title: alpha-SNAP and NSF are required in a priming step during the human sperm acrosome reaction.
abstract: The acrosome is a membrane-limited granule that overlies the nucleus of the mature spermatozoon. In response to physiological or pharmacological stimuli it undergoes a special type of Ca2+-dependent exocytosis termed the acrosome reaction (AR), which is an absolute prerequisite for fertilization. Aided by a streptolysin-O permeabilization protocol developed in our laboratory, we have previously demonstrated requirements for Rab3A, N-ethylmaleimide-sensitive factor (NSF), several soluble NSF-attachment protein receptor (SNARE) proteins, and synaptotagmin VI in the human sperm AR. Here, we show that alpha-soluble NSF-attachment protein (alpha-SNAP), a protein essential for most fusion events through its interaction with NSF and the SNARE complex, exhibits a direct role in the AR. First, the presence of alpha-SNAP is demonstrated by the Western blot of human sperm protein extracts. Immunostaining experiments reveal an acrosomal localization for this protein. Second, the Ca2+ and Rab3A-triggered ARs are inhibited by anti-alpha-SNAP antibodies. Third, bacterially expressed alpha-SNAP abolishes exocytosis in a fashion that depends on its interaction with NSF. Fourth, we show a requirement for alpha-SNAP/NSF in a prefusion step early in the exocytotic pathway, after the tethering of the acrosome to the plasma membrane and before the efflux of intra-acrosomal Ca2+. These results suggest a key role for alpha-SNAP/NSF in the AR, and strengthen our understanding of the molecular players involved in the vesicle-to-plasma membrane fusion taking place during exocytosis.
authors.affiliation: Laboratorio de Biología Celular y Molecular, Instituto de Histología y Embriología (IHEM-CONICET), Facultad de Ciencias Médicas, CC 56, Universidad Nacional de Cuyo, 5500 Mendoza, Argentina. ctomes@fcm.uncu.edu.ar
authors.forename: C N
authors.initials: CN
authors.lastname: Tomes
authors.forename: G A
authors.initials: GA
authors.lastname: De Blas
authors.forename: M A
authors.initials: MA
authors.lastname: Michaut
authors.forename: E V
authors.initials: EV
authors.lastname: Farré
authors.forename: O
authors.initials: O
authors.lastname: Cherhitin
authors.forename: P E
authors.initials: PE
authors.lastname: Visconti
authors.forename: L S
authors.initials: LS
authors.lastname: Mayorga
mesh_terms: D000177:Acrosome / Q000737:chemistry / Q000378:metabolism; D020101:Acrosome Reaction / Q000502:physiology*; D000906:Antibodies / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism / Q000494:pharmacology; D005089:Exocytosis / Q000187:drug effects / Q000502:physiology; D006801:Humans; D008297:Male; D050600:SNARE Proteins; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; D013094:Spermatozoa / Q000737:chemistry / Q000378:metabolism*; D033921:Vesicular Transport Proteins / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism* / Q000502:physiology*; D020826:rab3A GTP-Binding Protein / Q000378:metabolism / Q000494:pharmacology
chemical_list: D000906:Antibodies; D050600:SNARE Proteins; D052067:Soluble N-Ethylmaleimide-Sensitive Factor Attachment Proteins; D033921:Vesicular Transport Proteins; D020826:rab3A GTP-Binding Protein; D002118:Calcium