title: Identification of a hyaluronidase, Hyal5, involved in penetration of mouse sperm through cumulus mass.
abstract: A glycosylphosphatidylinositol (GPI)-anchored hyaluronidase, PH-20, on the sperm surface has long been believed to assist sperm penetration through the cumulus mass surrounding the eggs. However, mouse sperm lacking PH-20 were still capable of penetrating the cumulus mass despite a delayed dispersal of cumulus cells. Intriguingly, a 55-kDa hyaluronan-hydrolyzing protein was abundantly present in wild-type and PH-20-deficient mouse sperm. In this study, we purified the 55-kDa mouse protein from soluble protein extracts released from epididymal sperm by acrosome reaction and identified as a hyaluronidase, Hyal5. Hyal5 was exclusively expressed in the testis and formed a 160-kbp gene cluster together with Hyalp1, Hyal4, and Ph-20 on mouse chromosome 6. Hyal5 was a single-chain hyaluronidase present on the plasma and acrosomal membranes of sperm presumably as a GPI-anchored protein. Moreover, hyaluronan zymography revealed that Hyal5 is enzymatically active in the pH range 5-7 and inactive at pH 3 and 4. Both Hyal5-enriched PH-20-free soluble protein extracts and PH-20-deficient mouse sperm were capable of dispersing cumulus cells from the cumulus mass. Cumulus cell dispersal was strongly inhibited by the presence of a hyaluronidase inhibitor, apigenin. These results suggest that in the mouse, Hyal5 may function principally as a "cumulus matrix depolymerase" in the sperm penetration through the cumulus mass and in the local hyaluronan hydrolysis near or on the surface of the egg zona pellucida to enable the proximal region of sperm tail to move freely. PH-20 may compensate in part for the functional roles of Hyal5.
authors.affiliation: Graduate School of Life and Environmental Sciences, University of Tsukuba, Tsukuba Science City, Ibaraki, Japan.
authors.forename: Ekyune
authors.initials: E
authors.lastname: Kim
authors.forename: Daichi
authors.initials: D
authors.lastname: Baba
authors.forename: Masanori
authors.initials: M
authors.lastname: Kimura
authors.forename: Misuzu
authors.initials: M
authors.lastname: Yamashita
authors.forename: Shin-ichi
authors.initials: S
authors.lastname: Kashiwabara
authors.forename: Tadashi
authors.initials: T
authors.lastname: Baba
mesh_terms: D000818:Animals; D001483:Base Sequence; D015815:Cell Adhesion Molecules / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D006821:Hyaluronoglucosaminidase / Q000235:genetics* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D015151:Immunoblotting; D008297:Male; D051379:Mice; D008969:Molecular Sequence Data; D020133:Reverse Transcriptase Polymerase Chain Reaction; D017422:Sequence Analysis, DNA; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013084:Sperm-Ovum Interactions / Q000502:physiology*; D013094:Spermatozoa / Q000201:enzymology*; D013737:Testis / Q000378:metabolism
chemical_list: D015815:Cell Adhesion Molecules; C510449:Hyal5 protein, mouse; D006821:Hyaluronoglucosaminidase; C066852:hyaluronidase PH-20