Simian Y Chromosomes: species-specific rearrangements of DAZ, RBM, and TSPY versus contiguity of PAR and SRY

The three human male specific expressed gene families DAZ, RBM, and TSPY are known to be repetitively clustered in the Y-specific region of the human Y Chromosome (Chr). RBM and TSPY are Y-specifically conserved in simians, whereas DAZ cannot be detected on the Y chromosomes of New World monkeys. The proximity of SRY to the pseudoautosomal region (PAR) is highly conserved and thus most effectively stabilizes the pseudoautosomal boundary on the Y (PABY) in simians. In contrast, the non-recombining part of the Y Chrs, including DAZ, RBM, and TSPY, was exposed to species-specific amplifications, diversifications, and rearrangements. Evolutionary fast fixation of any of these variations was possible as long as they did not interfere with male fertility. Received: 18 August 1997 / Accepted: 13 November 1997     

Identification in chromosome 8q11 of a region of homology with the g1 amplicon of the Y chromosome and functional analysis of the BEYLA gene

The male-specific region (MSY) of the Y chromosome contains genes involved mainly in male sex determination and in spermatogenesis. The majority of genes involved in male fertility are localized in multiple copies in the long arm of the Y chromosome, within specific regions defined as "ampliconic regions." It has been suggested that these genes derived from X-linked or autosomal ancestors during evolution, providing a benefit for male fertility when transposed onto the Y chromosome. So far, the autosomal origin has been demonstrated only for two MSY genes, DAZ and CDY. In the present study we report on the identification within chromosome 8q11.2 of a region homologous to the g amplicon, containing the VCY2 (approved gene symbol BPY2), TTTY4, and TTTY17 genes. A search for ancestor genes within the 8q11.2 region allowed us to identify a gene named BEYLA and to characterize the genomic organization and the expression patterns of this gene.     

Heterogeneity of pericentric inversions of the human y chromosome

Pericentric inversions of the human Y chromosome (inv(Y)) are the result of breakpoints in Yp and Yq. Whether these breakpoints occur recurrently on specific hotspots or appear at different locations along the repeat structure of the human Y chromosome is an open question. Employing FISH for a better definition and refinement of the inversion breakpoints in 9 cases of inv(Y) chromosomes, with seemingly unvarying metacentric appearance after banding analysis, unequivocally resulted in heterogeneity of the pericentric inversions of the human Y chromosome. While in all 9 inv(Y) cases the inversion breakpoints in the short arm fall in a gene-poor region of X-transposed sequences proximal to PAR1 and SRY in Yp11.2, there are clearly 3 different inversion breakpoints in the long arm. Inv(Y)-types I and II are familial cases showing inversion breakpoints that map in Yq11.23 or in Yq11.223, outside the ampliconic fertility gene cluster of DAZ and CDY in AZFc. Inv(Y)-type III shows an inversion breakpoint in Yq11.223 that splits the DAZ and CDY fertility gene-cluster in AZFc. This inversion type is representative of both familial cases and cases with spermatogenetic impairment. In a further familial case of inv(Y), with almost acrocentric morphology, the breakpoints are within the TSPY and RBMY repeat in Yp and within the heterochromatin in Yq. Therefore, the presence of specific inversion breakpoints leading to impaired fertility in certain inv(Y) cases remains an open question.     

The cloning of size-heterogeneous,Y-specific repetitive DNAs and their clinical application

Four Y-specific DNAs of different sizes were isolated by screening a human Y-chromosome gene-library. After determining their structural characteristics, the possibility for their clinical application was examined. The results are as follows: 1. The 4 cloned DNAs had inserts of 3.3, 2.0, 1.9 and 1.4 kb; they were subcloned into plasmid pUC19, and designated pKY-2, pKY-3, pKY-4 and pKY-5, respectively. 2. All four clones hybridized specifically with the Y-specific 3.5 kb DNA but not with female DNA. However, they did not cross-hybridize with the 2.1 kb fragment known to be another Y-specific repetitive DNA of human genomes. 3. The newly cloned probes were applied to the detection of Y-specific DNA in abnormal Y-chromosomes [Turner Syndrome (45, X/46, X, -X, + marker), 46, XY-female, mosaicism (46, XY/47, XYY), 46, XX-male and super male (47, XYY)]. 4. Since the newly cloned DNAs did not hybridize with other eukaryotic genome DNAs such as monkey, rat, mouse, chicken, frog, or fish, their human gene specificities were confirmed.     

