鸡和人性染色体的趋同进化

1、Convergent Evolution of Chicken Z and Human X Chromosomes by Expansion and Gene AcquisitionDaniel W. Bellott1, Helen Skaletsky1, Tatyana Pyntikova1, Elaine R. Mardis2, Tina Graves2,Colin Kremitzki2, Laura G. Brown1, Steve Rozen1, Wesley C. Warren2, Richard K. Wilson2,and David C Page11 Howard Hughes

2、 Medical Institute, Whitehead Institute, and Department of Biology,Massachusetts Institute of Technology, 9 Cambridge Center, Cambridge, Massachusetts 02142,USA2 The Genome Center, Washington University School of Medicine, 4444 Forest Park Boulevard,St. Louis Missouri 63108, USAAbstract摘要In birds, a

3、s in mammals, one pair of chromosomes differs between the sexes. In birds, males are ZZ and females ZW. In mammals, males are XY and females XX. Like the mammalian XY pair, the avian ZW pair is believed to have evolved from autosomes, with most change occurring in the chromosomes found in only one s

4、ex the W and Y chromosomes15. By contrast, the sex chromosomes found in both sexes the Z and X chromosomes are assumed to have diverged little from their autosomal progenitors2. Here we report findings that overturn this assumption for both the chicken Z and human X chromosomes. The chicken Z chromo

5、some, which we sequenced essentially to completion, is less gene-dense than chicken autosomes but contains a massive tandem array containing hundreds of duplicated genes expressed in testes. A comprehensive comparison of the chicken Z chromosome to the finished sequence of the human X chromosome dem

6、onstrates that each evolved independently from different portions of the ancestral genome.正如在哺乳动物中那样，在鸟类中，不同性别的个体有一对染色体不同。鸟类雄性的性染色体组成为ZZ，雌性则为ZW。在哺乳动物中，则是雄性为XY，雌性为XX。与哺乳动物XY这对染色体相同，大部分变化发生在只有一种性别具有的染色体上W和Y染色体，因而鸟类的ZW染色体也被认为是由常染色体演化而来。与之相反，两种性别都具有的染色体Z和X染色体，被认为与它们的常染色体祖先并没有很大差别。这里将报告我们推翻先前关于鸡Z和人X染色体设想

7、的发现。我们对于鸡Z染色体的测序工作基本完成，发现其基因密集程度低于常染色体，但却含有大量的串联重复序列，它们包含在睾丸中表达的数以百计的重复基因。通过对鸡Z染色体和已完成的人类X染色体进行综合比对，我们发现它们是由原始染色体组的不同部分独立演化而来的。Despite this independence, the chicken Z and human X chromosomes share features that distinguish them from autosomes: the acquisition and amplification of testis-expressed g

8、enes, as well as a low gene density resulting from an expansion of intergenic regions. These features were not present on the autosomes from which the Z and X chromosomes originated but were instead acquired during the evolution of the Z and X as sex chromosomes. We conclude that the avian Z and mam

9、malian X chromosomes followed convergent evolutionary trajectories, despite their evolving with opposite (female vs. male) systems of heterogamety. More broadly, in birds and mammals, sex chromosome evolution involved not only gene loss in sex-specific chromosomes, but also marked expansion and gene

10、 acquisition in sex chromosomes common to males and females.除了独立起源，鸡Z和人X染色体还有一些将它们与常染色体区分开的共同特征：睾丸表达基因的获得和表达，以及基因间隔区域扩大导致的低基因密度。这些特征并没有出现在Z和X染色体所起源的常染色体上，而是Z和X染色体在逐渐演化形成的过程中获得的。我们推断，尽管鸟类Z和哺乳动物X染色体进化形成了相反的配子异型系统，但它们遵循着趋同进化的轨迹。更广泛地说，鸟类和哺乳动物的性染色体进化不仅涉及特定性染色体上基因的丢失，而且包括性染色体上的显著扩增和基因的获得，这一特点是为雌性和雄性所共有的。正

