A Nonsense Mutation in TMEM95 Encoding a Nondescript Transmembrane Protein Causes Idiopathic Male Subfertility in Cattle

Abstract

Genetic variants underlying reduced male reproductive performance have been identified in humans and model organisms, most of them compromising semen quality. Occasionally, male fertility is severely compromised although semen analysis remains without any apparent pathological findings (i.e., idiopathic subfertility). Artificial insemination (AI) in most cattle populations requires close examination of all ejaculates before insemination. Although anomalous ejaculates are rejected, insemination success varies considerably among AI bulls. In an attempt to identify genetic causes of such variation, we undertook a genome-wide association study (GWAS). Imputed genotypes of 652,856 SNPs were available for 7962 AI bulls of the Fleckvieh (FV) population. Male reproductive ability (MRA) was assessed based on 15.3 million artificial inseminations. The GWAS uncovered a strong association signal on bovine chromosome 19 (P = 4.08×10−59). Subsequent autozygosity mapping revealed a common 1386 kb segment of extended homozygosity in 40 bulls with exceptionally poor reproductive performance. Only 1.7% of 35,671 inseminations with semen samples of those bulls were successful. None of the bulls with normal reproductive performance was homozygous, indicating recessive inheritance. Exploiting whole-genome re-sequencing data of 43 animals revealed a candidate causal nonsense mutation (rs378652941, c.483C>A, p.Cys161X) in the transmembrane protein 95 encoding gene TMEM95 which was subsequently validated in 1990 AI bulls. Immunohistochemical investigations evidenced that TMEM95 is located at the surface of spermatozoa of fertile animals whereas it is absent in spermatozoa of subfertile animals. These findings imply that integrity of TMEM95 is required for an undisturbed fertilisation. Our results demonstrate that deficiency of TMEM95 severely compromises male reproductive performance in cattle and reveal for the first time a phenotypic effect associated with genomic variation in TMEM95.

A novel synonymous SNP (A47A) of the <i>TMEM95</i> gene is significantly associated with the reproductive traits related to testis in male piglets

“…Although the novel A47A mutation in the pig TMEM95 gene was a synonymous mutation, it may impart an effect on fitness, splice regulation, and miRNA binding (Parmley et al, 2007). Several computational analyses have also indicated that synonymous mutations impact mRNA stability (Chamary et al, 2005;Quax et al, 2015;Lorenz et al, 2011).…”

“…Several computational analyses have also indicated that synonymous mutations impact mRNA stability (Chamary et al, 2005;Quax et al, 2015;Lorenz et al, 2011). Synonymous mutations affected protein folding and function (Parmley et al, 2007;Gartner et al, 2013). Therefore, the relationships between the SNP locus (A47A) and testis measurement traits in piglet were carried out.…”

“…Although Large White (LW) and Landrace (LD) pigs are the most popular breeds in many countries, especially in western China, they have some reproductive trait defects (Bergfelder-Drüing et al, 2015), e.g., reproductive barriers. It is well known that reproduction traits are determined by both females and males (Beerda et al, 2008;Pausch et al, 2014); however, to date, numerous studies on reproduction traits of female pigs have been published, but the study of male reproductive traits is limited. Owing to the more important role of males in reproduction in the pig industry, male reproduction should be emphasized in the current pig industry (Mack et al, 2014).…”

“…TMEM95 is located on the acrosomal membrane of the sperm head involved in the acrosome reaction (Pausch et al, 2014). Spermatozoa of mt/mt animals, deficiency of TMEM95, demonstrated no fluorescence at the acrosomal membrane, implying that successful fertilization by spermatozoa of mt/mt animals might be compromised, which was consistent with the equatorial segment of the acrosome and contact of the spermatozoon with the cell membrane of the oocyte (Bedford et al, 1979;Palermo et al, 1997;Pausch et al, 2014;Ramasamy et al, 2014). Therefore, TMEM95 is hypothesized to affect spermatogenesis, and it will be possibly associated with male reproductive traits.…”

“…Bovine TMEM95 has a highly conserved single-pass type I transmembrane protein consisting of 183 amino acids (aa) with a predicted extracellular N-terminal signal peptide , a 23-aa transmembrane domain and an 8-aa intracellular C-terminal domain (Pausch et al, 2014). Importantly, the premature stop codon (C161X) of bovine TMEM95 is located within the predicted transmembrane domain and truncates the protein, thus this mutation could cause idiopathic male subfertility in cattle (Pausch et al, 2014). However, its real function and novel genetic variations are unknown in many other species.…”