (wt/ collected from the sides ofuncrowded half-pint culture bottles. vol) starch/0. 1 M Tris citrate gel system, pH 6.9, (14) run at Homogenates were prepared from 25 mg oflarvae per ml of 10 300 V (25-35 mA) for 32 hr. Gels were stained for GPDH (14). mM phosphate buffer (pH 8.2) for all enzymes except dihydroxyphenylalanine (dopa) decarboxylase (DDC), for which 0.1 RESULTS M Tris (pH 7.2) was used. After centrifugation, enzyme activity The effect ofaneuploidy for small genetic segments on enzyme (except for DDC) was monitored by the change in absorption activity has been extensively examined in a number of species. at 340 nm by the production or utilization ofNADH or NADPH; Indeed, changes in enzyme activity associated with aneuploids DDC activity was determined from the amount of '4Co2 re- are often used to locate the structural gene of an enzyme or a leased from [1-14C] dopa, corrected for spontaneous decarbox- dose-dependent locus regulating its expression (2, 15). Because ylation in the presence ofheat-inactivated extract (12). All assays little is known about the effect of autosomal trisomy on gene were optimized with respect to all components. The conditions expression, we examined seven gene-enzyme systems located for each assay were as follows: phosphoglycerate kinase (PGK; within and outside a trisomic region, namely chromosome 2L. EC2.7.2.3), 0.1 MTris, pH 7.6/0.9mM EDTA/15mM NADH/ This was accomplished by measuring enzyme levels in diploid 2.2 mM ATP/13 mM glycerate-3-phosphate/3.2 mM MgSOg parental stocks and their trisomic offspring. The results for the 2.5 units of glyceraldehyde-3-phosphate dehydrogenase (EC two diploid parental stocks, Oregon R and C(2L)+,F(2R)bw/ 1.2.1.12) per ml; a-glycerol-3-phosphate dehydrogenase (NA F(2R)bw, and for their trisomy 2L offspring are presented in D+) (aGPDH EC 1.1.1.8), 0.1 M glycine, pH 9.5/2.5 mM glyc- Table 1. From previous findings with segmental aneuploids for erol-3-phosphate/10 mM NAD+; cytoplasmic malate dehydro- autosomal regions, a simple prediction is that genes duplicated genase (cMDH; EC 1.1.1.37), 50 mM phosphate, pH 7.3/0.25 in trisomies would show strict dose dependence. Accordingly, mM NADH/5 mM oxaloacetic acid; alcohol dehydrogenase the expected enzyme level would simply represent the sum of (ADH; EC 1.1.1.1.), 0.1 M glycine phosphate buffer, 1.7 g/ the activities of each of the three gene copies. Alternatively, a liter, pH 9.0/2.5 mM NAD+/2% ethanol; dopa decarboxylase compensatory mechanism may exist for controlling the expres- (DDC; EC 4.1.1.26), 0.1 M Tris, pH 7.2/0.12 mM pyridoxal- sion of the genes in this trisomic arm. The simplest prediction 5'-phosphate/25 mM phenylthiourea/0. 1 ILCi (1 Ci = 3.7 X would be that compensation acts on all three alleles equally, Downloaded by guest on October 1, 2021 1202 Genetics: Devlin et al. Proc. Natl. Acad. Sci. USA 79 (1982)

1.0- 0.10- a 0 "aa) 2 -- - 0) 00 K 0) 0.66 0.66- a) d) K an qa0 a) K 1 1 0.33 0.33 - K K I I I K Pgk Gpdh cMdh Adh Ddc C(2L)lt C(2L)lt C(2L)lt C(2L)lt C(2L)lt Gene-enzyme system Df(2L)GdhA BO-1-4 AdhN2 Diploid Trisomy Trisomy Diploid Trisomy FIG. 1. Enzyme activities in trisomy 2L individuals relative to expected levels. Level 2 gene-dose expressed represents the expected com- FIG. 2. Expression of enzyme loci on tne C(2L)lt chromosome in pensated level of expression; level 3 gene-dose expressed represents diploids and in trisomies possessing either a null allele or a deficiency the expected noncompensated estimate. forthe loci measured. Only the genotype of the left arm of chromosome 2 is given. Activities from loci on the C(2L)lt chromosome are graphed relative to the diploid level. All standard errors were less than 5% of in a resulting diploid level ofactivity, approximately two-thirds the mean. (Left) Gpdh. (Right) Adh. that of the noncompensated level. The observed and expected enzyme levels are presented in Table 1. The results for the five For the observed autosomal dosage compensation to be a loci on chromosome 2L are summarized in Fig. 1. The enzyme valid phenomenon, the compensating loci must be dose de- activities ofthree loci, Pgk, Gpdh, and cMdh, are clearly at the pendent in diploid strains with small duplications, at the same level expected if compensation occurs. These loci are located stage ofdevelopment. Small duplications ofthe structural genes in approximately the distal two-thirds of the