Evidence for distinct serine protease activities with a potential role in processing the sperm protein fertilin

Lawrence Lum, Carl P. Blobel

Research output: Contribution to journalArticle

87 Citations (Scopus)

Abstract

The guinea pig sperm protein fertilin (previously termed PH-30) plays an important role in sperm-egg fusion, and was the first recognized membrane- anchored metalloprotease/disintegrin protein. Fertilin is a heterodimeric glycoprotein which undergoes at least two distinct proteolytic processing steps. Fertilin α is processed first, in the testis, whereas fertilin β is processed separately during sperm maturation in the epididymis. The final processing of fertilin β occurs immediately adjacent to its predicted integrin ligand domain, and exposes an epitope recognized by a fusion blocking monoclonal antibody. Here, we demonstrate that one or more serine protease activities associated with testicular sperm can process fertilin β in vitro in a fashion that closely mimics the processing pattern observed in vivo during epididymal sperm maturation. In contrast, several proteases that were added to testicular sperm did not mimic the pattern observed in vivo. These findings raise the intriguing possibility that a fertilin β converting protease(s) active in vivo may originate from sperm, instead of from the epididymal epithelium. Further, we show that fertilin α is most likely processed intracellularly in the secretory pathway based on three observations: (i) only processed fertilin α, but not the precursor pro-α can be cell-surface biotinylated; (ii) some processed fertilin α is sensitive to endoglycosidase H, suggesting cleavage occurs prior to the medial Golgi apparatus; (iii) a reanalysis of the N-terminus of processed fertilin α showed that the proteolytic cleavage site is next to four arginine residues, a consensus sequence for intracellular subtilysin type pro-protein convertases. The N-terminal sequence analysis further showed that processed fertilin α contains an intact membrane anchored disintegrin domain, and not a truncated disintegrin domain as reported previously (Blobel, C. P., Wolfsberg, T. G., Turck, C. W., Myles, D. G., Primakoff, P., and White, J. M., Nature 356, 248-252, 1992). Proteolytic processing is thought to play an important role in regulating the function of fertilin, and the present study represents a first step toward a better understanding of protease activities involved in the maturation of fertilin, and potentially other sperm surface proteins.

Original languageEnglish (US)
Pages (from-to)131-145
Number of pages15
JournalDevelopmental Biology
Volume191
Issue number1
StatePublished - Nov 1 1997

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Serine Proteases
Spermatozoa
Disintegrins
Sperm Maturation
Peptide Hydrolases
Fertilins
Blocking Antibodies
Membranes
Epididymis
Glycoside Hydrolases
Secretory Pathway
Consensus Sequence
Golgi Apparatus
Metalloproteases
Integrins
Ovum
Sequence Analysis
Arginine
Testis
Epitopes

ASJC Scopus subject areas

  • Developmental Biology

Cite this

Evidence for distinct serine protease activities with a potential role in processing the sperm protein fertilin. / Lum, Lawrence; Blobel, Carl P.

In: Developmental Biology, Vol. 191, No. 1, 01.11.1997, p. 131-145.

Research output: Contribution to journalArticle

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abstract = "The guinea pig sperm protein fertilin (previously termed PH-30) plays an important role in sperm-egg fusion, and was the first recognized membrane- anchored metalloprotease/disintegrin protein. Fertilin is a heterodimeric glycoprotein which undergoes at least two distinct proteolytic processing steps. Fertilin α is processed first, in the testis, whereas fertilin β is processed separately during sperm maturation in the epididymis. The final processing of fertilin β occurs immediately adjacent to its predicted integrin ligand domain, and exposes an epitope recognized by a fusion blocking monoclonal antibody. Here, we demonstrate that one or more serine protease activities associated with testicular sperm can process fertilin β in vitro in a fashion that closely mimics the processing pattern observed in vivo during epididymal sperm maturation. In contrast, several proteases that were added to testicular sperm did not mimic the pattern observed in vivo. These findings raise the intriguing possibility that a fertilin β converting protease(s) active in vivo may originate from sperm, instead of from the epididymal epithelium. Further, we show that fertilin α is most likely processed intracellularly in the secretory pathway based on three observations: (i) only processed fertilin α, but not the precursor pro-α can be cell-surface biotinylated; (ii) some processed fertilin α is sensitive to endoglycosidase H, suggesting cleavage occurs prior to the medial Golgi apparatus; (iii) a reanalysis of the N-terminus of processed fertilin α showed that the proteolytic cleavage site is next to four arginine residues, a consensus sequence for intracellular subtilysin type pro-protein convertases. The N-terminal sequence analysis further showed that processed fertilin α contains an intact membrane anchored disintegrin domain, and not a truncated disintegrin domain as reported previously (Blobel, C. P., Wolfsberg, T. G., Turck, C. W., Myles, D. G., Primakoff, P., and White, J. M., Nature 356, 248-252, 1992). Proteolytic processing is thought to play an important role in regulating the function of fertilin, and the present study represents a first step toward a better understanding of protease activities involved in the maturation of fertilin, and potentially other sperm surface proteins.",
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