Saccharomyces cerevisiae has been used as a model system to investigate the mechanisms of pre-mRNA splicing but only a few examples of alternative splice site usage have been described in this organism. Using RNA-Seq analysis of nonsense-mediated mRNA decay (NMD) mutant strains, we show that many S. cerevisiae intron-containing genes exhibit usage of alternative splice sites, but many transcripts generated by splicing at these sites are non-functional because they introduce premature termination codons, leading to degradation by NMD. Analysis of splicing mutants combined with NMD inactivation revealed the role of specific splicing factors in governing the use of these alternative splice sites and identified novel functions for Prp17p in enhancing the use of branchpoint-proximal upstream 3' splice sites and for Prp18p in suppressing the usage of a non-canonical AUG 3'-splice site in GCR1. The use of non-productive alternative splice sites can be increased in stress conditions in a promoter-dependent manner, contributing to the down-regulation of genes during stress. These results show that alternative splicing is frequent in S. cerevisiae but masked by RNA degradation and that the use of alternative splice sites in this organism is mostly aimed at controlling transcript levels rather than increasing proteome diversity.
The X-linked transcription factor FOXP3 is expressed by epithelial cells of organs including the breast, where it is considered a tumour suppressor. The chemokine receptor CXCR4 also regulates the development of breast cancer by stimulating cell migration towards CXCL12-expressing sites of metastatic spread. During activation, human T cells show reciprocal regulation of FOXP3 and CXCR4. This study was designed to examine the role FOXP3 plays in metastatic breast cancer, with a particular focus on its potential to regulate CXCR4. Human breast cancer samples showed significantly decreased FOXP3 protein expression but an increased number of CXCR4 transcripts. In comparison with normal primary breast epithelial cells, FOXP3 was down-regulated at both transcript and protein levels in the breast cancer cell lines MCF-7 and MDA-MB-231. In the invasive MDA-MB-231 cells, the remaining FOXP3 was located predominately within the cytoplasm. Following stable FOXP3 overexpression in MDA-MB-231 cells, significant decreases were observed in the expression of ErbB2/HER2, SKP2, c-MYC, and CXCR4. In contrast, an increase in p21 expression led to inhibition of cell proliferation, with a greater proportion in the G1 phase of the cell cycle suggesting the induction of senescence. Specific knockdown of FOXP3 in normal human breast epithelial cells with siRNA significantly increased ErbB2/HER2, SKP2, c-MYC, and CXCR4, and decreased p21 expression. These cells also showed a significantly increased chemotactic response towards CXCL12, consistent with a role for FOXP3 in the regulation of cell migration. Results from this study are consistent with FOXP3 functioning as an important tumour suppressor in breast cancer. Indeed, the potential functions of FOXP3 in breast epithelium can now be extended to include regulation of CXCR4 expression and response to the pro-metastatic chemokine CXCL12.
Although dimorphic sexes have evolved repeatedly in multicellular eukaryotes, their origins are unknown. The mating locus (MT) of the sexually dimorphic multicellular green alga Volvox carteri specifies the production of eggs and sperm and has undergone a remarkable expansion and divergence relative to MT from Chlamydomonas reinhardtii, which is a closely related unicellular species that has equal-sized gametes. Transcriptome analysis revealed a rewired gametic expression program for Volvox MT genes relative to Chlamydomonas and identified multiple gender-specific and sex-regulated transcripts. The retinoblastoma tumor suppressor homolog MAT3 is a Volvox MT gene that displays sexually regulated alternative splicing and evidence of gender-specific selection, both of which are indicative of cooption into the sexual cycle. Thus, sex-determining loci affect the evolution of both sex-related and non-sex-related genes.
Regulatory T cells (Tregs) effectively ameliorate graft-versus-host disease (GVHD). The mechanisms underlying Treg therapeutic effect on GVHD are not fully elucidated. This study investigates whether Treg prevention of GVH tissue damage is associated with blocking CD8 effector T-cell tissue invasion, a question not yet addressed in humans.
The transcription factor FOXP3 is widely known for its role in the development and function of immunoregulatory T cells. However, it has been discovered recently that FOXP3 is also expressed in epithelial cells of the normal human breast, ovary and prostate. Aggressive cancer of these epithelial tissues often correlates with abnormal expression of FOXP3, which can be either absent or underexpressed at transcript or protein levels. It is becoming clear that this failure of normal FOXP3 expression can result in dysregulation of the expression of a range of oncogenes which have been implicated in the development and metastasis of cancer. Recent evidence suggests that FOXP3 might also regulate chemokine receptor expression, providing a possible explanation for the chemokine-driven, tissue-specific spread that is characteristic of many cancers. This review first summarises the general structure, function and properties of FOXP3. This is followed by an analysis of the tumour-suppressive properties of this transcription factor, with particular reference to the development and chemokine-mediated spread of human breast cancer. A final section focuses on potential applications of this new knowledge for therapeutic intervention.
We used RNA sequencing to query the Chlamydomonas reinhardtii transcriptome for regulation by CO(2) and by the transcription regulator CIA5 (CCM1). Both CO(2) and CIA5 are known to play roles in acclimation to low CO(2) and in induction of an essential CO(2)-concentrating mechanism (CCM), but less is known about their interaction and impact on the whole transcriptome. Our comparison of the transcriptome of a wild type versus a cia5 mutant strain under three different CO(2) conditions, high CO(2) (5%), low CO(2) (0.03 to 0.05%), and very low CO(2) (<0.02%), provided an entry into global changes in the gene expression patterns occurring in response to the interaction between CO(2) and CIA5. We observed a massive impact of CIA5 and CO(2) on the transcriptome, affecting almost 25% of all Chlamydomonas genes, and we discovered an array of gene clusters with distinctive expression patterns that provide insight into the regulatory interaction between CIA5 and CO(2). Several individual clusters respond primarily to either CIA5 or CO(2), providing access to genes regulated by one factor but decoupled from the other. Three distinct clusters clearly associated with CCM-related genes may represent a rich source of candidates for new CCM components, including a small cluster of genes encoding putative inorganic carbon transporters.
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