34 research outputs found
Deficiency in mouse Y chromosome long arm gene complement is associated with sperm DNA damage
Deficiency in the Multicopy Sycp3-Like X-Linked Genes Slx and Slxl1 Causes Major Defects in Spermatid Differentiation
Slx and Slxl1 are genes present in multiple copies on the mouse X chromosome. Using transgenically-delivered small interfering RNAs to disrupt their function, we show that Slx and Slxl1 are important for normal sperm differentiation and male fertility
The role of sperm DNA damage in the origin of infertility associated with Y chromosome long arm deletions in the mouse model
Ph.D. University of Hawaii at Manoa 2013.Includes bibliographical references.In mouse and man Y chromosome deletions are frequently associated with spermatogenic defects. Mice with severe non-pairing Y chromosome long arm (NPYq) deficiencies are infertile in vivo and in vitro. We have previously shown that sperm from these males, although having grossly malformed heads, were able to fertilize oocytes via intracytoplasmic sperm injections (ICSI) and yield live offspring. However, in continuing ICSI trials we noted a reduced efficiency when cryopreserved sperm were used and with epididymal sperm as compared to testicular sperm. Our initial study tested if NPYq deficiency is associated with sperm DNA damage-a known cause of poor ICSI results. We observed that epididymal sperm from mice with severe NPYq deficiency are impaired in oocyte activation ability, and have an increased incidence of oocyte arrest and paternal chromosome breaks. Comet assays revealed increased DNA damage in both epididymal and testicular sperm, and transmission electron microscopy showed sperm having impaired membrane integrity and abnormal chromatin condensation. We therefore concluded that the increased DNA damage associated with NPYq deficiency might be a consequence of disturbed chromatin remodeling taking place during spermiogenesis.
There are four distinct multi-copy genes found in the long arm of the Y chromosome. One of them, Sly, is known to control the expression of sex chromosome genes after meiosis; Sly deficiency results in a remarkable upregulation of sex chromosome genes. Sly deficiency has also been shown to be the underlying cause of sperm head anomalies and infertility associated with NPYq gene loss. We therefore hypothesized that Sly is our target gene. To test this, we examined mice with transgenically (RNAi) silenced Sly. Our analysis of Sly-deficient mice demonstrated similar 'sperm DNA damage' phenotype. This confirmed that lack of Sly is responsible for the sperm DNA damage/chromatin packaging defects observed in mice with NPYq deletions.
This project provides the first evidence of DNA damage in sperm from mice with NPYq deficiencies and that the multi-copy NPYq-encoded Sly gene plays a key role in processes regulating chromatin remodeling and thus maintaining DNA integrity in sperm
Mouse Y-Encoded Transcription Factor Zfy2 Is Essential for Sperm Formation and Function in Assisted Fertilization.
Spermatogenesis is a key developmental process allowing for a formation of a mature male gamete. During its final phase, spermiogenesis, haploid round spermatids undergo cellular differentiation into spermatozoa, which involves extensive restructuring of cell morphology, DNA, and epigenome. Using mouse models with abrogated Y chromosome gene complements and Y-derived transgene we identified Y chromosome encoded Zfy2 as the gene responsible for sperm formation and function. In the presence of a Zfy2 transgene, mice lacking the Y chromosome and transgenic for two other Y-derived genes, Sry driving sex determination and Eif2s3y initiating spermatogenesis, are capable of producing sperm which when injected into the oocytes yield live offspring. Therefore, only three Y chromosome genes, Sry, Eif2s3y and Zfy2, constitute the minimum Y chromosome complement compatible with successful intracytoplasmic sperm injection in the mouse
Progeny genotype frequencies.
<p>Frequency of the offspring genotypes obtained after assisted reproduction with X<sup><i>E</i>,<i>Z2</i></sup>Y*<sup>X</sup><i>Sry</i> males. Four predominant genotypes (grey bars) and one rare genotype derived from untypical sex chromosome segregation (black bar on the right side of the dashed line) were observed. These genotypes are expected from X<sup><i>E</i>,<i>Z2</i></sup>Y*<sup>X</sup><i>Sry</i> males. Number of genotyped progeny was 49 for ROSI and 18 for ICSI. See also <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1005476#pgen.1005476.s007" target="_blank">S2 Table</a>.</p
Mouse X and Y chromosomes, variant sex chromosomes, and mouse genotypes relevant to this study.