chromosome arm. for all five enzymes were made, and enzyme assays were per- In contrast, the enzymes produced by the two proximally located loci, Adh and Ddc, show dose-dependent levels of activ- formed on mature third instar larvae (Table 2). The enzyme activities in these aneuploids showed dose dependence; that is, ity, indicating an absence ofdosage compensation. ADH activity the levels in the strains with three doses were significantly is significantly higher than expected, even for a noncompen- higher than those of their diploid parents. sated gene. Thus, it appears that some loci in trisomic individ- Simplistically, compensation may occur by one of two mech- uals are capable of or are directed toward compensation, anisms: first, each allele may be expressed at two-thirds of the whereas others are not, and this difference appears to be re- normal diploid level or, second, one allele may be completely gionally distributed along the chromosome arm. Results from reciprocal crosses were statistically indistinguishable (a = 0.05). Table 2. Dose dependence of enzyme levels in small segmental The structural genes for IDH on chromosome 3L and 6PGD aneuploids in late third instar larvae on the X chromosome were monitored to observe what effect, Dupli- Translo- if any, trisomy 2L might have on the activities ofunlinked loci. Gene cation cation Dose Activity Ratio The enzyme activities of these loci in trisomies was approximately 15% below the expected diploid level (Table 1). Pgk 22D-24A G120 2 0.559 ± 0.018 (5) R136 2 0.535 ± 0.009 (7) 1.31 A direct assessment ofthe effect oftrisomy on the expression DpG120-R136 3 0.714 ± 0.033 (8) of a gene can be made by comparing the activity of the same two alleles in both diploid and trisomy 2L strains. This was ac- Gpdh 25F-26B D106 2 0.113 ± 0.008 (5) complished by comparing the enzyme activity ofindividual loci G105 2 0.136 ± 0.008 (5) 1.66 on the same C(2L) chromosome in C(2L)-bearing diploid strains Dp D106-G105 3 0.207 ± 0.007 (5) with a trisomic strain in which the standard chromosome carried either a null allele or a deficiency for the enzyme locus in ques- cMdh 30F-31EF L52 2 1.53 ± 0.098 (3) tion. The results are shown in Fig. 2. For Gpdh, the expression A162 2 1.44 ± 0.130 (3) 1.29 of the two active alleles is reduced by approximately 1/3rd in Dp L52-A162 3 1.91 ± 0.040 (5) the trisomic strains carrying either a null allele or a deficiency for the structural gene, clearly demonstrating that the expres- Adh 34B-35D G74 2 39.1 ± 0.670 (6) sion ofthis gene-enzyme system is compensated in trisomy 2L P58 2 36.7 ± 0.680 (6) 1.79 individuals. In an analogous experiment involving the noncom- Dp G74-P58 3 68.0 ± 1.60 (6) pensating locus, Adh, the expression of the two active alleles in the C(2L) chromosome was unaffected in trisomic individuals Ddc 36EF-37D D219 2 11.03 ± 0.85 (4) bearing an Adh null allele on the standard second chromosome. H174 2 11.70 ± 0.76 (4) 1.41 These results demonstrate that the response of compensating Dp D219-H174 3 15.98 + 0.36 (4) and noncompensating genes is independent of the number of Activity units are the same as in Table 1. Ratio represents the en- active alleles, an observation that has been long known for the zyme levels found in duplication-bearing larvae relative to the mean X chromosome. of that found in their parents. Downloaded by guest on October 1, 2021 Genetics: Devlin et aL Proc. Natd Acad. Sci. USA 79 (1982) 1203

(50.1) and Ddc (53.9), remain dose dependent in whole-arm +ve trisomies. Thus, autosomal dosage compensation appears to be an all-or-none phenomenon acting at the level of the individual gene. The results ofthe five gene-enzyme systems studied suggest that the distal two-thirds ofthe chromosome compensates while the proximal one-third does not (Fig. 1). One can speculate that A~~~~~~~A this regionality in the compensatory response reflects either a difference in gene organization or a difference in chromosome structure capable of responding to compensation. However, this regionality may be simply an artifact of the small number of loci sampled. Although several loci on the X chromosome of B A C Tm Tf A A B TmTfB A C D. melanogaster are not dosage compensated, they are not lo- B CaC cated in distinct regions (19-22). In contrast, regionality for sex- linked dosage compensation has been observed in a related spe- FIG. 3. Electrophoretic analysis of Gpdh variants in