11、文A century ago, Herman Muller proposed the first theory of sex chromosome evolution that the X and Y chromosomes of Drosophila evolved from an ordinary pair of autosomes, and that genes on the Y chromosome had gradually deteriorated while their counterparts on the X were preserved1. In the 1960s, Su

12、sumu Ohno applied Mullers theory to the sex chromosomes of vertebrates, arguing that while the sex-specific W and Y chromosomes of birds and mammals had degenerated, the content of the Z and X chromosomes remained intact2. Four decades on, comparisons of the human X and Y chromosomes have underscore

13、d the dramatic evolutionary changes on the Y chromosome,35 but the assumption that the X chromosome has been evolutionarily stable remains unexamined.一个世纪前，Herman Muller提出了性染色体进化的第一个理论果蝇的X和Y染色体是由一对常染色体进化而来，而且Y染色体上的基因逐渐改变但它们在X染色体上相应的基因则被保留下来1。在20世纪60年代，Susumu Ohno将Muller的理论应用于脊椎动物的性染色体进化，认为尽管鸟类和哺乳动物的

14、“性别决定染色体”W和Y染色体已经退化，但Z和X染色体上的基因仍然是完好无缺的2。40年以来，通过对比人类X和Y染色体，Y染色体巨大的进化上的改变已经被揭示并且得到重视3-5，但关于X染色体在进化上保持稳定的假想却一直没有得到检验。The evolutionary relationship between the mammalian X chromosome and the avian Z chromosome has been the subject of much speculation, but it also remains unresolved. Ohno conjectured t

15、hat the X chromosomes of mammals were orthologous to the Z chromosomes of birds2. However, comparative mapping of 30 Z-linked genes indicated that the chicken Z chromosome was orthologous to human chromosomes 5, 8, 9, and 18, not to the human X chromosome6,7. These findings were supported by the dra

16、ft sequence of the chicken genome, but only about one third of the sequence of the Z chromosome was present in the assembly, leaving open the possibility that regions of orthology between the avian Z and mammalian X had yet to be detected8. The recent discovery that a subset of the five platypus X c

17、hromosomes contain orthologs of genes on the chicken Z chromosome renewed speculation that the avian Z and mammalian X share a common origin912. To accommodate the results of comparative gene mapping experiments, some have proposed that the chicken Z and human X were derived from different portions

18、of an ancestral proto sex chromosome which broke apart, leaving Z orthologs autosomal in mammals and X orthologs autosomal in birds11,12.虽然关于哺乳动物X染色体和鸟类Z染色体的进化关系已经有了很多假说，但都没能得到验证。Ohno猜想哺乳动物X染色体和鸟类的Z染色体属于直系同源染色体2。但是，对30个Z染色体连锁基因的对比作图显示，鸡的Z染色体与人类5，8，9号染色体以及18号染色体直系同源，而不是人类的X染色体6-7。这些发现有鸡的基因组序列草图支持，但当时

19、Z染色体序列只装配好了三分之一，仍然存在鸟类Z染色体和哺乳动物X染色体的同源区域未被发现的可能8。近期发现，5个鸭嘴兽的X染色体上含有鸡Z染色体的同源基因。这一发现也使鸟类Z和哺乳动物X染色体具有同一起源的推测重新得到人们重视9-12。为了解释对比基因作图与该发现的矛盾，一些人提出鸡Z染色体和人X染色体起源于同一祖先性染色体的不同部分，是由其断裂形成的，因此Z染色体才会和哺乳动物常染色体同源而X染色体和鸟类的常染色体同源11,12。To reconstruct and compare the evolutionary trajectories of the avian Z and mammal

20、ian X chromosomes, we have produced the finished sequence of the chicken Z chromosome, the first for any Z chromosome (Supplementary Figures 13). The resulting sequence spans roughly 80 megabases (Mb), is complete apart from four gaps, and is accurate to about one nucleotide per megabase. The chicke

