A. Touré

Patterns, mechanisms, and functions of translation regulation in mammalian spermatogenic cells

Translational regulation is a fundamental aspect of the atypical patterns of gene expression in mammalian meiotic and haploid spermatogenic cells. Every mRNA is at least partially translationally repressed in meiotic and haploid spermatogenic cells, but the extent of repression of individual mRNA species is regulated individually and varies greatly. Many mRNA species, such as protamine mRNAs, are stored in translationally repressed free-mRNPs in early haploid cells and translated actively in late haploid cells. However, translation does not regulate developmental expression of all mRNAs. Some mRNAs appear to be partially repressed for the entire period that the mRNA is expressed in meiotic and haploid cells, while other mRNAs, some of which are expressed at high levels, are almost totally inactivated in free-mRNPs and/or generate little or no protein. This distinctive phenomenon can be explained by the hypothesis that translational repression is used to prevent the potentionally deleterious effects of overproduction of proteins encoded by overexpressed mRNAs. Translational regulation also appears to be frequently altered by the widespread usage of alternative transcription start sites in spermatogenic cells. Many ubiquitously expressed genes generate novel transcripts in somatic spermatogenic cells containing elements, uORFs and secondary structure that are inhibitory to mRNA translation, while the ribosomal proten L32 mRNA lacks a repressive element that is present in somatic cells. Very little is known about the mechanisms that regulate mRNA translation in spermatogenic cells, largely because few labs have utilized in vivo genetic approaches, although there have been important insights into the repression and activation of protamine 1 mRNA, and the role of Y-box proteins and poly(A) lengthening in mRNA-specific translational activation mediated by the cytoplasmic poly(A) element binding protein and a testis-specific isoform of poly(A) polymerase. A very large literature by evolutionary biologists suggests that the atypical patterns of gene expression in spermatogenic cells are the consequence of the powerful and unusual selective pressures on male reproductive success.

Specific arrests of spermatogenesis in genetically modified and mutant mice.

In naturally occurring mutant mice but also in mice genetically modified for the study of other organs, relatively often a spermatogenic arrest is seen. In a number of cases the arrests appear to be very specific causing apoptosis of germ cells at a particular step in their development, while before this step cells progress normally. These steps include: proliferation/migration of primordial germ cells, the production of differentiating spermatogonia by gonocytes, the regulation of stem cell renewal/differentiation, the differentiation of Aal into A1 spermatogonia, proliferation of A1-A4 spermatogonia, germ cell density regulation, start of meiosis, epithelial stage IV checkpoint of pachytene spermatocytes, the first meiotic division, the formation of the acrosomic vesicle in spermatids and several other steps in spermatid development. In addition, there are many mice that have not been studied in enough detail for a proper categorization. In this review an overview is given of the various mutations and genetically modified mice showing a direct effect on specific spermatogenic cell types. In addition, the relevance of these models to our understanding of the spermatogenic process is discussed.

Expression Analysis of the Mouse Multi-Copy X-Linked Gene Xlr-Related, Meiosis-Regulated (Xmr), Reveals That Xmr Encodes a Spermatid-Expressed Cytoplasmic Protein, SLX/XMR

Abstract The mouse multi-copy X-linked gene Xlr-related, meiosis-regulated (Xmr/Slx) has previously been described as encoding a testis-specific nuclear protein expressed during male meiotic prophase, and during which it becomes concentrated in the inactive X and Y chromatin domain. These conclusions were based on Western blot and immunolocalization analysis using an antibody raised against a related lymphocyte protein, XLR; however, our recently published RNA in situ for Xmr revealed that transcripts are predominantly or exclusively postmeiotic, and this is supported by a growing body of microarray data. This led us to reanalyze the expression of Xmr, both at the RNA level by RT-PCR and by RNA fluorescence in situ hybridization, and at the protein level by using antibodies raised against XMR that do not recognize XLR. In agreement with our previous RNA in situ data, our further transcription analysis showed almost exclusive expression in spermatids, and Western blot and immunostaining with the XMR antibodies showed that the protein is cytoplasmic and restricted to spermatids. Furthermore, the previously used XLR antibody was shown not to cross-react with XMR, and it is suggested that the meiotically expressed nuclear protein recognized by this antibody is another member of the complex Xlr superfamily. As a result of these findings, the gene previously known as Xmr is now officially know as Slx, Sycp3-like, X-linked.

A protein encoded by a member of the multicopy Ssty gene family located on the long arm of the mouse Y chromosome is expressed during sperm development

Multicopy Y-chromosomal genes in human and mouse have been postulated to play a role in spermatogenesis. The mouse Y long arm (Yq) carries hundreds of supposedly intronless copies of Ssty, for which no protein has hitherto been identified; mice lacking Yq are sterile with grossly abnormal sperm. We have now identified an Ssty-encoded protein (Ssty1) that is expressed in spermatids. The protein is absent from spermatids of mice that lack Yq, but is not reduced in mice with a two-thirds reduction of Ssty copies, implying that most do not produce this protein. Furthermore, no protein was produced by a strongly transcribed intronless Ssty transgene, raising doubts as to the protein-encoding potential of these intronless genes. We have now identified an intron-containing copy that is also present in multiple copies on Yq. One or more intron-containing copies are retained in the Ssty-deficient mice and may be the source of the Ssty1 protein.

