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17 September 2014 Hyaluronidase 2: A Novel Germ Cell Hyaluronidase with Epididymal Expression and Functional Roles in Mammalian Sperm
Mark J. Modelski, Gladys Menlah, Yipei Wang, Soma Dash, Kathie Wu, Deni S. Galileo, Patricia A. Martin-DeLeon
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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.

Mark J. Modelski, Gladys Menlah, Yipei Wang, Soma Dash, Kathie Wu, Deni S. Galileo, and Patricia A. Martin-DeLeon "Hyaluronidase 2: A Novel Germ Cell Hyaluronidase with Epididymal Expression and Functional Roles in Mammalian Sperm," Biology of Reproduction 91(5), (17 September 2014). https://doi.org/10.1095/biolreprod.113.115857
Received: 12 November 2013; Accepted: 1 September 2014; Published: 17 September 2014
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