Identification and characterization of espin, an actin-binding protein localized to the F-actin-rich junctional plaques of Sertoli cell ectoplasmic specializations

James R. Bartles*, Allison Wierda, Lili Zheng

*Corresponding author for this work

Research output: Contribution to journalArticle

105 Scopus citations

Abstract

Ectoplasmic specializations are membrane-cytoskeletal assemblages found in Sertoli cells at sites of attachment to elongate spermatids or neighboring Sertoli cells. They are characterized in part by the presence of a unique junctional plaque which contains a narrow layer of parallel actin bundles sandwiched between the Sertoli cell plasma membrane and an affiliated cistern of endoplasmic reticulum. Using a monoclonal antibody, we have identified 'espin,' a novel actin-binding protein localized to ectoplasmic specializations. By immunogold electron microscopy, espin was localized to the parallel actin bundles of ectoplasmic specializations at sites where Sertoli cells contacted the heads of elongate spermatids. The protein was also detected at the sites of ectoplasmic specializations between neighboring Sertoli cells. Espin exhibits an apparent molecular mass of ~ 110 kDa in SDS gels. It is encoded by an ~ 2.9 kb mRNA, which was found to be specific to testis among the 11 rat organs and tissues examined. On the basis of cDNA sequence, espin is predicted to be an 836 amino acid protein which contains 8 ankyrin-like repeats in its N-terminal third, a potential P-loop, two proline-rich peptides and two peptides which contain clusters of multiple glutamates bracketed by arginines, lysines and glutamines in a pattern reminiscent of the repetitive motif found in the protein trichohyalin. The ankyrin-like repeats and a 66 amino acid peptide in the C terminus show significant sequence similarity to proteins encoded by the forked gene of Drosophila. A fusion protein containing the C-terminal 378 amino acids of espin was found to bind with high affinity (K(d) = ~ 10 nM) to F-actin in vitro with a stoichiometry of ~ 1 espin per 6 actin monomers. When expressed by transfected NRK fibroblasts, the same C-terminal fragment of espin was observed to decorate actin fibers or cables. On the basis of its structure, localization and properties, we hypothesize that espin is involved in linking actin filaments to each other or to membranes, thereby potentially playing a key role in the organization and function of the ectoplasmic specialization.

Original languageEnglish (US)
Pages (from-to)1229-1239
Number of pages11
JournalJournal of cell science
Volume109
Issue number6
StatePublished - Jun 1 1996

Keywords

  • Ankyrin repeat
  • Cytoskeleton
  • Espin
  • Forked
  • Junction
  • Seminiferous epithelium
  • Spermiogenesis

ASJC Scopus subject areas

  • Cell Biology

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