trisomies. B, cies, D. miranda (23). A, and C represent mobility variants for Gpdh. Lanes B, A, and C are To further examine the response of compensated and non- from diploid homozygous individuals; lanes A/B, A/C, and B/C are compensated loci in trisomies, the same two active alleles for from diploid heterozygotes; lanes designated T are from trisomies pos- bothAdh and for Gpdh were measured in diploid and in trisomic sessing all three variants produced from reciprocal crosses (where Tm andTf represent trisomic offspring produced from compound-free male strains (Fig. 2). In the trisomic strains, the standard chromo- and female parents, respectively). +ve, Mobility towards anode. some possessed either a null allele or a deletion that eliminated the activity ofthe third allele. ForAdh, the activity ofeach allele was identical both in trisomic and in diploid strains, as expected repressed while the other two are fully active. To distinguish of any noncompensated locus. The identical expression ofAdh between these two alternatives, trisomies possessing three elec- alleles in trisomic and diploid strains suggests that any physi- trophoretic variants for Gpdh were produced. Starch gel elec- ological differences between them is minimal. In contrast, the trophoresis of this enzyme in trisomies demonstrated that all expression of each Gpdh allele in the trisomic strain was re- three alleles of this compensated gene were expressed (Fig. 3). duced by 1/3rd relative to the expression of this same allele in the diploid strain. This is precisely the response expected from DISCUSSION a compensated locus and suggests that autosomal dosage com- Dosage compensation is one of the few well-documented ex- pensation occurs at the level ofgene transcription. Indeed, anal- amples of repression (mammals) or modulation (invertebrates) ysis of trisomies possessing three electrophoretic variants for of gene transcription in higher animals. In Drosophila, X chro- aGPDH (Fig. 3) demonstrated that all three loci were mosome-linked loci transcribe at different and predictable rates expressed. depending on the genotype (1). The total X-linked transcription To examine the influence oftrisomy 2L on unlinked loci, the within a diploid nucleus is constant, regardless of the number levels ofIDH from chromosome 3L and 6PGD from the X chro- of X chromosomes. Thus, the transcriptional rate of a single X mosome were measured, In the trisomies, each displayed a chromosome-linked structural gene varies in direct proportion level somewhat lower than expected (approximately 15%). to the number of X chromosome-linked templates present in Whether this reflects a general effect of trisomy on nonlinked the nucleus. For example, individuals with one or with three genes, differences in gender, or simply genetic variation (24, X chromosomes transcribe X-linked loci at twice or at two-thirds 25) (isogenic strains were not used) awaits a more extensive sur- the rate of diploid females, respectively. In addition, the total vey ofother unlinked loci and other trisomies. Becausewe have level ofX chromosome-linked transcription is directed to match not established that the decreases in IDH and 6PGD activities that of the autosomes and is directly dependent on their num- are a specific effect of trisomy, we hesitate to use them as in- ber. For example, X-linked transcription in individuals pos- ternal standards. We suggest, therefore, that in dealing with sessing two X chromosomes and three sets of autosomes (a tri- grossly altered genotypes such as trisomies, any changes in en- ploid intersex) is 50% greater than normal diploid females (two zyme activity should be interpreted initially as locus specific X chromosomes and two sets of autosomes). rather than reflective of total genomic activity. The observation that individuals trisomic for the X chromo- Our data raise two questions. First, by what mechanism does some show dosage compensation (16, 17) is of particular rele- autosomal dosage compensation occur and, second, why should vance to this study. Whereas diploid individuals with one, two, it exist for autosomal aneuploids? Autosomal and X chromo- or three X chromosomes express sex-linked traits equally, males some-linked dosage compensation appear analogous in many or females having small duplications or deletions of X chro- ways; therefore, it is tempting to assume that their molecular mosome genes do