Mouse Y-Encoded Transcription Factor Zfy2 Is Essential for Sperm Head Remodelling and Sperm Tail Development

International audienceA previous study indicated that genetic information encoded on the mouse Y chromosome short arm (Yp) is required for efficient completion of the second meiotic division (that generates haploid round spermatids), restructuring of the sperm head, and development of the sperm tail. Using mouse models lacking a Y chromosome but with varying Yp gene complements provided by Yp chromosomal derivatives or transgenes, we recently identified the Y-encoded zinc finger transcription factors Zfy1 and Zfy2 as the Yp genes promoting the second meiotic division. Using the same mouse models we here show that Zfy2 (but not Zfy1) contributes to the restructuring of the sperm head and is required for the development of the sperm tail. The preferential involvement of Zfy2 is consistent with the presence of an additional strong spermatid-specific promotor that has been acquired by this gene. This is further supported by the fact that promotion of sperm morphogenesis is also seen in one of the two markedly Yp gene deficient models in which a Yp deletion has created a Zfy2/1 fusion gene that is driven by the strong Zfy2 spermatid-specific promotor, but encodes a protein almost identical to that encoded by Zfy1. Our results point to there being further genetic information on Yp that also has a role in restructuring the sperm head

Addition of Y*^X to X^<i>E</i>O<i>Sry</i> or X^<i>E</i><i>Sxr</i>^<i>b</i>O models does not improve spermiogenic progression.

<p>Periodic acid Schiff/hematoxylin stained testis sections illustrating the extent of spermiogenic progression. Roman numerals denote estimated tubule stages. <b>(A)</b> In X^<i>E</i>O<i>Sry</i>, the predominantly diploid spermatids do not elongate and the acrosomes (stained dark pink) remain randomly orientated relative to the basement membrane of the tubule (insets in VIII and XII). The spermatid nuclei show signs of pycnosis by stage XII (inset) and the cells have been eliminated by stages II-IV. The abundant round cells at stages II-IV are the new generation of round spermatids with early stages of acrosome development (dark pink ‘acrosomal granules’ in inset) [for more details see Vernet et al 2012]. In X^<i>E</i>Y*^X<i>Sry</i> the block to spermiogenesis remains with elimination of the arrested cells once again evident by stages II-IV (see inset). <b>(B)</b> In X^<i>E</i><i>Sxr</i>^<i>b</i>O, at stage VIII the spermatids have not elongated and they are randomly orientated relative to the tubule basement membrane. However, as previously reported (Vernet et al 2012), spermatid elongation is delayed rather than absent, and is apparent by stage XII. Nuclear condensation is also delayed as it is not evident at stage XII, but many of the elongating spermatids survive to stages II-IV at which point nuclear condensation is now evident. In X^<i>E</i>Y*^X<i>Sxr</i>^<i>b</i> spermatid elongation and nuclear condensation is similarly delayed, but there appear to be fewer elongating spermatids surviving to stages II-IV [note the now evident haploid (h) as well as diploid (d) spermatids]. Scale bar is 40 μm (insets are x3 magnification).</p

Levels of <i>Cypt</i>-dependent transcripts for XY<i>Sxr</i>^<i>b</i>, XY, X^{<i>E</i>,<i>Z2</i>}Y*^X<i>Sry</i> and X^<i>E</i>Y*^X<i>Sxr</i>^<i>b</i>.

