Male infertility-linked point mutation reveals a vital binding role for the C2 domain of sperm PLCζ

• DOI: 10.1042/bcj20161057
• Publication Date: 2017-03-08T01:20:23Z
• Authors: Michail Nomikos; Panagiotis Stamat...; Jessica R. Sanders; Konrad Beck; Brian L. Calver; Luke Buntwal; Morgan Lofty; Zili Sideratou; Karl Swann; F. Anthony Lai
• Keywords: Male; 0303 health sciences; Microinjections; Phenylalanine; Gene Expression; C2 Domains; Mice; 03 medical and health sciences; Phosphoinositide Phospholipase C; Amino Acid Substitution; Phosphatidylinositol Phosphates; Fertilization; Liposomes; Oocytes; Animals; Humans; Calcium; Cattle; Female; Calcium Signaling; Isoleucine; Infertility, Male

Sperm-specific phospholipase C zeta (PLCζ) is widely considered to be the physiological stimulus that evokes intracellular calcium (Ca2+) oscillations that are essential for the initiation of egg activation during mammalian fertilisation. A recent genetic study reported a male infertility case that was directly associated with a point mutation in the PLCζ C2 domain, where an isoleucine residue had been substituted with a phenylalanine (I489F). Here, we have analysed the effect of this mutation on the in vivo Ca2+ oscillation-inducing activity and the in vitro biochemical properties of human PLCζ. Microinjection of cRNA or recombinant protein corresponding to PLCζI489F mutant at physiological concentrations completely failed to cause Ca2+ oscillations and trigger development. However, this infertile phenotype could be effectively rescued by microinjection of relatively high (non-physiological) amounts of recombinant mutant PLCζI489F protein, leading to Ca2+ oscillations and egg activation. Our in vitro biochemical analysis suggested that the PLCζI489F mutant displayed similar enzymatic properties, but dramatically reduced binding to PI(3)P and PI(5)P-containing liposomes compared with wild-type PLCζ. Our findings highlight the importance of PLCζ at fertilisation and the vital role of the C2 domain in PLCζ function, possibly due to its novel binding characteristics.