Related Experiment Videos
Ribosomal protein L30 is dispensable in the yeast Saccharomyces cerevisiae
D M Baronas-Lowell1, J R Warner
1Department of Cell Biology, Albert Einstein College of Medicine, Bronx, New York 10461.
Abstract:
In the yeast Saccharomyces cerevisiae, L30 is one of many ribosomal proteins that is encoded by two functional genes. We have cloned and sequenced RPL30B, which shows strong homology to RPL30A. Use of mRNA as a template for a polymerase chain reaction demonstrated that RPL30B contains an intron in its 5' untranslated region. This intron has an unusual 5' splice site, C/GUAUGU. The genomic copies of RPL30A and RPL30B were disrupted by homologous recombination. Growth rates, primer extension, and two-dimensional ribosomal protein analyses of these disruption mutants suggested that RPL30A is responsible for the majority of L30 production. Surprisingly, meiosis of a diploid strain carrying one disrupted RPL30A and one disrupted RPL30B yielded four viable spores. Ribosomes from haploid cells carrying both disrupted genes had no detectable L30, yet such cells grew with a doubling time only 30% longer than that of wild-type cells. Furthermore, depletion of L30 did not alter the ratio of 60S to 40S ribosomal subunits, suggesting that there is no serious effect on the assembly of 60S subunits. Polysome profiles, however, suggest that the absence of L30 leads to the formation of stalled translation initiation complexes.
Insights
In yeast Saccharomyces cerevisiae, the ribosomal protein L30 is produced by two genes, RPL30A and RPL30B. Even without L30, yeast cells can grow, indicating its dispensability for cell viability.
Area of Science:
- Molecular Biology
- Yeast Genetics
- Ribosome Biogenesis
Background:
- The ribosomal protein L30 in Saccharomyces cerevisiae is encoded by two genes, RPL30A and RPL30B.
- RPL30B was cloned and sequenced, revealing strong homology to RPL30A and an unusual intron in its 5' untranslated region.
Purpose of the Study:
- To investigate the functional redundancy and essentiality of the RPL30A and RPL30B genes in yeast.
- To characterize the impact of L30 depletion on ribosomal function and cell growth.
Main Methods:
- Cloning and sequencing of RPL30B.
- Disruption of RPL30A and RPL30B genes via homologous recombination.
- Analysis of growth rates, mRNA, ribosomal protein composition, and polysome profiles in mutant strains.
Main Results:
- RPL30A is the primary source of L30 protein.
- Yeast cells lacking L30 protein (disrupted RPL30A and RPL30B) are viable with only a moderate decrease in growth rate.
- L30 depletion does not significantly affect the assembly of ribosomal subunits but leads to stalled translation initiation complexes.
Conclusions:
- The ribosomal protein L30 is not essential for yeast cell viability.
- The absence of L30 impacts translation initiation, suggesting a role in translational regulation or fidelity.