title: Hyaluronidase 2: a novel germ cell hyaluronidase with epididymal expression and functional roles in mammalian sperm.
abstract: To initiate the crucial cell adhesion events necessary for fertilization, sperm must penetrate extracellular matrix barriers containing hyaluronic acid (HA), a task thought to be accomplished by neutral-active hyaluronidases. Here we report that the ~57 kDa hyaluronidase 2 (HYAL2) that in somatic tissues has been highly characterized to be acid-active is present in mouse and human sperm, as detected by Western blot, flow cytometric, and immunoprecipitation assays. Immunofluorescence revealed its presence on the plasma membrane over the acrosome, the midpiece, and proximal principal piece in mice where protein fractionation demonstrated a differential distribution in subcellular compartments. It is significantly more abundant in the acrosome-reacted (P = 0.04) and soluble acrosomal fractions (P = 0.006) (microenvironments where acid-active hyaluronidases function) compared to that of the plasma membrane where neutral hyaluronidases mediate cumulus penetration. Using HA substrate gel electrophoresis, immunoprecipitated HYAL 2 was shown to have catalytic activity at pH 4.0. Colocalization and coimmunoprecipitation assays reveal that HYAL2 is associated with its cofactor, CD44, consistent with CD44-dependent HYAL2 activity. HYAL2 is also present throughout the epididymis, where Hyal2 transcripts were detected, and in the epididymal luminal fluids. In vitro assays demonstrated that HYAL2 can be acquired on the sperm membrane from epididymal luminal fluids, suggesting that it plays a role in epididymal maturation. Because similar biphasic kinetics are seen for HYAL2 and SPAM1 (Sperm adhesion molecule 1), it is likely that HYAL2 plays a redundant role in the catalysis of megadalton HA to its 20 kDa intermediate during fertilization.
authors.affiliation: Department of Biological Sciences, University of Delaware, Newark, Delaware.
authors.forename: Mark J
authors.initials: MJ
authors.lastname: Modelski
authors.affiliation: Department of Biological Sciences, University of Delaware, Newark, Delaware.
authors.forename: Gladys
authors.initials: G
authors.lastname: Menlah
authors.affiliation: Department of Biological Sciences, University of Delaware, Newark, Delaware.
authors.forename: Yipei
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Biological Sciences, University of Delaware, Newark, Delaware.
authors.forename: Soma
authors.initials: S
authors.lastname: Dash
authors.affiliation: Department of Biological Sciences, University of Delaware, Newark, Delaware.
authors.forename: Kathie
authors.initials: K
authors.lastname: Wu
authors.affiliation: Department of Biological Sciences, University of Delaware, Newark, Delaware.
authors.forename: Deni S
authors.initials: DS
authors.lastname: Galileo
authors.affiliation: Department of Biological Sciences, University of Delaware, Newark, Delaware pdeleon@udel.edu.
authors.forename: Patricia A
authors.initials: PA
authors.lastname: Martin-DeLeon
mesh_terms: D000818:Animals; D015815:Cell Adhesion Molecules / Q000235:genetics / Q000502:physiology*; D004822:Epididymis / Q000201:enzymology / Q000378:metabolism*; D005260:Female; D005306:Fertilization; D058851:GPI-Linked Proteins / Q000235:genetics / Q000502:physiology; D015870:Gene Expression; D005854:Germ Cells / Q000201:enzymology / Q000378:metabolism*; D006801:Humans; D006820:Hyaluronic Acid / Q000378:metabolism; D006821:Hyaluronoglucosaminidase / Q000235:genetics / Q000502:physiology*; D008297:Male; D008322:Mammals; D051379:Mice; D008810:Mice, Inbred C57BL; D013084:Sperm-Ovum Interactions; D013094:Spermatozoa / Q000201:enzymology / Q000378:metabolism*
chemical_list: D015815:Cell Adhesion Molecules; D058851:GPI-Linked Proteins; D006820:Hyaluronic Acid; C473096:Hyal2 protein, human; C469221:Hyal2 protein, mouse; D006821:Hyaluronoglucosaminidase
keywords: acrosome reaction; epididymis; fertilization; sperm maturation; zona pellucida