Acidic hyaluronidase activity is present in mouse sperm and is reduced in the absence of SPAM1: Evidence for a role for hyaluronidase 3 in mouse and human sperm

Abstract

The molecular mechanisms underlying sperm penetration of the physical barriers surrounding the oocyte have not been completely delineated. Although neutral-active or "reproductive" hyaluronidases (hyases), exemplified by Sperm Adhesion Molecule 1 (SPAM1), are thought to be responsible for hyaluronan digestion in the egg vestments and for sperm-zona binding, their roles in mouse sperm have been recently questioned. Here we report that acidic "somatic" Hyaluronidase 3 (HYAL3) exists in two isoforms in human (~47 kDa, ~55 kDa) and mouse (~44, 47kDa) sperm where it resides on the plasma membrane over the head and midpiece. Mouse isoforms are differentially distributed in the soluble (SAP), membrane (MBP), and acrosomereacted (AR) fraction where they are most abundant. Comparisons of zymography of Hyal3 null and wild-type (WT) AR and MBP fractions show significant HYAL3 activity at pH 3 and 4, and less at 7. At pH 4, a second acid-active hyase band at ~57 kDa is present in the AR fraction. HYAL3 activity was confirmed using immunoprecipitated HYAL3 and spectrophotometry. In total proteins, hyase activity was higher at pH 6 than at 4 where Spam1 nulls had significantly (P<0.01) diminished activity, indicating that murine SPAM1 has acidic activity. Although fully fertile, Hyal3 null sperm showed delayed cumulus penetration and reduced acrosomal exocytosis. HYAL3, similar to SPAM1 with which it shares 74.6% structural similarity, exists in epididymal tissue/fluid from which it is acquired by caudal mouse sperm in vitro. Our results indicate for the first time the concerted activity of both neutral-and acid-active hyaluronidases in sperm.

Oviductosome-Sperm Membrane Interaction in Cargo Delivery

“…Confocal Analysis of Indirect Immunofluorescence Sperm Staining-Immunofluorescence staining was performed as previously described (29,30). Briefly, capacitated sperm were fixed with 4% paraformaldehyde (cat.…”

HYAL-2–WWOX–SMAD4 Signaling in Cell Death and Anticancer Response

“…HYAL-3 lacks an intrinsic hyaluronidase activity in somatic cells in vivo , whereas it participates in the hyaluronan metabolism by augmenting the activity of HYAL-1 (Hemming et al, 2008). HYAL-3 is abundant in the testis, and the enzymatic activity has been detected in spermatozoa (Reese et al, 2010). PH-20, also known as SPAM1, is a GPI-anchored hyaluronidase, which is expressed in the lysosome-derived acrosome on a sperm's surface.…”

Germ-cell hyaluronidases: their roles in sperm function

“…This challenges the classification of HYAL3 as a somatic hyase. While the enzymatic activity of HYAL3 has been clearly detected in spermatozoa (Reese et al. , 2010), it is debatable whether it exists in somatic cells (Lokeshwar et al.…”

“…However, more recent studies indicate that in mice, both HYALP1 (Miller et al. , 2007) and HYAL3 contribute to this activity, and that an acid‐active 57 kDa hyase is also present in spermatozoa (Reese et al. , 2010).…”

“…, 2010). HYAL3, which also exists in human spermatozoa, increasing the number in humans to two, shares the highest similarity with SPAM1 (76.5%) at the acidic hyase domain (Reese et al. , 2010).…”