Cloning of cDNAs with possible association with senescence and immortalization of human cells

Abstract

Normal human diploid fibroblasts (HDF) have a finite life span in vitro and have been used as a model system for the study of in vivo aging. Little is known about how changes in gene expression may affect the immortalization of human fibroblasts. We looked for cDNA clones whose mRNAs were differentially expressed between mortal senescent SV40-transformed human fibroblasts (B-32) and the immortal counterparts (B-32F) derived from B-32 cells. We identified three cDNA isolates by subtractive differential hybridization with 32P-labeled cDNA probes from B-32 cells and B-32F cells. Nucleotide sequence analysis of these cDNA clones revealed that they were homologous to the human vimentin, a human mitochondrial gene and a human gene of unknown nature. Slot blot and Northern blot analyses demonstrated that the former two were preferentially expressed in senescent B-32 cells and the last one was less expressed in B-32F immortal cells.