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A conditional yeast mutant deficient in mRNA transport from nucleus to cytoplasm
T Kadowaki1, Y Zhao, A M Tartakoff
1Institute of Pathology, Case Western Reserve University, Cleveland, OH 44106.
Abstract:
Transport of mRNA from nucleus to cytoplasm is critical for eukaryotic gene expression; however, the mechanism of export is unknown. Selection and screening procedures have therefore been used to obtain a family of temperature-sensitive conditional mutants of Saccharomyces cerevisiae that accumulate poly(A)+ RNA in the nucleus when incubated at 37 degrees C, as judged by in situ hybridization. In one such mRNA transport mutant, mtr1-1, RNA synthesis continues, the export of poly(A)+ RNA is inhibited, intranuclear poly(A)+ is remarkably stable, and protein synthesis gradually stops. Thus, there is no tight coupling between RNA synthesis and export. The export lesion is reversible. Although mRNA export is clearly not a default option, neither inhibition of protein synthesis, inhibition of mRNA splicing, nor inhibition of poly(A)-binding protein function blocks export of the average poly(A)+, as judged by in situ hybridization. Further analysis of the family of mtr mutants should help map the path of RNA transport.
Insights
Researchers identified temperature-sensitive mutants in yeast to study messenger RNA (mRNA) nuclear export. One mutant, mtr1-1, shows blocked mRNA export, revealing insights into eukaryotic gene expression mechanisms.
Area of Science:
- Molecular Biology
- Cell Biology
- Genetics
Background:
- Nuclear mRNA export is essential for eukaryotic gene expression, but the underlying mechanisms remain largely unknown.
- Understanding mRNA transport is crucial for deciphering how genetic information flows from the nucleus to the cytoplasm.
Purpose of the Study:
- To identify and characterize mutants affecting mRNA export in Saccharomyces cerevisiae.
- To investigate the relationship between RNA synthesis, mRNA export, and protein synthesis.
Main Methods:
- Utilized selection and screening to isolate temperature-sensitive mutants of Saccharomyces cerevisiae.
- Employed in situ hybridization to assess poly(A)+ RNA localization in wild-type and mutant strains at different temperatures.
- Analyzed RNA synthesis, mRNA export, and protein synthesis rates in identified mutants.
Main Results:
- Identified a family of mutants accumulating poly(A)+ RNA in the nucleus at the restrictive temperature (37°C).
- The mtr1-1 mutant demonstrated inhibited poly(A)+ RNA export, stable intranuclear RNA, and gradual cessation of protein synthesis.
- Demonstrated that RNA synthesis and mRNA export are not tightly coupled, and the export defect is reversible.
Conclusions:
- The mtr1-1 mutant provides a valuable tool for dissecting the molecular pathways of mRNA nuclear export.
- Further analysis of these mRNA transport mutants is expected to elucidate the intricate route of RNA transport in eukaryotes.