<p>The RT-PCR bands quantified are those from the RT-PCR assay in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0145398#pone.0145398.s002" target="_blank">S1 Fig</a>. The transcripts expected for each genotype (n = 2 represented by two bars on the chart) are indicated above each genotype label (2/1 denotes <i>Cypt</i>-dependent <i>Zfy2/1</i> transcripts and 2 denotes <i>Cypt-</i>dependent <i>Zfy2</i> transcripts). <b>(A)</b> The PCR primers for this assay amplified <i>Zfy2/1</i> and <i>Zfy2</i> transcripts; only XY<i>Sxr</i>^<i>b</i> has both transcripts. It can be seen that the level of <i>Cypt-Zfy2/1</i> transcripts in X^<i>E</i>Y*^X<i>Sxr</i>^<i>b</i> is comparable to the level of <i>Cypt-Zfy2</i> transgene transcripts in X^{<i>E</i>,<i>Z2</i>}Y*^X<i>Sry</i>. <b>(B)</b> For the left panel the PCR primers are specific for <i>Zfy2/1</i>; the second and third genotypes lack the <i>Zfy2/1</i> fusion gene so the low level signal represents ‘background’. For the right panel the PCR primers are specific for <i>Zfy2</i>; in this case it is the fourth genotype that lacks <i>Zfy2</i>. The genotypes lacking the Y long arm (Yq-) have substantially higher transcript levels than the genotypes with a complete Y (Yq+).</p

Sperm head and tail morphogenesis in X^{<i>E</i>,<i>Z2</i>}Y*^X<i>Sry</i> males as compared to controls.

<p><b>(A)</b> Electron micrographs of developing sperm heads from 6 week old XY, XY*^X<i>Sxr</i>^<i>a</i> and X^{<i>E</i>,<i>Z2</i>}Y*^X<i>Sry</i> males. Spermatids at round to elongating transitional stage in X^{<i>E</i>,<i>Z2</i>}Y*^X<i>Sry</i> present no apparent ultrastructural defects (bottom left picture). However, vacuoles (V) appear in the cytoplasm of elongating and condensing spermatids. Irregular spread of the acrosomal cap (arrows) distorting the spermatid nuclei is observed in X^{<i>E</i>,<i>Z2</i>}Y*^X<i>Sry</i> and XY*^X<i>Sxr</i>^<i>a</i>. It was again evident that the sperm heads in X^{<i>E</i>,<i>Z2</i>}Y*^X<i>Sry</i> fail to elongate properly (stars). <b>(B)</b> Electron micrographs of sperm tail sections showing a normal 9x2+2 axoneme pattern with a central microtubule pair (p) in addition to the nine outer doublets (d) in all three genotypes. Scale bars: A = 1 μm, B = 0.5 μm (Insets = 3x magnification).</p

Mapping of <i>Prssly</i> and <i>Teyorf1</i> to <i>Sxr</i>^<i>a</i> and <i>Sxr</i>^<i>b</i>.

<p><i>Prssly</i> and <i>Teyorf1</i> map to the Yp-derived <i>Sxr</i>^<i>a</i> chromosomal fragment (here attached distal to the PAR of one X of an XX<i>Sxr</i>^<i>a</i> male). As expected from the known breakpoints for the Δ^<i>Sxr-b</i> deletion, <i>Prssly</i> and <i>Teyorf1</i> are also present in <i>Sxr</i>^<i>b</i>, whereas the <i>Tspy</i> pseudogene is absent.</p

The XO and XY*^X mouse models.

<p><b>(A)</b> XY. The Y short arm (Yp) gene complement of an XY male (represented to scale in the magnified view) comprises nine single copy genes, two duplicated genes and one multi copy gene. The pseudoautosomal region (PAR) located distally on the Y long arm mediates pairing and crossing over with the X PAR during meiosis to generate the XY sex bivalent. Centromeres are represented by a dot on the chromosome. <b>(B–D)</b> The diminishing Yp gene complements for the three XO male mouse models that lack the Y long arm. <b>(B)</b> X<i>Sxr</i>^<i>a</i>O. The Yp-derived <i>Sxr</i>^<i>a</i> attached distal to the X PAR provides an almost complete Yp gene complement. <b>(C)</b> X^{<i>Eif2s3y</i>}<i>Sxr</i>^<i>b</i>O. The <i>Sxr</i>^<i>a</i>-derived deletion variant <i>Sxr</i>^<i>b</i> has a 1.3 Mb deletion (Δ^<i>Sxr-b</i>) removing 6 single copy genes and creating a <i>Zfy2/1</i> fusion gene spanning the deletion breakpoint (†). The deleted gene <i>Eif2s3y</i> is necessary for normal spermatogonial proliferation, so an X-located <i>Eif2s3y</i> transgene has been added. <b>(D)</b> X^{<i>Eif2s3y</i>}OSry. This model has only two Yp genes—the testis determinant <i>Sry</i> provided as an autosomally located transgene and the spermatogonial proliferation factor <i>Eif2s3y</i> provided as the X-located transgene. E. Y*^X. This mini sex-chromosome is an X chromosome with a deletion from just proximal to <i>Amelx</i> to within the DXHXF34 repeat adjacent to the X centromere († marks the deletion breakpoint). This X chromosome derivative has a complete PAR that can pair and crossover with the PAR of X<i>Sxr</i>^<i>a</i>, X<i>Sxr</i>^<i>b</i> or X to form a ‘minimal sex bivalent’. Scale bar for magnified views is 150 kb.</p