21、n Z chromosome contains 1000 genes (Supplementary Table 1). This makes the Z chromosome less gene-dense than any chicken autosome, with 11 genes per megabase, which is less than half of the chicken autosomal average of 25 genes per megabase (Table 1)8. Conversely, the density of interspersed repeats

22、 is 60% higher in the Z chromosome than in chicken autosomes (Supplementary Figure 1, Table 1). Most of these repeats are LINE elements, whose abundance in the Z chromosome is 70% higher than in autosomes (Supplementary Figure 1, Table 1). As a result, the Z chromosome is structurally distinct from

23、the rest of the chicken genome.为了重建和对比鸟类Z和哺乳动物X染色体的进化路线，我们首次完成了鸡Z染色体全部碱基对的测序工作（补充图形1-3）。整个序列总长约8千万碱基对（80Mbp），被分为5个部分，精确到约每一百万碱基对有一个错误。鸡的Z染色体包含约1000个基因（补充表1），平均每1Mbp11个基因。这也使得Z染色体具有比任何鸡常染色体更低的基因密度，乃至比常染色体平均密度每1Mbp25个基因的一半还低（表1）8。相反地是，Z染色体散在重复序列的密度比常染色体高百分之六十（补充图1，表1）。这些重复大多属于长散在核序列，它在Z染色体上的丰度比常染色体高百分

24、之七十（补充图1，表1）。因此，Z染色体在结构上与鸡其它染色体是有显著不同的。The Z chromosomes most prominent feature is a previously unrecognized tandem array of testis-expressed genes, extending over 11 megabases at the distal end of the long arm (Fig. 1, Supplementary Figure 4). This array constitutes nearly 15% of the Z chromosome,

25、onefifth of all chicken segmental duplications, and 1% of the entire chicken genome (Fig. 1A &B)8. This sequence was initially reported as heterochromatin13, but we find three genes are present in each repeat unit, and a smaller flanking array contains a fourth (Fig. 1C & D, Supplementary Fi

26、gure 5, Supplementary Table 2). Together, these four gene families total hundreds of copies and comprise almost one third of the protein-coding genes on the Z chromosome (Fig. 1D, Supplementary Table 2). All four gene families are expressed predominantly in the testis (Fig. 1E). We have termed this

27、massive array of testis-expressed genes the Z amplicon.Z染色体最显著的特征是具有先前未被发现的串联排列的睾丸表达基因，它位于长臂末端，总长超过了11Mbp（图1，补充图4）。这段序列占Z染色体总长的15%，占鸡重复序列的1/15，以及占鸡整个基因组的1%（图1A &B)8。它原先被认为属于异染色质13，但我们在每个重复单元中发现了3个基因，而且在其侧翼的一小段序列中发现了第四个（图1 C & D，补充图5，补充表2）。这四个基因家族共具有数以百计的拷贝数而且包含了Z染色体上近三分之一的蛋白质编码基因（图1D，补充表2），它

28、们主要在睾丸中表达（图1E）。我们把如此大量排列的睾丸表达基因称为Z扩增子。With the finished sequence of the Z chromosome in hand, we set out to test Ohnos hypothesis that the avian Z and mammalian X chromosomes are orthologous. To reconstructand visualize evolutionary relationships between chicken and human chromosomes, we systematica

29、lly plotted the locations of orthologous gene pairs (Supplementary Fig. 6 & 7). We find that none of the 1000 genes on the chicken Z chromosome has an ortholog on the human X chromosome (Fig. 2A & B, Supplementary Table 1). The Z chromosome is orthologous only to portions of human autosomes

30、5, 9, and 18 (Fig. 2A). Contrary to initial reports7, the Z chromosome is not orthologous to human chromosome 8 (Supplementary Fig. 6). In reciprocal fashion, the human X chromosome is orthologous only to portions of chicken autosomes 1 and 4, not to the Z chromosome (Fig. 2B)5. Based on this compre

