A mouse protein that localizes to acrosome and sperm tail is regulated by Y-chromosome

Abstract

BackgroundAcrosomal proteins play crucial roles in the physiology of fertilization. Identification of proteins localizing to the acrosome is fundamental to the understanding of its contribution to fertilization. Novel proteins are still being reported from acrosome. In order to capture yet unreported proteins localizing to acrosome in particular and sperm in general, 2D-PAGE and mass spectrometry analysis of mouse sperm proteins was done.ResultsOne of the protein spots identified in the above study was reported in the NCBI database as a hypothetical protein from Riken cDNA 1700026L06 that localizes to chromosome number 2. Immunofluorescence studies using the antibody raised in rabbit against the recombinant protein showed that it localized to mouse acrosome and sperm tail. Based on the localization of this protein, it has been named mouse acrosome and sperm tail protein (MAST, [Q7TPM5 (http://www.ncbi.nlm.nih.gov/protein/Q7TPM5)]). This protein shows 96% identity to the rat spermatid specific protein RSB66. Western blotting showed that MAST is expressed testis-specifically. Co-immunoprecipitation studies using the MAST antibody identified two calcium-binding proteins, caldendrin and calreticulin as interacting partners of MAST. Caldendrin and calreticulin genes localize to mouse chromosomes 5 and 8 respectively. In a Yq-deletion mutant mouse, that is subfertile and has a deletion of 2/3rd of the long arm of the Y chromosome, MAST failed to localize to the acrosome. Western blot analysis however, revealed equal expression of MAST in the testes of wild type and mutant mice. The acrosomal calcium-binding proteins present in the MAST IP-complex were upregulated in sperms of Yq-del mice.ConclusionsWe have identified a mouse acrosomal protein, MAST, that is expressed testis specifically. MAST does not contain any known motifs for protein interactions; yet it complexes with calcium-binding proteins localizing to the acrosome. The misexpression of all the proteins identified in a complex in the Yq-del mice invokes the hypothesis of a putative pathway regulated by the Y chromosome. The role of Y chromosome in the regulation of this complex is however not clear from the current study.

Quantitative Proteomic Analysis of Human Airway Cilia Identifies Previously Uncharacterized Proteins of High Abundance

“…In mouse caudal sperm, SPACA9 localizes to sperm head, and is also present on the midpiece and principle piece of sperm tails. This localization is regulated by Y chromosome (71). …”

Y chromosomal noncoding RNAs regulate autosomal gene expression via piRNAs in mouse testis

“…Few other autosomal proteins are also deregulated in different strains of Y-deleted mice besides the ones identified in the proteomics screen; caldendrin is upregulated in XY RIII qdel sperm (22) and acrosin is downregulated in B10.BR-Y del sperm. (33).…”

“…MAST a novel protein that is absent in the sperm proteome localizes to both the acrosome and sperm tail, indicating a role in the penetration of the egg and sperm motility (22). Bioinformatic analysis of the transcript (Riken cDNA 1700001L19) of the hypothetical protein from spot A predicts it as a cilia related gene (44).…”

“…Studies from Burgoyne's lab reported morphological abnormalities in sperms from XY RIII qdel mice (9). In order to understand the connection between the Y-deletion and sperm abnormalities, we performed comparative sperm proteome analysis between normal mice and Y-long arm deletion mutant XY RIII qdel mice by 2D-PAGE and mass spectrometry using protocols standardized in the laboratory (22). This analysis identified 8 protein spots that were differentially expressed in the pI ranges of 4-7 and 5-8 ( Fig.…”

“…µg of small RNA from each tissue was resolved on a 12% Urea PAGE gel in 0.5 x TBE running buffer and transferred onto Hybond N + membrane. Decade marker (Ambion) was labeled and loaded according to the manufacturer's instructions [10][11][12][13][14][15][16][17][18][19][20][21][22][23][24][25]. …”