<p>(A) The mouse Y chromosome contains ~90 Mb of male specific DNA and ~0.7 Mb constituting the pseudoautosomal region (PAR) situated at the end of the long arm. The PAR is the region of homology with the X that mediates pairing and recombination between the X and Y in normal males. The remaining non-pairing male specific part of Y (NPY) contains several genes and gene families. On the short arm (NPYp), there are single<b>-</b>copy genes: <i>Prssly</i>, <i>Teyorf1</i>, <i>Uba1y</i>, <i>Smcy/Kdm5d</i>, <i>Eif2s3y</i>, <i>Uty</i>, <i>Dby/Ddx3y</i>, <i>Usp9y</i>, <i>Sry</i>, duplicated gene <i>Zfy</i> (<i>Zfy1</i> and <i>2</i>), duplicated gene <i>H2al2y</i>, and a multi-copy gene <i>Rbmy</i>. The non-pairing region of the long arm (NPYq), representing ~90% of all NPY, contains mostly repetitive sequences, and encodes multiple copies of 5 distinct genes that are expressed in spermatids: <i>Ssty1</i> and <i>Ssty2</i>, <i>Sly</i>, <i>Srsy</i>, <i>Rbm31y</i> [<a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1005476#pgen.1005476.ref006" target="_blank">6</a>]. Y*<sup>X</sup> is an X chromosome derivative encoding PAR, X centromere and near centromeric region. <i>Sxr</i><sup>a</sup> is a sex reversal variant Tp(Y)1Ct<sup><i>Sxr-a</i></sup> encoding almost intact NPYp complement but with <i>Rbmy</i> gene family reduced. <i>Sxr</i><sup>b</sup> <i>is a Sxr</i><sup>a</sup> derivative with a 1.3 Mb deletion that has removed the majority of the NPYp gene complement and created a <i>Zfy2/1</i> fusion gene. (B) The mice used in this study and their Y chromosome contribution. The X chromosome located <i>Eif2s3y</i> and autosomally located <i>Sry</i> transgenes, are shown in light blue frames. The <i>Zfy2</i> transgene, shown in brown frame, is located on the X chromosome in the <i>Hrpt</i> locus in close proximity to the <i>Eif2s3y</i> transgene. The genotype designations without the <i>Zfy2</i> transgene are shown above the diagrammatic representation of sex chromosomes and with the <i>Zfy2</i> transgene below them (brown font). <i>Sxr</i><sup>a</sup> and <i>Sxr</i><sup>b</sup> gene content is shown in A. n/a = mice with transgenic <i>Zfy2</i> addition were either not produced or not examined in this study.</p
Sperm headshape analysis.
<p>(A) The distribution of specific headshape defect categories among testicular sperm from X<sup><i>E</i>,<i>Z2</i></sup>Y*<sup>X</sup><i>Sry</i> (n = 41 sperm from 3 males), X<sup><i>E</i></sup><i>Sxr</i><sup>b</sup>Y*<sup>X</sup> (n = 19 sperm from 2 males), and XY*<sup>X</sup><i>Sxr</i><sup>a</sup> (44 sperm from 3 males) males. (B) The categories of headshape defects. Normal: represents a normal shape of testicular sperm head. A ("dolphin"): sperm head is elongated and has some curvature reminiscent of crescent shape typical for mouse sperm, small hooked tip can be differentiated. B ("mushroom"): sperm head is elongated but the curvature is not present, hint of a hooked tip can sometimes be observed. C ("cupcake"): sperm head is no longer elongated, the caudal side is wider than in A and B categories, and opens up to a wide dorsal side; hint of a hooked tip can be sometimes be seen. D ("egg"): sperm head has an oval shape with no mark of a hooked tip. The head is less elongated than in category B; E ("ball"): sperm head has a round shape with no hint of a hooked tip, is smaller than in all other categories, and is strongly stained indicative of high DNA condensation; F ("drumstick"): sperm head is elongated, has no traces of a hooked tip, is longer and thinner than in category A&B but shorter than in G. G ("club"): sperm head is clearly elongated with no hint of a hooked tip, and very poorly condensed. Scale = 5 μm.</p