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A mouse thymidylate synthase pseudogene derived from an aberrantly processed RNA molecule
1Department of Biochemistry, Ohio State University, Columbus 43210.
Abstract:
A DNA fragment containing a mouse-thymidylate-synthase(TS) processed pseudogene was cloned and analyzed. Comparison with the sequences of the mouse TS-encoding gene (ts) and cDNA revealed that the pseudogene started at one of the normal 5' termini of TS mRNA, ended with a poly(A) tail, and was flanked by 16-nucleotide (nt) direct repeats. The region corresponding to the open reading frame was 97.3% identical to that of the cDNA. Two unusual features were observed. First, the poly(A) tail of the pseudogene was located 2 kb downstream from the normal location. Second, the final 10 nt of intron 5 were retained in the 'coding region' of the pseudogene. Therefore, it appears that the pseudogene was derived from a nonfunctional TS 'mRNA' that was aberrantly spliced and polyadenylated. Analysis of the sequence of intron 5 of the ts gene revealed the presence of an alternative 3' splice site 10 nt upstream from the normal splice site. S1-nuclease protection assays showed that about 10% of TS mRNA isolated from mouse cells was spliced at the alternative site.
Insights
Researchers identified a mouse thymidylate synthase (TS) processed pseudogene with unusual features, suggesting it originated from aberrantly spliced and polyadenylated TS mRNA. This pseudogene provides insights into gene expression regulation.
Area of Science:
- Molecular Biology
- Genetics
- Bioinformatics
Background:
- Thymidylate synthase (TS) is a crucial enzyme in DNA synthesis.
- Processed pseudogenes are non-functional DNA copies of mRNA, lacking introns and regulatory elements.
- Understanding pseudogene formation can shed light on gene expression and regulation.
Purpose of the Study:
- To clone and analyze a mouse thymidylate synthase (TS) processed pseudogene.
- To investigate the origin and unusual features of the identified TS pseudogene.
- To explore the implications of aberrant splicing and polyadenylation in pseudogene formation.
Main Methods:
- DNA cloning and sequencing of the mouse TS pseudogene.
- Sequence comparison between the pseudogene, TS gene, and TS cDNA.
- Analysis of intron sequences and alternative splicing sites.
- S1-nuclease protection assays to detect alternative splicing in mouse cells.
Main Results:
- A mouse TS processed pseudogene was identified, sharing high sequence identity with TS cDNA.
- The pseudogene exhibited an unusually located poly(A) tail and retained intronic sequences.
- An alternative 3' splice site in intron 5 of the TS gene was identified.
- Approximately 10% of mouse TS mRNA utilized this alternative splice site.
Conclusions:
- The mouse TS pseudogene likely arose from aberrantly spliced and polyadenylated TS mRNA.
- Aberrant splicing and polyadenylation contribute to the formation of processed pseudogenes.
- The findings highlight the complexity of gene expression and the potential for non-canonical processing events.