31、hensive analysis, we conclude that genes that are sex-linked in chickens are autosomal in humans, and vice versa, in broad agreement with earlier comparative mapping experiments6,7.由于拥有了完整的Z染色体基因序列，我们开始检验Ohno关于鸟类Z和哺乳动物X染色体直系同源的假说。为了重建鸡和人染色体形象化的进化关系，我们系统地绘制了同源基因对位置的图谱（补充图6 & 7)。我们发现鸡Z染色体上这近1000个基

32、因没有一个与人X染色体上基因同源（图2A & B，补充表1）。Z染色体只与人常染色体5，9和18号上部分基因同源（图2A）。与最初的报告相反7，Z染色体不与人8号染色体同源（补充图6）。与之相对应的是，人X染色体只与鸡常染色体1号和4号部分同源，而不是与Z染色体（图2B）5。基于综合分析，我们得出如下结论：鸡中与性别有关的基因与人常染色体基因同源，反之亦然，这也与早先的对比作图实验结果相符合6，7。Although the Z and X chromosomes show no signs of orthology, it is conceivable that they were r

33、ecruited from different portions of a proto sex chromosome in the common ancestor of birds and mammals11. Some investigators have raised this possibility based on comparative gene mapping in the platypus11. However, the platypus does not form an outgroup to birds and mammals, and cannot resolve whic

34、h is the ancestral state: a platypus like linkage of Z-orthologous genes and X-orthologous genes, or the separation we observe in chicken and human. Others have attempted to resolve this question with comparisons to an outgroup genome that is far from complete12. Instead, we compared the Z and X chr

35、omosomes to the genomes of the four closest out-group species whose genomes are sequenced and assembled. Each species represents a different order of teleost fish, which diverged from land vertebrates over 450 million years ago14. After they diverged from birds and mammals, but before they diverged

36、from each other, these fish species experienced a whole genome duplication, complicating the identification of 1:1 orthologs14. Nevertheless, we observe that most orthologs of Z and X-linked genes occupy separate portions of each fish genome (Supplementary Fig. 811). For example, three-spine stickle

37、back linkage groups 13 and 14 carry the bulk of Z-orthologous genes, while X-orthologous genes mostly reside on stickleback linkage groups 1, 4, 7, and 16 (Fig. 2C). Since we observe that Z orthologous genes are separated from X-orthologous genes in birds, mammals, and each of these four fish, we co

38、nclude that the Z and the X chromosomes have evolved independentlyfrom distinct portions of the ancestral vertebrate genome.尽管并没有迹象表明Z和X染色体同源，但它们仍然可能是起源于鸟类和哺乳动物共同祖先的原始性染色体的不同部分11。一些研究者基于鸭嘴兽的对比基因图谱提出了该假说。但鸭嘴兽并没有构成鸟类和哺乳动物的一个外类群，而且鸭嘴兽中Z同源基因和X同源基因的组合体与我们在鸡和人中观察到的分开部分，哪一个处于祖先地位这一问题并没有得到解决。其他人也试图通过将它们与一个并

39、不完整的外类群基因组进行对比来解决这个问题12。而我们则将Z和X染色体与最靠近的四个外类群物种基因组进行比对，这四个物种的基因组都是已经测序和装配好的。每个物种代表了一个不同目的硬骨鱼，它们都是在4.5亿年前就与陆生脊椎动物分离14。在它们与鸟类及哺乳动物分离后，且在它们之间相互分离前这段时间，这些鱼类经历了一次全基因组的加倍。然而，我们观察到大多数Z和与X相关基因占据着每个鱼基因组不同部分，且这些部分相互分开（补充图8-11）。例如，三棘鱼13和14号染色体上的连锁群携带着大量Z染色体的同源基因，而X染色体的同源基因则大多数存在于1，4，7，和16号染色体的连锁群上（图2C）。我们观察到Z染

