title: Male infertility-linked point mutation reveals a vital binding role for the C2 domain of sperm PLCζ.
abstract: 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 thein vivoCa2+oscillation-inducing activity and thein vitrobiochemical properties of human PLCζ. Microinjection of cRNA or recombinant protein corresponding to PLCζI489Fmutant 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 PLCζI489Fprotein, leading to Ca2+oscillations and egg activation. Ourin vitrobiochemical analysis suggested that the PLCζI489Fmutant 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.
authors.affiliation: College of Medicine, Qatar University, PO Box 2713, Doha, Qatar mixosn@yahoo.com lait@cf.ac.uk.|College of Biomedical and Life Sciences, School of Biosciences, Cardiff University, Cardiff, U.K.
authors.forename: Michail
authors.initials: M
authors.lastname: Nomikos
authors.affiliation: College of Biomedical and Life Sciences, School of Biosciences, Cardiff University, Cardiff, U.K.
authors.forename: Panagiotis
authors.initials: P
authors.lastname: Stamatiadis
authors.affiliation: College of Biomedical and Life Sciences, School of Biosciences, Cardiff University, Cardiff, U.K.
authors.forename: Jessica R
authors.initials: JR
authors.lastname: Sanders
authors.affiliation: College of Biomedical and Life Sciences, School of Dentistry, Cardiff University, Cardiff, U.K.
authors.forename: Konrad
authors.initials: K
authors.lastname: Beck
authors.affiliation: College of Biomedical and Life Sciences, School of Biosciences, Cardiff University, Cardiff, U.K.
authors.forename: Brian L
authors.initials: BL
authors.lastname: Calver
authors.affiliation: College of Biomedical and Life Sciences, School of Biosciences, Cardiff University, Cardiff, U.K.
authors.forename: Luke
authors.initials: L
authors.lastname: Buntwal
authors.affiliation: College of Biomedical and Life Sciences, School of Biosciences, Cardiff University, Cardiff, U.K.
authors.forename: Morgan
authors.initials: M
authors.lastname: Lofty
authors.affiliation: National Center for Scientific Research 'Demokritos', 15310 Aghia Paraskevi, Greece.
authors.forename: Zili
authors.initials: Z
authors.lastname: Sideratou
authors.affiliation: College of Biomedical and Life Sciences, School of Biosciences, Cardiff University, Cardiff, U.K.
authors.forename: Karl
authors.initials: K
authors.lastname: Swann
authors.affiliation: College of Biomedical and Life Sciences, School of Biosciences, Cardiff University, Cardiff, U.K. mixosn@yahoo.com lait@cf.ac.uk.
authors.forename: F Anthony
authors.initials: FA
authors.lastname: Lai
mesh_terms: D019943:Amino Acid Substitution; D000818:Animals; D000070538:C2 Domains; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling; D002417:Cattle; D005260:Female; D005306:Fertilization; D015870:Gene Expression; D006801:Humans; D007248:Infertility, Male / Q000235:genetics*; D007532:Isoleucine / Q000737:chemistry / Q000378:metabolism; D008081:Liposomes / Q000737:chemistry / Q000378:metabolism; D008297:Male; D051379:Mice; D008845:Microinjections; D009865:Oocytes / Q000166:cytology / Q000378:metabolism; D010649:Phenylalanine / Q000737:chemistry / Q000378:metabolism; D018129:Phosphatidylinositol Phosphates / Q000737:chemistry / Q000378:metabolism; D054801:Phosphoinositide Phospholipase C / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D017354:Point Mutation; D011485:Protein Binding; D018075:RNA, Complementary / Q000008:administration & dosage / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000008:administration & dosage / Q000235:genetics / Q000378:metabolism; D013094:Spermatozoa / Q000378:metabolism / Q000473:pathology
chemical_list: D008081:Liposomes; D018129:Phosphatidylinositol Phosphates; D018075:RNA, Complementary; D011994:Recombinant Proteins; D007532:Isoleucine; D010649:Phenylalanine; C499235:PLCZ1 protein, human; D054801:Phosphoinositide Phospholipase C; D002118:Calcium
keywords: calcium signaling; fertilisation; infertility