Analysis of the role of the X/Y homologous gene family Xmr/Sly in murine spermiogenesis

Spermatogenesis is a complex, continuous developmental process that involves the formation of highly differentiated haploid sperm from diploid spermatogonial stem cells. Deletion analysis has established that genes on the Y chromosome are essential for normal sperm production in humans, mice and Drosophila. In mice, deletions of the Y chromosome long arm (Yq) are associated with abnormal sperm head formation, reduced fertilising ability, up-regulation of spermatid-expressed sex-linked genes, and an offspring sex ratio distortion in favour of females. The severity of the testicular phenotypes correlates with the extent of the deletion, with mice lacking the Yq being sterile. It has been suggested that deficiency of a Yq-encoded multicopy genetic element (the ‘spermiogenesis factor’) is responsible for these phenotypes, and one potential candidate gene is Sly, a member of the Xlr superfamily. This family also includes Xmr, an X-linked multicopy gene found to be up-regulated (along with other X- and Y-encoded genes) in the Yq deletion models. To access the candidacy of Sly as the ‘spermiogenesis factor’, the expression of this gene was examined in the testis at the transcript and protein levels. Sly encodes a protein that is expressed in round and early elongating spermatids. SLY interacts with the acrosomal protein DKKL1, suggesting that SLY may be involved in the development or function of the acrosome, a structure that contains digestive enzymes required for fertilisation. Secondly, the role of Xmr in spermatogenesis was investigated. Xmr has been reported to encode a meiotic protein that localises to the transcriptionally silent sex body in pachytene spermatocytes. However, evidence is provided that this meiotic protein is not XMR, with the antibody used in previous studies unable to recognise XMR. Instead, Xmr is predominantly transcribed in spermatids were it encodes a cytoplasmic protein of unknown function. Next, the up-regulation of X- and Y-linked genes in spermatids from the Yq deletion mice was studied in detail. This up-regulation is due, at least in part, to increased gene transcription, and this is accompanied by changes to the epigenetic profile of the sex chromosome and centromeric heterochromatin in round spermatids. In addition, the protein levels of the up-regulated genes are also increased in the testis from Yq deletion mice, and thus the testicular phenotypes exhibited by these mice may be the result of over-expression of one or more X- and Y-encoded proteins, rather than a direct effect of Yq-linked gene deficiency. Finally, mice transgenic for Sly were generated and characterised to determine if loss of this gene alone is responsible for the spermiogenic defects observed in mice with Yq deletions. Xmr transgenic mice were also produced to examine any potential regulatory interaction between Sly and Xmr and investigate if over-expression of Xmr contributes to the phenotypes exhibited by Yq deletion mice