show dose-dependent levels of gene expres- bases are similar. Two mechanisms for sex-linked dosage com- sion (1, 18). pensation have been hypothesized, and both are presumed to In contrast to X chromosome-linked loci, it has generally act at the level oftranscription. One model is based on repres- been assumed that autosomal loci are dose dependent regard- sion and the other is based on activation of X chromosome- less ofthe chromosomal constitution. Genes in small duplicated linked gene expression. The first model postulates the existence segments of the autosomes, like their X chromosome-linked ofX chromosome-linked loci, themselves not dosage compen- counterparts, produce dose-dependent levels of gene product sated, which modulate the activity of other X chromosome- (Table 2). However, in individuals trisomic for the entire left linked genes by partially repressing transcription (19). As the arm ofchromosome 2, three distally mapping enzyme loci, Pgk number of X chromosomes increases, the concentration of the (7.6), Gpdh (20.5), and cMdh (37.2), show levels of activity ex- repressor substance increases and the amount of transcription pected of a diploid. Thus, a compensatory mechanism appears from each X chromosome-linked template decreases (compen- to be acting. However, not all genes on the trisomic chromo- sation). Evidence has been presented for (26) and against (17) some arm are compensated. Two proximally located loci, Adh the existence ofthese repressor loci on the X chromosome. The Downloaded by guest on October 1, 2021 1204 Genetics: Devlin et al. Proc. Natl Acad. Sci. USA 79 (1982)

second model suggests that the transcription ofX chromosome- pressed in trisomies. In all ofthese respects, autosomal dosage linked genes is modulated by nonlinked loci that produce a lim- compensation resembles sex-linked dosage compensation in ited amount of a diffusible activator (27, 28). According to this Drosophila. This prompts us to suggest that the basis for dosage model, the X chromosome-linked genes compete for the acti- compensation may be a normal process for maintaining home- vator. Increases or decreases in the number of X chromosomes ostasis which operates in euploid as well as aneuploid strains. have no effect on the total amount of transcription from the X- linked genes because the amount ofactivator substance remains We thank Dr. R. Macdntyre for kindly providing the Gpdh variants, Dr. D. T. Suzuki for his many helpful comments and extensive review- constant. Recently, evidence has been presented for the latter ing of the manuscript, and Drew Robinson for typing the manuscript. model of sex-linked dosage compensation (18, 27). This investigation was supported by grants to D.G.H. from the Natural Either model can adequately account for the compensation Science and Engineering Research Council (A-5853) and to T.A.G. from we found in trisomy 2L strains. Experiments are in progress to the Natural Science and Engineering Research Council (A-1764) and test the application of these two models to autosomal dosage British Columbia Health Care Foundation (12-80-2). This report is compensation. Indeed, Rawls and Lucchesi (15) have reported taken in part from material to be submitted in partial fulfillment of re- that regions of chromosome 2L are capable of reducing the ac- quirements for the degree of Doctor of Philosophy in Zoology at the tivity of aGPDH, indicating regulatory loci may exist on chro- University of British Columbia by R.H.D. mosome 2L. 1. Lucchesi, J. C. (1978) Science 202, 711-716. It is not surprising that a mechanism exists to compensate for 2. O'Brien, S. J. & Macdntyre, R. (1976) in The Genetics and Biol- the imbalance in X chromosome-linked templates which occurs ogy ofDrosophila, eds. Wright, T. & Ashburner, M. (Academic, between males and females in many higher organisms. How- New York), Vol. 2b, pp. 395-551. ever, autosomes generally do not vary in number and, there- 3. Lindsley, D. L., Sandler, L., Baker, B. S., Carpenter, A. T. C., fore, it is not obvious why autosomal dosage compensation Denell, R. E., Hall, J. C., Jacobs, P. A., Gabor Miklos, G. L., Davis, B. K., Gethmann, R. C., Hardy, R. W., Hessler, A., should exist. The fact that survival and compensation can occur Miller, S. M., Nozawa, H., Parry, D. M. & Gould-Somero, M. for both X chromosomal and autosomal trisomies leads us to (1972) Genetics 71, 157-184. speculate that in Drosophila, perhaps all forms of dosage com- 4. Stewart, B. & Merriam, J. R. (1973) Drosoph. Inf. Serv. 50, pensation simply reflect the existence of a homeostatic mech- 167-170. anism that operates at all times. Such a control mechanism 5. Lindsley, D. L. & Grell, E. (1967) Carnegie Inst Washington would be responsible for modulating the overall level of gene PubI 677, 1-477. 