Molecular cloning, tissue distribution, and hormonal control in the ovary of Cx41 mRNA, a novel Xenopus connexin gene transcript

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Abstract
Gap junctions (GJ) are aggregates of intercellular channels, composed of connexin (Cx) protein, between adjacent cells. The vertebrate ovarian follicle contains homocellular (granulosa cell‐granulosa cell) and heterocellular (granulosa cell‐oocyte) GJ. However, the function of GJ during final oocyte differentiation (maturation) is controversial. The objectives of this study are to reexamine the number and identity of Cx genes that are expressed in the Xenopus ovary, and to examine the potential role of GJ in oocyte maturation by determining the temporal association between changes in ovarian Cx mRNA content and the process of maturation. We used reverse transcriptase‐polymerase chain reaction to amplify ovarian cDNA fragments using degenerate Cx primers. We amplified three Cx‐like fragments: one was novel and two corresponded to known Cx of Xenopus ovaries (Cx38 and 43). The novel fragment was used to screen an ovarian cDNA library. One positive clone was identified and its nucleotide sequence determined. Its deduced amino acid sequence showed that it corresponded to a novel Cx, Cx41, belonging to the Group II class of Cx. Xenopus Cx41 showed the highest homology to rat Cx37 (65% identity, 80% similarity). Also, the last 10 C‐terminal amino acids of Cx41 were identical to those of rat, mouse, and human Cx37. Cx41 transcripts were detected by riboprobe mapping in ovarian somatic cells, heart, leg muscle, liver and eye, but not in brain or in oocytes of any developmental stage. Full‐grown follicles incubated in vitro with human chorionic gonadotropin became committed to mature within 1–4 hr, and physical signs of maturation (germinal vesicle breakdown) were seen at 4–5 hr. Significant reductions in the levels of Cx41 and 43, but not 38 transcripts were seen at 4 hr, after oocytes had committed to mature. Thus, if availability of Cx mRNA determines availability of Cx protein and GJ, our results would suggest that irreversible commitment to maturation occurred prior to major declines in follicular GJ during the periovulatory period. The present study is the first to report the presence of at least two hormone‐responsive Cx gene transcripts (Cx41 and 43 in Xenopus) in ovaries of a single animal species.