MOLECULAR-CLONING AND DEVELOPMENTAL EXPRESSION OF AN MESSENGER-RNA ENCODING THE 27 KDA OUTER DENSE FIBER PROTEIN OF RAT SPERMATOZOA

We have isolated a cDNA (ODF27), encoding the major 27 kDa protein of rat sperm outer dense fibers (ODF), by screening a testicular lambda-g t11 phage cDNA library with an affinity-purified anti-27 kDa ODF polyc lonal antibody. A cyanogen bromide derived internal amino acid (a.a.) sequence of the 27 kDa ODF protein was identical to an internal region of the deduced a.a. sequence of this cDNA. The cDNA encodes a protein with a high proportion of a repetitive motif, Cys-Gly-Pro, at the car boxy-terminal end, reminiscent of the testis-specific Mst(3)CGP protei ns of Drosophila melanogaster (Schafer et al., 1993. Mol Cell Biol 13: 1708-1718). Nick translation probes of the ODF27 cDNA recognized two c omplementary mRNAs of 1.2 and 1.5 kb in the rat testis. Developmental Northern blot analysis revealed that these mRNAs are first transcribed in round spermatids. In situ hybridization confirmed the haploid expr ession of these transcripts and demonstrated that they are found in th e cytoplasm of spermatids throughout most of the duration of spermioge nesis. They reach a peak in steps 8-10 of spermiogenesis at the time t ranscription ceases, remain at high levels from steps 11 to 15, and di minish in steps 16-18 at the time ODF protein synthesis and assembly a re shown to be maximum. The translation of these transcripts, therefor e, appears to be post-transcriptionally controlled. A literature and N CBI database search revealed that the nucleotide sequence of the 027 c DNA is homologous to the rat gene RT7 (Van Der Hoorn et al., 1990. Dev Biol 142:147-154) and to the rat testis-specific cDNA rts 5/1 (Burfei nd and Hoyer-Fender, 1991. Dev Biol 148:195-204), which encodes a 27 k Da polypeptide. (C) 1994 Wiley-Liss, Inc.