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Molecular cloning, sequencing, chromosomal localization and expression of mouse p21 (Waf1)
1Laboratory of Genetics, NCI/NIH, Bethesda, Maryland 20892.
Abstract:
The recent discovery that expression of Waf1 (p21), an inhibitor of cyclin-dependent kinases, is induced by the tumor suppressor p53 provides an important linkage between growth suppression and the cell cycle. We report here the cloning and sequencing of a mouse p21 cDNA that contains the entire coding region. Hybridization of the mouse p21 probe in Southern blot analyses confirms that p21 is a single-copy gene and that the corresponding locus, Waf1, lies proximal to H-2 on mouse chromosome 17. In northern analyses, the expression of p21 is found in most normal mouse tissues, but a surprising lack of correlation is found between mRNA levels of p21 and p53. In order to determine which regions of p21 are most evolutionarily conserved, we have compared the cDNA sequences for the entire p21 coding region in 13 different mouse strains or species and the human p21 sequence. We conclude that two regions (corresponding to human codons 21-60 and 130-164) are strongly conserved in p21 and that these regions may represent domains that are especially critical to a functional p21 protein.
Insights
Researchers cloned and sequenced mouse Waf1 (p21) cDNA, finding it
Area of Science:
- Molecular Biology
- Genetics
- Cell Biology
Background:
- The tumor suppressor p53 induces Waf1 (p21), linking growth suppression and cell cycle regulation.
- Understanding p21's role is crucial for cell cycle research.
Purpose of the Study:
- To clone and sequence mouse Waf1 (p21) cDNA.
- To investigate the genetic location and evolutionary conservation of the p21 gene.
Main Methods:
- cDNA cloning and sequencing of mouse p21.
- Southern blot analysis to determine gene copy number and locus.
- Northern blot analysis to assess gene expression.
- Comparative sequence analysis across mouse strains/species and human.
Main Results:
- The entire coding region of mouse p21 cDNA was successfully cloned and sequenced.
- p21 is a single-copy gene located proximal to H-2 on mouse chromosome 17.
- p21 mRNA expression is widespread in normal mouse tissues, with no clear correlation to p53 mRNA levels.
- Two specific regions within the p21 coding sequence (codons 21-60 and 130-164) show strong evolutionary conservation.
Conclusions:
- The mouse p21 gene is confirmed and characterized.
- Conserved regions in p21 likely indicate functionally critical domains for its activity.