47,X,idic(Y),inv dup(Y): a non-mosaic case of a phenotypically normal boy with two different Y isochromosomes and neocentromere formation

A de novo aberrant karyotype with 47 chromosomes including 2 different-sized markers was identified during prenatal diagnosis. Fluorescence in situ hybridization (FISH) with a Y painting probe tagged both marker chromosomes which were supposed to be isochromosomes of the short and the long arm, respectively. A normal boy was born in time who shows normal physical and mental development. To characterize both Y markers in detail, we postnatally FISH-mapped a panel of Y chromosomal probes including SHOX (PAR1), TSPY, DYZ3 (Y centromere), UTY, XKRY, CDY, RBMY, DAZ, DYZ1 (Yq12 heterochromatin), SYBL1 (PAR2), and the human telomeric sequence (TTAGGG)(n). The smaller Y marker turned out to be an isochromosome containing an inverted duplication of the entire short arm, the original Y centromere, and parts of the proximal long arm, including AZFa. The bigger Y marker was an isochromosome of the rest of the Y long arm. Despite a clearly visible primary constriction within one of the DAPI- and DYZ1-positive heterochromatic regions, hybridization of DYZ3 detected no Y-specific alphoid sequences in that constriction. Because of its stable mitotic distribution, a de novo formation of a neocentromere has to be assumed.     

对法医学相关的DYS549、DYS527和DYS459基因座在男性不育症人群中的缺失、重复调查

位于Y染色体无精症因子区域(Azoospermia factor, AZF)的基因座位点DYS549、DYS527和DYS459在法医学鉴定和家系分析中被广泛应用。但是,在男性不育患者中,DYS549、DYS527和DYS459位点很可能会表现出特殊的基因型,对应用Y染色体短串联重复序列(Y chromosome short tandem repeat, Y-STR)进行个体识别的结果产生干扰。因此,文章应用14个Y-STR基因座复合扩增体系和Y染色体AZFc区DAZ、CDY1基因的拷贝数检测等方法,探讨男性不育症中法医学相关的3个Y-STR基因座的异常分型,对个体识别和家系分析中的DNA检验异常结果提供合理的解释。在240例男性非梗阻性无精、严重少精、先天性双侧输精管缺如(CBVAD)患者中,采用改良的多重PCR体系进行AZF区域微缺失的序列标签位点(Sequence tagged sites, STSs)检测,发现AZF微缺失40例(AZFa：2例;AZFb：2例;AZFc：30例;AZFb+c：6例),AZF的总缺失率为16.67%。应用14 Y-STR复合扩增体系对上述AZF微缺失的阳性患者样本进行检测,发现所有AZFb缺失患者存在DYS549等位基因缺失,AZFc缺失患者存在DYS527、DYS459等位基因缺失,AZFb+c缺失患者存在DYS549、DYS527和DYS459等位基因缺失。在AZF微缺失阴性的不育症患者中,通过检测DAZ、CDY1基因拷贝数发现10例AZFc部分复制的患者(1例为先天性输精管缺如,2例非梗阻性无精症,7例严重少精子症),占所调查不育人群的4.17%。男性不育人群AZF区域3个Y-STR基因座多态性会造成等位基因缺失或者重复,这些异常分型是由于临床遗传缺陷造成的而不是实验偏差。阐明Y-STR在男性不育人群中的异质性可以更好地完善Y-STR数据库和解释STR实验结果。     

High-Throughput Screening for Spermatogenesis Candidate Genes in the AZFc Region of the Y Chromosome by Multiplex Real Time PCR Followed by High Resolution Melting Analysis

Microdeletions in the AZF region of the Y chromosome are among the most frequent genetic causes of male infertility, although the specific role of the genes located in this region is not fully understood. AZFa and AZFb deletions impair spermatogenesis since no spermatozoa are found in the testis. Deletions of the AZFc region, despite being the most frequent in azoospermic patients, do not correlate with spermatogenic failure. Therefore, the aim of this work was to develop a screening method to ascertain the presence of the main spermatogenesis candidate genes located in the AZFc region in the light of the identification of those responsible for spermatogenic failure. DAZ, CDY, BPY2, PRY, GOLGA2LY and CSGP4LY genes were selected on the basis of their location in the AZFc region, testis-only expression, and confirmed or predicted protein codification. AMEL and SRY were used as amplification controls. The identification of Real Time PCR products was performed by High Resolution Melting analysis with SYTO 9 as intercalating dye. The herein described method allows a rapid, simple, low-cost, high-throughput screening for deletions of the main AZFc genes in patients with spermatogenic failure. This provides a strategy that would accelerate the identification of spermatogenesis candidate genes in larger populations of patients with non-obstructive idiopathic azoospermia.     