40、色体同源基因和在鸟，哺乳动物，四种鱼中的X染色体同源基因是分离的，因此得出结论，Z和X染色体是从脊椎动物祖先染色体组的不同部分独立进化而来的。Although we rejected the hypothesis that the avian Z and mammalian X chromosomes share a common origin, we discovered that the chicken Z and human X chromosomes share common features. Like the chicken Z chromosome, the human X ch

41、romosome has a low gene density; there are half as many genes per megabase on the X chromosome as on the average human autosome (Fig. 3A, Table 1A)5. Other investigators have observed that low gene density is often associated with increased interspersed repeat content, specifically LINEs5,15. We als

42、o observe this association on the Z and X chromosomes (Supplementary Fig. 1, Table 1A).尽管我们否认了鸟类Z和哺乳动物X染色体具有相同起源的假说，但我们发现鸡的Z和人的X染色体具有一些共同的特征。与鸡Z染色体很像的是，人类X染色体也有一个较低的基因密度，大约只有人类常染色体平均基因密度的一半（图3A，表1A）5。其他研究者观察到低基因密度往往伴随着更多的散在重复序列，尤其是长散在核序列（LINEs）5，15。我们也在Z和X染色体上观察到了它们的这一联系（补充图1，表1A）。Two scenarios coul

43、d account for these features of the chicken Z and human X chromosomes. Either the Z and X chromosomes arose from autosomes pre-adapted for the role of sex chromosomes, or they arose from ordinary autosomes that convergently evolved into specialized sex chromosomes. If the Z and X chromosomes arose f

44、rom pre-adapted autosomes, then the structural features shared by the Z and X chromosomes should also be found on the orthologous autosomal regions. We tested this theory by comparing each sex chromosome to the orthologous autosomes in the other species (Fig. 2, Table 1, Supplementary Tables 3 &

45、 4). As a group, the autosomal regions that correspond to the Z and the X chromosomes are typical of their respective genomes (Table 1B). Although these regions show a slight deficit in gene density relative to the average within their respective genomes, the difference is too small to account for t

46、he extremely low gene density of the Z and X chromosomes. Because the orthologous autosomes in the other species do not share the structural features common to the Z and X chromosomes, we infer that these convergent features arose during the process of sex chromosome evolution, and not before.鸡Z和人类X

47、染色体的这一特点的形成有两种可能性。Z和X染色体可能起源于已经具有性染色体功能的常染色，也可能起源于普通的常染色体，后来趋同进化为性染色体。如果它们是起源于已经改变的常染色体，那么它们所共有的结构上的特点也应该可以在与它们同源的常染色体区域上被找到。我们通过比较每个性染色体与它们在其他生物中的同源常染色体来检验这个理论（图2，表1，补充表3 & 4）。总的来说，与Z和X染色体对应的常染色体区域具有它们各自基因组的特点（表1B）。尽管这些区域的基因密度与它们各自基因组中的平均水平相比略显不足，但如果考虑到Z和X染色体上极低的基因密度，这个差距还是太小了。由于其它物种中同源染色体并不具有与

48、Z和X染色体相同的结构特点，我们推测这些趋同的特点形成于性染色体进化的过程中，而不是之前。To explain the paucity of genes on the Z and X chromosomes, we looked for evidence that both chromosomes lost genes during sex chromosome evolution. Instead, we observed that both the Z and the X chromosomes gained protein-coding genes. We compared the ge

49、ne content of the Z and the X chromosomes to the orthologous autosomes from the other species as a surrogate for the ancestral gene content of the Z and X chromosomes (Fig. 2,Fig. 3B, Table 1, Supplementary Tables 3 & 4). We found that only a few dozen genes present on the orthologous autosomes

