The present study examined 1) whether the estrogen-regulated destabilization of albumin mRNA occurs in the nuclear or extranuclear fraction of the liver cell, and 2) whether the selective posttranscriptional regulation of albumin mRNA stability might result from covalent changes introduced in the processing or polyadenylation of the primary transcript. The disappearance of albumin mRNA after estrogen is restricted to the extranuclear fraction of the cell. Transient changes in steady state levels of the mature nuclear transcript were observed that mirrored the transient estrogen-induced changes previously reported for albumin gene transcription. When assayed 24 h after estrogen (when albumin RNA is virtually undetectable in the extranuclear fraction) the steady state levels of both the primary and mature albumin transcripts found in the nucleus were the same as observed in control animals. Estrogen had no effect on the splicing or selection of polyadenylation sites on the 3'-UTR as determined by high resolution gel analysis of the 3'-UTR and DNA sequencing of cDNA clones isolated from a liver library from an estrogen-treated male Xenopus. Most eukaryotic mRNAs have poly(A) tracts several hundred residues in length, and recent studies have demonstrated that a change in the stability of a number of mRNAs correlates directly with the degree of polyadenylation. Albumin contrasts sharply with this, first because it has an exceptionally short poly(A) tail of 17 residues, and second because the degree of polyadenylation is totally unrelated to its destabilization in response to estrogen. These findings indicate that a unique pathway is involved in the regulation of albumin RNA stability by estrogen in Xenopus.

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The role of Ca 2+ in volume regulation remains obscure. Before it can be investigated, however, the time courses of osmolyte and cell volume regulation and the effect of Ca 2+ must be simultaneously specified in a suitable cell type. We have tested the red blood cells of Noetia ponderosa in that context. Our results show that the regulation of cell volume of the erythrocytes following hypoosmotic stress has two components. The first is an efflux of intracellular K + and Cl~ (but not Na + ) that begins immediately with the onset of hypoosmotic exposure. The second component, an efflux of taurine, follows the first, but only after many minutes. In addition, clam erythrocyte volume regulation is dependent on external [Ca 2+ ]. Volume recovery is potentiated in hypoosmotic media containing elevated Ca 2+ levels. Taurine efflux from clam erythrocytes in hypoosmotic conditions is reduced in Ca 2+ -free media and potentiated in high Ca 2+ media. In contrast, the effluxes of K + and Cl are not sensitive to extracellular Ca 2+ levels in either isosmotic or hypoosmotic media. Thus, the effluxes of ionic and organic osmolytes from these cells are controlled by mechanisms that differ in response time and Ca 2+ sensitivity. These results suggest that the clam cells have an unexceptional volume regulatory mechanism and should therefore make an excellent model with which to study the role of Ca 2+ in that process.

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In adult Xenopus serum, albumin gene expression is regulated by estrogen through the selective destabilization of its mRNA during the vitellogenic response. The present study reports the cDNA sequence of both the 68K and 74K Xenopus albumin mRNAs, their derived amino acid sequence, and the regulation of albumin gene expression during embryogenesis. Albumin mRNA has a 39 nucleotide 5' untranslated region terminating in a consensus translation initiation site. The derived amino acid sequence yields a 24-amino acid hydrophobic leader sequence (terminating in Lys-Arg) that shares significant homology with the leader peptide of rat albumin. Overall there is 37% sequence identity between rat and frog albumin, with exact conservation of all but one Cys residue and the Pro residues responsible for the three domain structure of the mature protein. The 74K albumin (unlike the 68K albumin) is glycosylated; a point mutation converting Lys256 to Asn introduces an N-linked glycosylation site that is similar to one found in the sequence of mammalian alpha-fetoproteins. A larval albumin-like protein was not detectable by silver staining in serum of tadpoles before the beginning of metamorphosis at stage 48. Albumin mRNA is absent from early tadpoles (stages 22-47); however, it is rapidly induced at stage 48 as one of the earliest manifestations of metamorphosis. Exposure of embryos to 10(-8) M T3, which regulates amphibian metamorphosis, resulted in the premature induction of albumin mRNA, such that it is evident by stage 43.

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The pathogenic dimorphic fungal organism Blastomyces dermatitidis exists as a budding yeast at 37°C and as a mycelium at 25°C. While the conversion of one morphological phase of B. dermatitidis to another has long been known to be a thermally dependent process, little of the accompanying biochemical or genetic events controlling the phase transition has been elucidated. Using differential cDNA library screening, we have identified one transcript, bysi, in B. dermatitidis that is expressed at very high levels in the yeast phase but whose levels diminish rapidly when yeast cells are transferred to 25°C to promote conversion to the mycelial phase. Although the 0.95-kb bysi transcript is absent in B. dermatitidis mycelia maintained at 25°C, transfer of mycelial cultures to 37°C results in the reappearance of bysI within 12 h. bysl codes for a protein of 18.6 kDa that contains multiple putative phosphorylation sites, a hydrophobic N terminus, and two 34-amino-acid domains with similarly spaced nine-amino-acid degenerative repeating motifs. Although the nature of the thermal dependency ofbysi expression and the function of the bysl protein are unknown, the strong expression of this transcript specifically in the yeast phase ofB. dermatitidis may prove to be very useful in the development of more specific and sensitive diagnostic methods for blastomycosis.