Association of the mouse infertility factor DAZL1 with actively translating polyribosomes

The DAZ (Deleted in AZoospermia) gene family was isolated from a region of the human Y chromosome long arm that is deleted in about 10% of infertile men with idiopathic azoospermia. DAZ and an autosomal DAZ-like gene, DAZL1, are expressed in germ cells only. They encode proteins with an RNA recognition motif and with either a single copy (in DAZL1) or multiple copies (in DAZ) of a DAZ repeat. A role for DAZL1 and DAZ in spermatogenesis is supported by their homology to a Drosophila male infertility protein Boule and by sterility of Dazl1 knock-out mice. The biological function of these proteins remains unknown. We found that DAZL1 and DAZ bound similarly to various RNA homopolymers in vitro. We also used an antibody against the human DAZL1 to determine the subcellular localization of DAZL1 in mouse testis. The sedimentation profiles of DAZL1 in sucrose gradients indicate that DAZL1 is associated with polyribosomes, and further capture of DAZL1 on oligo(dT) beads demonstrates that the association is mediated through the binding of DAZL1 to poly(A) RNA. Our results suggest that DAZL1 is involved in germ-cell specific regulation of mRNA translation.     

Origin of Amerindian Y-chromosomes as inferred by the analysis of six polymorphic markers

We analysed the frequency of six Y-specific polymorphisms in 105 Amerindian males from seven different populations, 42 Caucasian males, and a small number of males of African, Chinese, and Melanesian origin. The combination of three of the six polymorphisms studied produced four different Y-haplogroups. The haplogroup A (non-variant) was the most frequent one. Eighty-five percent of Amerindians showing haplogroup A have the alphoid II (αhII) and the DYS19A Y-specific markers, an association that is found only in 10% of Caucasians and that has not been detected in Asiatics and Africans. Haplogroups C (YAP+) and D (YAP+ plus an A → G transition in the locus DYS271) are of African origin. Four percent of Amerindians and ∼12% of Caucasians showed haplogroup C; ∼1% of Amerindians and ∼2% of Caucasians had haplogroup D. Haplogroup B is characterized by a C → T transition in nucleotide position 373 of the SRY gene domain; this haplogroup is found in Caucasians (∼12%) and Amerindians (∼4%). None of the Amerindians exhibiting the haplogroups B, C, or D show the haplotype αhII/DYS19A. By haplotyping the Alu insert and the DNA region surrounding the insert in YAP+ individuals, we could demonstrate that Amerindian Y chromosomes bearing African markers (haplogroups C and D) are due to recent genetic admixture. Most non-αhII/DYSl9A Amerindian Y-chromosomes in haplogroup A and most cases in haplogroup B are also due to gene flow. We show that haplotype αhII/DYS19A is in linkage disequilibrium with a C → T transition in the locus DYS199. Our results suggest that most Amerindian Y-chromosomes derive from a single paternal lineage characterized by the αhII/DYS19A/DYS199T Amerindian-specific haplotype. The analysis of a larger sample of native American Y-chromosomes will be required in order to confirm or correct this hypothesis. Am J Phys Anthropol 102:79–89, 1997. © 1997 Wiley-Liss, Inc.     

DAZ family proteins exist throughout male germ cell development and transit from nucleus to cytoplasm at meiosis in humans and mice

The human DAZ gene family is expressed in germ cells and consists of a cluster of nearly identical DAZ (deleted in azoospermia) genes on the Y chromosome and an autosomal homolog, DAZL (DAZ-like). Only the autosomal gene is found in mice. Y-chromosome deletions that encompass the DAZ genes are a common cause of spermatogenic failure in men, and autosomal homologs of DAZ are essential for testicular germ cell development in mice and DROSOPHILA: Previous studies have reported that mouse DAZL protein is strictly cytoplasmic and that human DAZ protein is restricted to postmeiotic cells. By contrast, we report here that human DAZ and human and mouse DAZL proteins are present in both the nuclei and cytoplasm of fetal gonocytes and in spermatogonial nuclei. The proteins relocate to the cytoplasm during male meiosis. Further observations using human tissues indicate that, unlike DAZ, human DAZL protein persists in spermatids and even spermatozoa. These results, combined with findings in diverse species, suggest that DAZ family proteins function in multiple cellular compartments at multiple points in male germ cell development. They may act during meiosis and much earlier, when spermatogonial stem cell populations are established.     

