Uncoupling yeast intron recognition from transcription with recursive splicing.

P J Lopez; B Séraphin

doi:10.1093/embo-reports/kvd065

Home/Archive/PubMed

PubMed · 11269499

Uncoupling yeast intron recognition from transcription with recursive splicing.

P J Lopez·B Séraphin

View original source ↗Export metadata ↓

Abstract

Pre-mRNA splicing has to be coordinated with other processes occurring in the nucleus including transcription, mRNA 3' end formation and mRNA export. To analyze the relationship between transcription and splicing, we constructed a network of nested introns. Introns were inserted in the 5' splice site and/or branchpoint of a synthetic yeast intron interrupting a reporter gene. The inserted introns mask the recipient intron from the cellular machinery until they are removed by splicing. Production of functional mRNA from these constructs therefore requires recognition of a spliced RNA as a splicing substrate. We show that recurrent splicing occurs in a sequential and ordered fashion in vivo. Thus, in Saccharomyces cerevisiae, intron recognition and pre-spliceosome assembly is not tightly coupled to transcription.

Explore related subjects

Genes, Fungal Introns Nucleic Acid Conformation Nucleic Acid Precursors Oligonucleotide Probes Plasmids RNA Splicing RNA, Fungal RNA, Messenger Saccharomyces cerevisiae Transcription, Genetic beta-Galactosidase Journal Article Research Support, Non-U.S. Gov't

Keep this discovery

Explore connections, maps & timelines

BibTeX RIS

P J Lopez, B Séraphin. 2000. Uncoupling yeast intron recognition from transcription with recursive splicing.. https://doi.org/10.1093/embo-reports%2Fkvd065

Cite the original work for its findings. Save a collection to share your selection of sources.

KEEP EXPLORING

Related citations

PubMed2006

A novel high-throughput screen reveals yeast genes that increase secretion of heterologous proteins.

The yeast Saccharomyces cerevisiae is an attractive host for the production of heterologous proteins. However, low-yield production of many proteins (from micrograms to milligrams/liter) leaves considerable room for optimization. By engineering the yeast cell via traceable genome-wide libraries, genes that can enhance protein expression level because of their roles in protein transcription, translation, folding, and trafficking processes can be readily identified. This report details a novel approach that combines yeast cDNA overexpression libraries with yeast surface display to allow the rapid flow cytometric screening of engineered yeast for gene products that improve the display of heterologous proteins. After optimization of the screening conditions, a genome-wide scan yielded five yeast gene products that promoted increased display levels of a single-chain T-cell receptor (scTCR). The display-enhancing genes included those coding for cell wall proteins (CCW12, CWP2, and SED1), a ribosomal subunit protein (RPP0), and an endoplasmic reticulum-resident protein (ERO1). Under the premise that yeast surface display levels could be used as a predictor of secretion efficiency, each display-enhancing gene product was tested for its ability to affect secretion levels of multiple scTCR and single-chain antibodies (scFv). All of the selected yeast gene products were shown to promote increased secretion of active protein (1.5-fold to 7.9-fold), with CCW12 and ERO1 being the most generalizable enhancers of scFv/scTCR secretion.

Genes, Fungal↗

PubMed2006

The fission yeast stress MAPK cascade regulates the pmp3+ gene that encodes a highly conserved plasma membrane protein.

In eukaryotic organisms, stress-activated mitogen-activated protein kinases (MAPK) play crucial roles in transmitting environmental signals to regulate gene expression for cellular stress adaptation. Here we report that, in the fission yeast Schizosaccharomyces pombe, Spc1/Sty1 MAPK and the Atf1 transcription factor regulate the stress-induced expression of Pmp3, a ubiquitous small membrane protein implicated in the modulation of the plasma membrane potential. The pmp3 null mutant, as well as the spc1 and atf1 mutants, is hypersensitive to the cationic antibiotic hygromycin B. Transcriptional regulation of the Pmp3-like genes by the stress-activated MAPK may also be conserved in other eukaryotes, including plants.

Genes, Fungal↗

PubMed2006

Genetic pleiotropy in Saccharomyces cerevisiae quantified by high-resolution phenotypic profiling.

Genetic pleiotropy, the ability of a mutation in a single gene to give rise to multiple phenotypic outcomes, constitutes an important but incompletely understood biological phenomenon. We used a high-resolution and high-precision phenotypic profiling approach to quantify the fitness contribution of genes on the five smallest yeast chromosomes during different forms of environmental stress, selected to probe a wide diversity of physiological features. We found that the extent of pleiotropy is much higher than previously claimed; 17% of the yeast genes were pleiotropic whereof one-fifth were hyper-pleiotropic. Pleiotropic genes preferentially participate in functions related to determination of protein fate, cell growth and morphogenesis, signal transduction and transcription. Contrary to what has earlier been proposed we did not find experimental evidence for slower evolutionary rate of pleiotropic genes/proteins. We also refute the existence of phenotypic islands along chromosomes but report on a remarkable loss both of pleiotropy and of phenotypic penetrance towards chromosomal ends. Thus, the here reported features of pleiotropy both have implications on our understanding of evolutionary processes as well as the mechanisms underlying disease.

Genes, Fungal↗