Exons 2 and 3 of alpha‐tropomyosin are spliced in a strict mutually exclusive manner. Exon 3 is a default choice, being selected in almost all cell types where the gene is expressed. The default selection arises from a competition between the two exons, in which the stronger branch point/pyrimidine tract elements of exon 3 win. Exon 2 is selected predominantly or exclusively only in smooth muscle cells. We show here that the basis for the smooth muscle‐specific switching of exon selection is inhibition of exon 3. Exon 3 is still skipped with smooth muscle specificity, even in the absence of exon 2. We have defined two conserved sequence elements, one in each of the introns flanking exon 3, that are essential for this regulation. Mutation of either element severely impairs regulated suppression of exon 3. No other exon or intron sequences appear to be necessary for regulation. We have also demonstrated skipping of exon 3 that is dependent upon both regulatory elements in an in vitro splicing assay. We further show that both splice sites of exon 3 must be inhibited in a concerted fashion to switch to selection of exon 2. This may relate to the requirement for negative elements on both sides of the exon.

Production of mRNA in eukaryotic cells involves not only transcription but also various processing reactions such as splicing. Recent experiments have indicated that there are direct physical connections between components of the transcription and processing machinery, supporting previous suggestions that pre-mRNA splicing occurs co-transcriptionally. Here we have used a novel functional approach to demonstrate co-transcriptional regulation of alternative splicing. Exon 3 of the alpha-tropomyosin gene is specifically repressed in smooth muscle cells. By delaying synthesis of an essential downstream inhibitory element, we show that the decision to splice or repress exon 3 occurs during a limited window of opportunity following transcription, indicating that splice site selection proceeds rapidly after transcription.

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Alternative splicing is a common mechanism for regulating gene expression in different cell types. In order to understand this important process, the trans‐acting factors that enforce the choice of particular splicing pathways in different environments must be identified. We have used the rat alpha‐tropomyosin gene as a model system of tissue‐specific alternative splicing. Exon 3 of alpha‐tropomyosin is specifically inhibited in smooth muscle cells allowing the alternative inclusion of exon 2. We have used a novel gene transfer and selection strategy to detect a gene whose expression in fibroblasts is sufficient to switch them to smooth muscle‐specific splicing of alpha‐tropomyosin and also alpha‐actinin. Extracts from the regulating fibroblasts contain an apparently novel 55 kDa protein which binds to RNA elements required for regulation of tropomyosin splicing. This protein is not detected in extracts of non‐regulating cells and is therefore a strong candidate cell‐specific splicing regulator. These experiments advance our understanding of smooth muscle splicing regulation as well as establishing a means for direct cloning of tissue‐specific splicing regulators which have so far been refractory to biochemical analysis.

Este estudio examina la viabilidad de la implementación de una empresa de consultoría en recursos humanos que ofrezca sus servicios por medio del aprovechamiento de las TIC que ofrece Internet. La necesidad de este tipo de servicio existe y ha quedado comprobada en Costa Rica en diversos estudios realizados por profesionales en recursos humanos,   las Mipymes requieren  los servicios de asesoría en recursos humanos para mejorar la gestión mediante la identificación de los problemas y la implementación de soluciones propuestas por asesores externos. Los profesionales que brindan servicios de asesoría deben ser personas que tengan experiencia en la materia y formación académica en el área de recursos humanos, además de habilidades tecnológicas que contribuyan a dar un servicio de calidad a los pequeños y medianos empresarios.