A large AZFc deletion removes DAZ3/DAZ4 and nearby genes from men in Y haplogroup N

Deletion of the entire AZFc locus on the human Y chromosome leads to male infertility. The functional roles of the individual gene families mapped to AZFc are, however, still poorly understood, since the analysis of the region is complicated by its repeated structure. We have therefore used single-nucleotide variants (SNVs) across approximately 3 Mb of the AZFc sequence to identify 17 AZFc haplotypes and have examined them for deletion of individual AZFc gene copies. We found five individuals who lacked SNVs from a large segment of DNA containing the DAZ3/DAZ4 and BPY2.2/BPY2.3 gene doublets in distal AZFc. Southern blot analyses showed that the lack of these SNVs was due to deletion of the underlying DNA segment. Typing 118 binary Y markers showed that all five individuals belonged to Y haplogroup N, and 15 of 15 independently ascertained men in haplogroup N carried a similar deletion. Haplogroup N is known to be common and widespread in Europe and Asia, and there is no indication of reduced fertility in men with this Y chromosome. We therefore conclude that a common variant of the human Y chromosome lacks the DAZ3/DAZ4 and BPY2.2/BPY2.3 doublets in distal AZFc and thus that these genes cannot be required for male fertility; the gene content of the AZFc locus is likely to be genetically redundant. Furthermore, the observed deletions cannot be derived from the GenBank reference sequence by a single recombination event; an origin by homologous recombination from such a sequence organization must be preceded by an inversion event. These data confirm the expectation that the human Y chromosome sequence and gene complement may differ substantially between individuals and more variations are to be expected in different Y chromosomal haplogroups.     

Comparative analysis of human masculinity

To study rapidly evolving male specific Y (MSY) genes we retrieved and analyzed nine such genes. VCY, HSFY and RBMY were found to have functional X gametologs, but the rest did not. Using chimpanzee orthologs for XKRY, CDY, HSFY, PRY, and TSPY, the average silent substitution is estimated as 0.017 +/- 0.006/site and the substitution rate is 1.42 x 10(-9)/site/year. Except for VCY, all other loci possess two or more pseudogenes on the Y chromosome. Sequence differences from functional genes show that BPY2, DAZ, XKRY, and RBMY each have one pseudogene for each one that is human specific, while others were generated well before the human-chimpanzee split, by means of duplication, retro-transposition or translocation. Some functional MSY gene duplication of VCY, CDY and HSFY, as well as X-linked VCX and HSFX duplication, occurred in the lineage leading to humans; these duplicates have accumulated nucleotide substitutions that permit their identification.     

Deletion of CDY1b copy of Y chromosome CDY1 gene is a risk factor of male infertility in Tunisian men

The relationship between male infertility and microdeletions in the Y chromosome that remove multiple genes varies among countries and populations. The aim of this study was to investigate the different types of Chromodomain protein, Y-linked 1 (CDY1) gene deletions and their effect on male infertility and spermatogenesis in Tunisian men. A total of 241 infertile men with different spermatogenic impairments and 115 fertile men were included in this study. We determined the prevalence of CDY1a and CDY1b copy deletions by PCR-RFLP using PvuII as restriction endonuclease. Results: Among the 356 Tunisian individuals, 93.25% had the two copies (CDY1a and CDY1b) of CDY gene (91.2% in infertile patients and 97.3% in fertile men). We also found that deletion of CDY1b was significantly more frequent in infertile patients (azoo/oligospermic and normospermic) than in fertile men (7% vs 1.7% respectively; p value = 0.02). However, deletion of CDY1a copy was very rare, and was detected in only one fertile man and four normospermic infertile patients. Our findings showed that deletion of CDY1b copy gene is a significant risk factor for male infertility independent of sperm concentration, whereas deletion of CDY1a gene seems to have no effect on fertility in the Tunisian population.     

Origin of YAP+ lineages of the human Y-chromosome

We screened a total of 841 Y-chromosomes representing 36 human populations of wide geographical distribution for the presence of a Y-specific Alu insert (YAP+ chromosomes). The Alu element was found in 77 cases. We tested 5 biallelic and 8 polyallelic markers in 70 out of the 77 YAP+ chromosomes. We could identify the existence of a hierarchical and chronological structuring of ancestral and derived YAP+ lineages, giving rise to 4 haplogroups, 14 subhaplogroups and 60 haplotypes. Moreover, we propose a monophyletic origin for each one of the YAP+ lineages. Out-of-Africa and out-of-Asia models have been suggested to explain the origin and evolution of ancestral and derived YAP+ elements. We analyze the evidence supporting these two hypotheses, and we conclude that the information available does not allow one to decide between the out-of-Asia or out-of-Africa models.     

DAZ家族新成员BOULE蛋白的结构与功能

BOULE蛋白是2001年发现的DAZ家族的新成员,是人类精子发生过程中减数分裂的关键调控因子. BOULE基因表达的改变或BOULE蛋白的缺乏可引起减数分裂阻滞和精子生成障碍,从而导致无精子症并产生不育. BOULE蛋白的一级结构中含有DAZ家族的特征结构域,包括DAZ重复和RNA结合域（RBM）,因此,将其列为继DAZ、DAZL之后DAZ家族的第3个成员.本文对BOULE的发现过程、结构和定位进行了总结回顾,并重点介绍了其在精子发生减数分裂中的作用及其作用机制.