50、are absent from the Z and X chromosomes (Fig. 3B).In contrast, hundreds of genes present on the Z and X chromosomes are absent from the orthologous autosomes (Fig. 3B). We conclude that both the Z and X chromosomes experienced substantial net gene gain.为了解释Z和X染色体上基因的缺乏，我们试图寻找它们在性染色体进化过程中丢失基因的证据。但相反的

51、是，我们观察到Z和X染色体都在进化过程中获得了蛋白质编码基因。我们将其它物种中Z和X染色体的同源常染色体上的基因作为Z和X染色体祖先基因的替代，并将Z和X染色体基因与其对比（图2，图3B，表1，补充表3 & 4）。我们发现只有很少一部分出现在同源常染色体上的基因在Z和X染色体上缺失了（图3B）。与此相反，数百个出现在Z和X染色体上的基因没有在其同源常染色体上找到（图3B）。我们推断，Z和X染色体都经历了净获得大量基因的过程。The majority of genes gained by the Z and X chromosomes are members of multi-copy

52、families (Fig. 3B, Supplementary Tables 3 & 4). On the chicken Z chromosome, these are the genes of the Z amplicon. The human X chromosome has gained thirteen different cancer-testis antigen gene families5. All of the Z amplicon genes are expressed predominantly in testis (Fig. 1E), as are the c

53、ancer-testis antigen genes of the human X chromosome16. The addition of these multi-copy gene families has biased the Z and X chromosomes toward testis-expressed genes (Fig. 3C). Both the Z and the X chromosomes have an elevated proportion of genes expressed in testis tissue compared to autosomes as

54、 measured by the number of genes with a testis EST in Unigene17 datasets (Fig. 3C). However, when multi-copy genes are removed, the remaining conserved single-copy genes show no bias (Fig. 3C). Others have observed a bias towards sex and reproduction related genes on the human X chromosome18. Our co

55、mparison suggests that the Z chromosome shares this bias. This bias was not a feature of the autosomes that gave rise to the sex chromosomes of birds and mammals; it arose by gene acquisition and amplification during sex chromosome evolution in each lineage.Z和X染色体获得的基因大多属于多拷贝基因（图3B，补充表3 & 4）。在鸡Z

56、染色体上，它们属于Z染色体基因的扩增子。人类X染色体已经获得了13个不同睾丸癌抗原基因家族5。所有Z染色体的扩增子基因主要在睾丸中表达（图1E），人类X染色体的睾丸癌抗原基因也是如此16。这些额外的多拷贝基因使得Z和X染色体偏向于睾丸表达基因图3C）。用具有非重复序列数据集17中睾丸表达序列标签的基因的数目来衡量，Z和X染色体与常染色体相比，其睾丸组织表达基因比例有了明显提升（图3C）。但是，当除去多拷贝基因，剩余的保守单拷贝基因则在Z和X染色体与常染色体中没有表现出偏差（图3C）。其他人也已经观察到对于性别的偏好和人X染色体上相关基因的扩增18。我们的对比显示Z染色体也存在同样的偏好。这样的

57、偏好并不是后来进化为鸟类和哺乳类性染色体的常染色体的特点，而是它们在各自独立的进化过程中基因的获得和扩增导致的。In light of this convergent gene gain, we looked for factors other than gene loss that could account for the low gene density of the Z and X chromosomes. Low gene density could result from Z-linked and X-linked genes that are larger than those

58、on autosomes, resulting in fewer genes in the same amount of sequence. However, we find that genes on both the Z and X chromosomes are smaller, on average, than autosomal genes (Table 1A). The only remaining explanation for the unusually low gene density of the Z and X chromosomes is a massive expan

59、sion of non-coding intergenic sequences which spread the genes further apart. We estimate that intergenic regions were expanded by about 40 Mb in the case of the Z chromosome and 80 Mb in the case of the X chromosome nearly half the present lengths of these chromosomes. No single class of non-coding sequence can account for this change, but the two-fold enrichment for LINEs on both the Z and X chromosomes (Table 1A) suggests that the doubling of intergenic sequence may have been driven by recurrent insertion and