CP β3, a Novel Isoform of an Actin-Binding Protein, Is a Component of the Cytoskeletal Calyx of the Mammalian Sperm Head

Abstract

In the mammalian sperm head, the nucleus is tightly associated with the calyx, a cell type-specific cytoskeletal structure. Previously, we have identified and characterized some basic proteins such as calicin and cylicins I and II as major calyx components of bovine and human spermatids and spermatozoa. Surprisingly we have now discovered another calyx constituent which by amino acid sequencing and cDNA cloning was recognized as a novel isoform of the widespread β subunit of the heterodimeric actin-binding “capping protein” (CP). This polypeptide, CP β3, of sperm calices, is identical with the β2 subunit present in diverse somatic cell types, except that it shows an amino-terminal extension of 29 amino acids and its mRNA is detected only in testis and, albeit in trace amounts, brain. This CP β3 mRNA contains the additional sequence, encoded by exon 1 of the gene, which is missing in β2 mRNAs. Antibodies specific for the β3 amino-terminal addition have been used to identify the protein by immunoblotting and to localize it to the calyx structure by immunofluorescence microscopy. We conclude that in spermiogenesis the transcription of the gene encoding the β1, β2, and β3 CP subunits is regulated specifically to include exon 1 and to give rise to the testis isoform CP β3, which is integrated into the calyx structure of the forming sperm head. This surprising finding of an actin-binding protein isoform in an insoluble cytoskeletal structure is discussed in relation to the demonstrated roles of actin and certain actin-binding proteins, such asLimulusα-scruin, in spermiogenesis and spermatozoa.