Age-related changes in DNA polymerase α expression

DNA polymerase α isozymes differing in specific activity and affinity of binding to DNA were purified from human fibroblasts derived from donors of different ages. Fetal-derived fibroblasts expressed a single, high-activity enzyme (A₂), with high affinity of binding to DNA. Adult-derived fibroblasts exhibited two forms of DNA polymerase α, one identical to the fetal enzyme, and a second with about tenfold less activity showing low affinity of binding to DNA (A₁). The ratio of DNA polymerase A₂/A₁ decreased dramatically with age from 100° A₂ in fetal-derived fibroblasts to about 94° A₁ in fibroblasts derived from a 66-year-old donor. The DNA binding affinity of polymerase α A₁ from adult-derived fibroblasts increased concominant with a significant increase in activity when the enzyme was treated with phosphatidylinositol-4-monophosphate (PIP), or with inositol-1, 4-bisphosphate (I(1,4)P₂). The enzyme reverted back to a less active form, with loss of the noncovalently bound I(1,4)P₂, as a function of time. When permeabilized human fibroblasts with low DNA excision repair capacity were treated with 7,8-dihydrodiol-9,10-epoxybenzo(a)-pyrene (BPDE) in the presence of ³²P-ATP, phosphatidylinositol, and cycloheximide, excision repair was initiated and ³²P-labeled DNA polymerase α was recovered in the absence of de novo protein synthesis. DNA synthesis associated with either scheduled DNA synthesis or BPDE-initiated excision repair declined as a function of increased age in human cells. The data suggest that the decline in both DNA excision repair-associated and mitogen-activated DNA synthesis may be correlated with decreased total intracellular levels of DNA polymerase and with the decline in polymerase α activity as a function of age, that DNA repair-associated initiation of DNA synthesis in adult-derived cells may increase with activation of a pool of low activity DNA polymerase α, and that DNA polymerase α activity increases as a function of enzyme interaction with a component of the PI phosphorylation cascade.