6. Grell, E. H. (1970) Genetics 65, 65-74. expression, even in normal diploids. What we observe as com- 7. Fitz-Earle, M. & Holm, D. G. (1977) Proceedings ofthe 15th In- pensation in trisomies may simply be this normal homeostatic ternational Congress of Entomology, Washington, DC, August mechanism forced to cope with extreme perturbations within 19-27, 1976, ed. White, D. (Entomological Society of America, the genome. Washington, DC), pp. 146-156. Further evidence for autosomal dosage compensation may 8. Fitz-Earle, M. & Holm, D. G. (1978) Genetics 89, 499-510. be found within the of the genus Dro- 9. Voelker, R. A., Ohnishi, S. & Langley, C. H. (1979) Biochem. evolutionary history Genet. 17, 769-783. sophila. Fusion of autosomal and X chromosome arms to form 10. Voelker, R. A., Ohnishi, S. & Langley, C. H. (1979) Biochem. X chromosomes of varying size and shape has occurred in the Genet. 17, 947-956. genus Drosophila (29) and appears to be a common mechanism 11. Woodruff, R. C. & Ashburner, M. (1979) Genetics 92, 117-132. ofspeciation. In fact, among present species, only the right arm 12. Hodgetts, R. B. & Konopka, R. J. (1973) J. Insect Physiot 19, of chromosome 3 and the fourth chromosome, as defined in D. 1211-1220. melanogaster, never appear as an X chromosome in other spe- 13. Lowry, 0. H., Rosebrough, N. J., Farr, A. L. & Randall, R. J. (1952) J. Biol Chem. 193, 265-275. cies. Although small differences in the genetic constitution of 14. Shaw, C. R. & Prasad, R. (1970) Biochem. Genet. 4, 297-320. a chromosome arm do exist between species, the majority of 15. Rawls, J. M., Jr., & Lucchesi, J. C. (1974) Genet. Res. Camb. 24, chromosomal material is identical. For example, D. pseudob- 59-72. scura has a metacentrix X chromosome comprised of the X and 16. Lucchesi, J. C., Rawls, J. M., Jr., & Maroni, G. (1974) Nature 3L chromosomal elements of D. melanogaster. Enzyme (30) (London) 248, 564-567. and heat shock (31) loci on the right arm ofthe D. pseudobscura 17. Stewart, B. R. & Merriam, J. R. (1975) Genetics 79, 635-647. 18. Maroni, G. & Lucchesi, J. C. (1980) Chromosoma 77, 253-261. X chromosome (melanogaster 3L chromosome) have been 19. Muller, J. H. (1950) Harvey Lect., Ser. 43, 1, 165-229. shown to be dosage compensated. This demonstrates that at 20. Smith, P. D. & Lucchesi, J. C. (1969) Genetics 61, 607-618. least one chromosomal element can display compensation or not 21. Stern, C. (1960) Can. J. Genet. Cytot 2, 105-118. (albeit between species), an ability which may have allowed the 22. Roberts, D. B. & Evans-Roberts, S. (1979) Nature (London) 280, evolution and modification of various X chromosomes. 691-692. There is also evidence for dosage compensation in the plant 23. Strobel, E., Pelling, C. & Arnheim, N. (1978) Proc. Natl Acad. Sci. USA 75, 931-935. t kingdom. For example, in Zea maize, whole arm duplications 24. Laurie-Ahlberg, C. C., Maroni, G., Bewley, G. C., Lucchesi, J. for chromosome 1L, which contains the Adh locus, do not show C. & Weir, B. S. (1980) Proc. Natl Acad. Sci. USA 77, 1073-1077. dose dependence for ADH activity (32). 25. Bijlsma, R. (1980) Biochem. Genet. 18, 699-715. In summary, we have demonstrated autosomal dosage com- 26. Birchler, J. A. (1980) Genetics 94, S9 (abstr.). pensation in whole arm trisomies of Drosophila melanogaster. 27. Maroni, G. & Plaut, W. (1973) Chromosoma 40, 361-377. However, we find that not all genes on the trisomic arm are 28. Schwartz, D. (1973) Genetics 75, 639-641. 29. Patterson, J. T. & Stone, W. S. (1952) Evolution in the Genus compensated. At the level ofthe individual gene, compensation Drosophila (MacMillan, New York). appears to be an all-or-none phenomenon. The compensatory 30. Abraham, I. & Lucchesi, J. (1974) Genetics 78, 1119-1126. response may take place at the level of transcription because 31. Pierce, D. A. & Lucchesi, J. (1980) Chromosoma 76, 245-254. all three alleles of the compensated locus, Gpdh, were ex- 32. Birchler, J. A. (1981) Genetics 97, 625-637. Downloaded by guest on October 1, 2021