Sequence family variant loss from the AZFc interval of the human Y chromosome, but not gene copy loss, is strongly associated with male infertility

Background: Complete deletion of the complete AZFc interval of the Y chromosome is the most common known genetic cause of human male infertility. Two partial AZFc deletions (gr/gr and b1/b3) that remove some copies of all AZFc genes have recently been identified in infertile and fertile populations, and an association study indicates that the resulting gene dose reduction represents a risk factor for spermatogenic failure. Methods: To determine the incidence of various partial AZFc deletions and their effect on fertility, we combined quantitative and qualitative analyses of the AZFc interval at the DAZ and CDY1 loci in 300 infertile men and 399 control men. Results: We detected 34 partial AZFc deletions (32 gr/gr deletions), arising from at least 19 independent deletion events, and found gr/gr deletion in 6% of infertile and 3.5% of control men (p>0.05). Our data provide evidence for two large AZFc inversion polymorphisms, and for relative hot and cold spots of unequal crossing over within the blocks of homology that mediate gr/gr deletion. Using SFVs (sequence family variants), we discriminate DAZ1/2, DAZ3/4, CDY1a (proximal), and CDY1b (distal) and define four types of DAZ-CDY1 gr/gr deletion. Conclusions: The only deletion type to show an association with infertility was DAZ3/4-CDY1a (p = 0.042), suggesting that most gr/gr deletions are neutral variants. We see a stronger association, however, between loss of the CDY1a SFV and infertility (p = 0.002). Thus, loss of this SFV through deletion or gene conversion could be a major risk factor for male infertility

Genetic diversity on the Comoros Islands shows early seafaring as major determinant of human biocultural evolution in the Western Indian Ocean

The Comoros Islands are situated off the coast of East Africa, at the northern entrance of the channel of Mozambique. Contemporary Comoros society displays linguistic, cultural and religious features that are indicators of interactions between African, Middle Eastern and Southeast Asian (SEA) populations. Influences came from the north, brought by the Arab and Persian traders whose maritime routes extended to Madagascar by 700–900 AD. Influences also came from the Far East, with the long-distance colonisation by Austronesian seafarers that reached Madagascar 1500 years ago. Indeed, strong genetic evidence for a SEA, but not a Middle Eastern, contribution has been found on Madagascar, but no genetic trace of either migration has been shown to exist in mainland Africa. Studying genetic diversity on the Comoros Islands could therefore provide new insights into human movement in the Indian Ocean. Here, we describe Y chromosomal and mitochondrial genetic variation in 577 Comorian islanders. We have defined 28 Y chromosomal and 9 mitochondrial lineages. We show the Comoros population to be a genetic mosaic, the result of tripartite gene flow from Africa, the Middle East and Southeast Asia. A distinctive profile of African haplogroups, shared with Madagascar, may be characteristic of coastal sub-Saharan East Africa. Finally, the absence of any maternal contribution from Western Eurasia strongly implicates male-dominated trade and religion as the drivers of gene flow from the North. The Comoros provides a first view of the genetic makeup of coastal East Africa