Cloning of senescent cell-derived inhibitors of DNA synthesis using an expression screen

Asao Noda, Yi Ning, Susan F. Venable, Olivia M. Pereira-Smith, James R. Smith

Research output: Contribution to journalArticle

Abstract

We here describe a rapid and simple expression screen method that has allowed us to isolate cDNAs coding for inhibitors of DNA synthesis from senescent human diploid fibroblasts. The assay involved transient transcriptional overexpression of a gene product encoded by a cDNA in a proliferating cell, on the assumption that this would be sufficient to block DNA synthesis in a short-term assay using tritiated thymidine autoradiography. Three cDNAs, referred to as senescent cell-derived inhibitors (sdi), that exhibit DNA synthesis-inhibitory activity when introduced into young cycling cells, were successfully identified. Expression of one of the cDNAs, sdi1, increased 10- to 20fold in senescent compared with young cells and the increase in RNA closely paralleled the onset of the senescent phenotype and loss of cell proliferation. sdi1 expression was also increased in young cells made nondividing (quiescent) by deprivation of growth factors or contact inhibition. Following serum stimulation, RNA levels of sdi1 in quiescent cells were initially increased, but then declined to low levels just prior to the entry of the cells into S phase. In contrast, RNA levels of sdi1 in senescent cells failed to decline, suggesting a role for this gene in maintaining the senescent phenotype. The sdi1 gene has been mapped to the p arm of chromosome 6.

Original languageEnglish (US)
Pages (from-to)90-98
Number of pages9
JournalExperimental Cell Research
Volume211
Issue number1
DOIs
StatePublished - 1994
Externally publishedYes

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Nucleic Acid Synthesis Inhibitors
Organism Cloning
Complementary DNA
RNA
Contact Inhibition
Genes
Phenotype
Chromosomes, Human, Pair 6
Autoradiography
Diploidy
S Phase
Thymidine
Intercellular Signaling Peptides and Proteins
Fibroblasts
Cell Proliferation
DNA

ASJC Scopus subject areas

  • Cell Biology

Cite this

Cloning of senescent cell-derived inhibitors of DNA synthesis using an expression screen. / Noda, Asao; Ning, Yi; Venable, Susan F.; Pereira-Smith, Olivia M.; Smith, James R.

In: Experimental Cell Research, Vol. 211, No. 1, 1994, p. 90-98.

Research output: Contribution to journalArticle

Noda, Asao ; Ning, Yi ; Venable, Susan F. ; Pereira-Smith, Olivia M. ; Smith, James R. / Cloning of senescent cell-derived inhibitors of DNA synthesis using an expression screen. In: Experimental Cell Research. 1994 ; Vol. 211, No. 1. pp. 90-98.
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