Stem cell antigen-1 is necessary for cell-cycle withdrawal and myoblast differentiation in C2C12 cells.

Extracellular signaling pathways regulating myoblast differentiation and cell-cycle withdrawal are not completely understood. Stem cell antigen-1 (Sca-1/Ly-6A/E) is a glycosylphosphatidylinositol-anchored membrane protein known for its role in T-cell activation, and recently described as a marker for regeneration-competent myoblasts. We previously determined that expression of Sca-1/Ly-6A is transiently upregulated during myocyte cell-cycle withdrawal; however, a specific function for Sca-1 in myogenesis has not been described. Here, we show that Sca-1 expression on the surface of a subpopulation of differentiating C2C12 myoblasts is maximal at the time of cell-cycle withdrawal, and that blocking Sca-1 with monoclonal antibodies or downregulating Sca-1 expression by antisense both promotes proliferation and inhibits myotube formation. Downregulating Sca-1 expression derepresses Fyn at the time of myoblast cell-cycle withdrawal, and dominant-negative and constitutively active Fyn mutants rescue and recapitulate the Sca-1 antisense phenotype, respectively. This suggests a Fyn-mediated mechanism for Sca-1 action. Thus, we demonstrate an unprecedented role for Sca-1 in early myogenesis in C2C12 cells, and propose a novel pathway from the myoblast cell surface to intracellular signaling networks controlling proliferation versus differentiation in mammalian muscle. These findings suggest that, beyond its role as a marker for muscle progenitors, Sca-1 may be an important therapeutic target for promoting muscle regeneration.

Distinct domains of human CDC5 direct its nuclear import and association with the spliceosome.

Genetic studies have shown that CDC5 proteins are essential for G2 progression and mitotic entry. CDC5 homologs in yeast and mammals are essential for pre-messenger ribonucleic acid (mRNA) processing. Other gene products also have been shown to play roles in both pre-mRNA splicing and cell cycle regulation, prompting the description of several models to explain the mechanism(s) linking these two processes. In this study, we demonstrate that the amino-terminus of human CDC5 directs nuclear import independent of consensus nuclear localization signals or phosphorylation, and that the carboxyl-terminus of human CDC5 preferentially associates with spliceosomal complexes in proximity of RNA transcription during interphase. hCDC5 colocalizes with Sm proteins in a cell cycle- and domain-dependent manner, and several proteins in the human CDC5-associated complex are identified that suggest potential roles for the complex in coupling pre-mRNA splicing to transcriptional activation and protein translation. These results indicate that human CDC5 may function in pre-mRNA processing and cell cycle progression through more than one mechanism.

Ultraviolet-B responses of nuclear genes encoding light-harvesting complex II proteins in pea (Pisum sativum) are altered by norflurazon- and photobleaching-induced chloroplast changes.

The role of functional and intact chloroplasts in mediating the ultraviolet-B (UV-B, 290-320 nm) regulation of two nuclear genes encoding light-harvesting complex II proteins in pea (Pisum sativum L. cv. Extra Early Alaska) was studied. Plants with chloroplasts lacking or containing carotenoids and functional photosystem II were obtained by growth under dim red light (0.2 µmol m-2 s-1 ) in the presence or absence of norflurazon (NF), an inhibitor of carotenoid biosynthesis. The NF-treatment resulted in an increase in AB80 (lhcb1* 2) mRNA but no substantial change in Cab-8 (lhcb1* 4) mRNA, indicating that the mRNA accumulations for AB80 and Cab-8 were differently correlated with the presence and absence of carotenoids. The mRNA levels for both Cab-8 and AB80 in the NF-treated plants were reduced to the same extent by partially photobleaching the chloroplasts with 3 h of higher intensity white light (W, 110 µmol m-2 s-1 ), suggesting that chloroplast integrity was equally important for transcript accumulation of both genes. The mRNAs of both Cab-8 and AB80 in non-NF-treated control plants were decreased by UV-B irradiation, with the level of AB80 mRNA reduced to a greater extent. The UV-B-induced mRNA reduction of both genes was inhibited by NF. The difference between the UV-B responses of the two genes was unaffected by NF, but was abolished by photobleaching the NF-treated plants prior to the UV-B irradiation. Therefore, the presence of carotenoids enhanced rather than prevented the UV-B down-regulation, and the difference in UV-B responses of the two genes may be dependent on chloroplast integrity.