title: Enhancement of trypsin-like enzymes by A23187 ionophore is crucial for sperm penetration through the egg vestment of the giant freshwater prawn.
abstract: We report the presence of trypsin-like enzymes preferring Boc-QAR-MCA substrate in sperm collected from different portions of male reproductive tracts of the giant freshwater prawn, Macrobrachium rosenbergii and compare enzyme activities before and after an A23187 calcium ionophore treatment. Fluorogenic enzyme assays revealed that testicular sperm lysates showed high trypsin-like enzyme activity but the activity was relatively low in vas deferens sperm lysates as well as in the live sperm. Upon sperm treatment with A23187, trypsin-like activity was greatly enhanced in distal vas deferens sperm. Substrate- and inhibitor-based localization studies indicated that the sperm trypsin-like enzymes were not of a soluble type but were rather of a membrane-borne type, localized at the anterior spike and upper part of the main body. Notable structural changes were also evident in A23187-induced sperm including extensive ruffling of the sperm membrane structure at the base of the main body thereby supporting the acrosome reaction response in this species. We further proved by substrate inhibition assays that the enhanced trypsin-like enzyme activity participates in sperm penetration through the vitelline envelope, a novel sperm-egg penetration mechanism that is unique in this species.
authors.affiliation: Department of Anatomy, Faculty of Science, Mahidol University, Rama VI Road, Ratchathewi, Bangkok, Thailand, 10400.
authors.forename: Atthaboon
authors.initials: A
authors.lastname: Watthammawut
authors.affiliation: Department of Anatomy, Faculty of Science, Mahidol University, Rama VI Road, Ratchathewi, Bangkok, Thailand, 10400.
authors.forename: Monsicha
authors.initials: M
authors.lastname: Somrit
authors.affiliation: Department of Anatomy, Faculty of Science, Mahidol University, Rama VI Road, Ratchathewi, Bangkok, Thailand, 10400.
authors.forename: Somluk
authors.initials: S
authors.lastname: Asuvapongpatana
authors.affiliation: Department of Anatomy, Faculty of Science, Mahidol University, Rama VI Road, Ratchathewi, Bangkok, Thailand, 10400. wattana.wee@mahidol.ac.th.
authors.forename: Wattana
authors.initials: W
authors.lastname: Weerachatyanukul
mesh_terms: D000818:Animals; D000001:Calcimycin / Q000494:pharmacology*; D057075:Enzyme Assays; D005456:Fluorescent Dyes / Q000378:metabolism; D005618:Fresh Water; D007476:Ionophores / Q000494:pharmacology*; D008297:Male; D008954:Models, Biological; D033541:Palaemonidae / Q000187:drug effects / Q000201:enzymology* / Q000648:ultrastructure; D013084:Sperm-Ovum Interactions / Q000187:drug effects*; D013094:Spermatozoa / Q000166:cytology / Q000187:drug effects / Q000201:enzymology* / Q000648:ultrastructure; D013379:Substrate Specificity / Q000187:drug effects; D014357:Trypsin / Q000378:metabolism*; D014361:Trypsin Inhibitors / Q000494:pharmacology
chemical_list: D005456:Fluorescent Dyes; D007476:Ionophores; D014361:Trypsin Inhibitors; D000001:Calcimycin; D014357:Trypsin
keywords: Acrosome reaction; Calcium; Shrimp; Sperm; Trypsin