The androgen receptor in spermatogenesis

Androgens are steroid hormones that are necessary for normal male phenotype expression, including the outward development of secondary sex characteristics as well as the initiation and maintenance of spermatogenesis. Many physiological actions of androgens are mediated by the androgen receptor (AR), a member of the nuclear receptor superfamily. AR functions as a ligand-dependent transcription factor, regulating expression of an array of target genes that are important in male pubertal development and fertility. In this review, the expression and necessity of AR in specific testicular cell types that are important in spermatogenesis will be discussed, and recent information obtained through the study of complete and cell type-specific AR null mouse models will be presented.

Meiotic recombination and spermatogenic impairment in Mus musculus domesticus carrying multiple simple Robertsonian translocations

We quantitatively analyzed the spermatogenic process, including evaluation of seminiferous tubules with defective cycles, rates of germ cell death and sperm morphology, in adult male mice with standard telocentric chromosomes (2n = 40, CD1 strain), homozygous (2n = 24, Mil II population) and heterozygous (2n = 24 × 40) for Robertsonian (Rb) rearrangements. The animals were analyzed at three different ages: three, five and seven months after birth. The number and position of crossover events were also determined by chiasmata counting and immunostaining with an antibody against mouse MLH1 protein. Our analysis of spermatogenesis confirms the impairment of the spermatogenic process in multiple simple heterozygotes due to both germ cell and abnormal sperm morphology. The detrimental effects exerted by Rb heterozygosities were found to be at least partially buffered with time: the frequency of defective tubules was lower and germ cell survival and sperm morphology better in 7-month-old animals than in the 3- and 5-month-old mice. While there are previously published data on germ cell death in multiple simple heterozygotes, this is the first report of a partial rescue of spermatogenesis with time. The mean frequency of MLH1 foci was lower in Rb homozygous and heterozygous mice than in mice carrying all telocentric chromosomes. The lower number of foci in Rb mice can be ascribed to a decrease in the number of multiple chiasmata and the maintenance of single chiasmata preferentially located in the terminal region of both the telocentric and metacentric chromosomes.

Septins at the annulus of mammalian sperm.

Abstract The annulus is an electron-dense ring structure connecting the midpiece and the principal piece of the mammalian sperm flagellum. Proteins from the septin family have been shown to localize to the annulus. A septin complex is assembled early in spermiogenesis with the cochaperone DNAJB13 and, in mature sperm, associates with Testis Anion Transporter 1; SLC26A8 (Tat1), a transmembrane protein of the SLC26 family. Studies in mice have shown that the annulus acts as a barrier to protein diffusion and controls correct organization of the midpiece. Consistent with these findings, absence of the annulus is associated with flagellum differentiation defects and asthenozoospermia in humans.

Pathways of post-transcriptional gene regulation in mammalian germ cell development

Male germ cell development is orchestrated by complex and disparate patterns of gene expression operating in different cell types. The mechanisms of gene expression underlying these have been dissected in the mouse because of its readily available genetics. These analyses have shown that as well as the traditional transcriptional mechanisms, post-transcriptional regulatory pathways of gene expression are essential for mouse spermatogenesis. Proteins essential for germ cell development have been identified which operate at different points throughout the life cycle of RNA from pre-mRNA splicing to translation and RNA decay in the cytoplasm. Recent data suggests that these post-transcriptional pathways respond to environmental cues via signalling pathways.

Insights into regulation of the mammalian cell cycle from studies on spermatogenesis using genetic approaches in animal models

The genetic hierarchy controlling mitosis and especially meiosis during gamete formation is not well understood, even in less complicated systems such as the yeasts. Meiotic divisions are obviously restricted to germ line cells and as such likely require mechanisms of cell cycle control that do not function and may not exist in somatic cells. While male and female germ cells have stages of cell cycle regulation in common, the timing of these events and the stage of development at which these events occur differ in the two sexes. Understanding the genetic program controlling the mitotic and meiotic divisions of the germ line represents a unique opportunity for providing insight into cell cycle control in vivo. Elucidating the key control points and proteins may also enhance our understanding of the etiology of infertility and provide new directions for contraception.

The role of N-glycans in spermatogenesis

Many proteins, in particular those in the plasma membranes, are glycosylated with carbohydrates, which are grouped into O-glycans and N-glycans. O-glycans are synthesized step by step by glycosyltransferases, whereas N-glycans are synthesized by en-bloc transfer of the so-called high-mannose-type oligosachharide from lipid-linked precursor to polypeptide. The high-mannose-type N-glycans are then modified by processing α-mannosidases. Alpha-mannosidase IIx (MX) was identified as the gene product of processing α-mannosidase II (MII)-related gene. MX apparently plays subsidiary role for MII in many cell types, as N-glycan patterns of MX null mouse tissues are not altered significantly. Surprisingly MX null male mice are infertile due to a failure of spermatogenesis. This review provides a brief overview of the in vivo role of N-glycans which are revealed by the gene knockout mouse approach, and introduce our studies on the MX gene knockout mouse. The MX gene knockout experiments unveiled a novel function of a specific N-glycan, which is N-acetylglucosamine-terminated and has a fucosylated triantennary structure, in the adhesion between germ cells and Sertoli cells. The study of MX is a good example of how the in vivo roles of an apparently redundant gene product are determined by the gene knockout approach.