Identification of single nucleotide polymorphisms in the p21 (CDKN1A) gene and correlations with longevity in the Italian population

Abstract

Longevity in humans is determined by multiple environmental and genetic factors. We have investigated possible associations between longevity and Single Nucleotide Polymorphisms (SNPs) in the p21 (CDKN1A) gene, a stress-inducible senescence-associated cell cycle inhibitor, expression of which upregulates genes implicated in several age-related diseases. By sequencing the promoter and exons of p21 in genomic DNA of ten individuals over 90 years old, we have identified 30 SNPs, many of which had not been previously characterized. A cluster of minor alleles within the -4547/-3489 bp region did not alter the basal activity or p53 responsiveness of the p21 promoter. We then compared the frequency of 41 p21 SNPs between 184 centenarians and 184 younger subjects in the Italian population. Rare alleles of two exon-derived SNPs, rs1801270 and rs1059234, were significantly under-represented among the centenarians; no significant differences were found for 39 non-exonic SNPs. SNP rs1801270 causes Ser to Arg substitution at amino acid 31 and SNP rs1059234 leads to a nucleotide change in the 3'-untranslated region. Previous studies showed that the rare alleles of these two SNPs may play a role in cancer. These p21 alleles may be potentially detrimental to longevity and therefore are rare in centenarians.

Keywords: CDKN1A; longevity; single nucleotide polymorphisms.

Figure 1.

Activities of the p21 promoter-luciferase constructs with the common (p21C-luc; black bars) or rare (p21R-luc; grey bars) allele SNP cluster in the -4547/-3489 bp region. (A) Three independent plasmid preparations of each of p21C-luc or p21R-Luc (R) were transfected into HCT116 wild type (WT) cells. Cells were harvested 48 h after transfection, and firefly luciferase activity was measured and normalized to Renilla luciferase expressed from a co-transfected construct. The bars show mean and standard deviation for triplicate transfections. (B) p21C-luc and p21R-luc plasmids were transfected in parallel into HCT116 WT and p53-/- cell lines, in triplicates as in A. The bars show mean and standard deviation of the ratio of normalized luciferase activities achieved with the same plasmid in the WT relative to p53-/- cells.

Figure 2.. Haplotype blocks distrubution in the p21 gene generated by Haploview.

The two SNPs showing significant differences in frequency between the centenarians and younger controls are bracketed. Every multimarker combination within this block including the two SNPs is significant on the omnibus test for frequency distribution among cases and controls. Table 3 gives the results of the haplotype test.