Evidence for a 90 kDa heat-shock protein gene expression in the amphibian oocyte

Abstract

In order to study expression of a 90-kDa heat-shock protein during amphibian oogenesis at physiological temperature, we isolated a Pleurodeles waltl hsc90 cDNA by screening an ovarian cDNA library with a chicken hsp90 cDNA probe. The cDNA thus obtained--named Pw90--shows a high homology level with the hsp90 gene in other species. RNase protection analysis led us to conclude that this sequence is part of the cognate gene hsc90 and is constitutively expressed in oocytes. Furthermore, results of quantitative Northern blot analysis, as well as in situ hybridizations on oocyte sections or lampbrush chromosome spreads, provide evidence for expression of hsc90 transcripts at every stage of oogenesis. Moreover, they point to the fact that an accumulation of transcripts occurs very early in oogenesis. Simultaneously, the expression of HSC90-related protein was analyzed on Western blots using a monoclonal antibody (AC88) and a polyclonal antibody (AP90Ct) raised against the Pleurodeles C-terminal part of HSC90. We provide evidence for a net accumulation of HSC90-related protein in oocytes. Immunolocalization shows that a nuclear transfer occurs in the course of oogenesis and leads to a concentration equilibrium between cytoplasm and nucleus in stage VI oocytes.