WBP2 shares a common location in mouse spermatozoa with WBP2NL/PAWP and like its descendent is a candidate mouse oocyte activating factor

Abstract

The sperm-borne oocyte-activating factor (SOAF) resides in the sperm perinuclear theca (PT). A consensus has been reached that SOAF most likely resides in the postacrosomal sheath (PAS), which is the first region of the PT to solubilize upon sperm-oocyte fusion. There are two SOAF candidates under consideration: PLCZ1 and WBP2NL. A mouse gene germline ablation of the latter showed that mice remain fertile with no observable phenotype despite the fact that a competitive inhibitor of WBP2NL, derived from its PPXY motif, blocks oocyte activation when coinjected with WBP2NL or spermatozoa. This suggested that the ortholog of WBP2NL, WBP2, containing the same domain and motifs associated with WBP2NL function, might compensate for its deficiency in oocyte activation. Our objectives were to examine whether WBP2 meets the developmental criteria established for SOAF and whether it has oocyte-activating potential. Immunoblotting detected WBP2 in mice testis and sperm and immunofluorescence localized WBP2 to the PAS and perforatorium of the PT. Immunohistochemistry of the testes revealed that WBP2 reactivity was highest in round spermatids and immunofluorescence detected WBP2 in the cytoplasmic lobe of elongating spermatids and colocalized it with the microtubular manchette during PT assembly. Microinjection of the recombinant forms of WBP2 and WBP2NL into metaphase II mouse oocytes resulted in comparable rates of oocyte activation. This study shows that WBP2 shares a similar testicular developmental pattern and location with WBP2NL and a shared ability to activate the oocyte, supporting its consideration as a mouse SOAF component that can compensate for a WBP2NL.

The perforatorium and postacrosomal sheath of rat spermatozoa share common developmental origins and protein constituents†

“…The molecular mass of WBP2NL is species-specific, as shown in previous studies with mouse (38 kDa), porcine (38 kDa), and bovine (32 kDa) spermatozoa [26]. WBP2 labeling was associated with an immunoreactive band at approximately 40 kDa, identical to that demonstrated in mouse spermatozoa [33]. The amount of sonication required to isolate the perforatorium has been shown to decrease the content of WBP2 in the PT, thereby accounting for a weaker than expected reactivity [33].…”

“…To confirm the perforatorial presence of these proteins, isolated perforatorial tips were loaded and probed with each antibody for immunoblotting analysis (Figure 5B). Note that the postacrosomal identities of these proteins have previously been shown in either bovine or murid sperm studies [25, 26, 33, 36].…”

“…The primary antibodies used in this report included a goat polyclonal anti-GSTO2 antibody (Santa Cruz Biotechnology, Y-12, sc-17943) as formerly validated [25], a rabbit polyclonal anti-WBP2 antibody (ProteinTech, 12030-1-AP) as previously used [33], a rabbit polyclonal affinity-purified anti-histone-H3 antibody (ProteinTech, 17168-1-AP), a mouse monoclonal anti-α-tubulin antibody (Sigma-Aldrich, T6074), a customized rabbit polyclonal affinity-purified anti-PERF15 antibody, and a customized rabbit polyclonal affinity-purified anti-WBP2NL antibody. Immune serum raised in a rabbit against the isolated perforatorial fraction [12] was affinity-purified on the western blot immobilized 15 kDa perforatorial polypeptide (PERF15), as previously described [11].…”

The emerging roles of WBP2 oncogene in human cancers

“…Following biochemical characterization of WBP2 resulting in the knowledge of its binding partners and involvement in steroid signaling [4,5], WBP2 has since been discovered to play roles in a variety of human diseases such as hearing loss and infertility [6,7]. However, it is in the area of cancer where the investigations have been most intense.…”

The eggstraordinary story of how life begins

“…Mating studies with Wbp2nl KO males showed no defects in fertility, with normal size litters being produced by these males. These findings along with the previously discussed data in human sperm suggest that PAWP plays no role in egg activation in mammals, although it remains remotely possible that loss of PAWP expression is being compensated functionally by upregulation of another gene product, or by the recently reported ortholog Wbp2 (Hamilton et al, ). Hachem et al () also examined the issue of whether PAWP expression was altered following Plcz1 gene KO.…”