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Mammalian genes coordinately regulated by growth arrest signals and DNA-damaging agents
A J Fornace1, D W Nebert, M C Hollander
1Radiation Oncology Branch, National Cancer Institute, Bethesda, Maryland 20892.
Abstract:
More than 20 different cDNA clones encoding DNA-damage-inducible transcripts in rodent cells have recently been isolated by hybridization subtraction (A. J. Fornace, Jr., I. Alamo, Jr., and M. C. Hollander, Proc. Natl. Acad. Sci. USA 85:8800-8804, 1988). In most cells, one effect of DNA damage is the transient inhibition of DNA synthesis and cell growth. We now show that five of our clones encode transcripts that are increased by other growth cessation signals: growth arrest by serum reduction, medium depletion, contact inhibition, or a 24-h exposure to hydroxyurea. The genes coding for these transcripts have been designated gadd (growth arrest and DNA damage inducible). Two of the gadd cDNA clones were found to hybridize at high stringency to transcripts from human cells that were induced after growth cessation signals or treatment with DNA-damaging agents, which indicates that these responses have been conserved during mammalian evolution. In contrast to results with growth-arrested cells that still had the capacity to grow after removal of the growth arrest conditions, no induction occurred in HL60 cells when growth arrest was produced by terminal differentiation, indicating that only certain kinds of growth cessation signals induce these genes. All of our experiments suggest that the gadd genes are coordinately regulated: the kinetics of induction for all five transcripts were similar; in addition, overexpression of gadd genes was found in homozygous deletion c14CoS/c14CoS mice that are missing a small portion of chromosome 7, suggesting that a trans-acting factor encoded by a gene in this deleted portion is a negative effector of the gadd genes. The gadd genes may represent part of a novel regulatory pathway involved in the negative control of mammalian cell growth.
Insights
Five growth arrest and DNA damage inducible (gadd) genes are induced by various growth cessation signals and DNA-damaging agents in rodent and human cells. These gadd genes are coordinately regulated and may be involved in controlling mammalian cell growth.
Area of Science:
- Molecular Biology
- Cell Biology
- Genetics
Background:
- DNA damage triggers cellular responses, including transient inhibition of DNA synthesis and cell growth.
- Previously, over 20 DNA-damage-inducible transcripts were identified in rodent cells.
- The regulation and function of these damage-inducible genes are not fully understood.
Purpose of the Study:
- To investigate whether DNA-damage-inducible transcripts respond to other growth cessation signals.
- To identify and characterize genes involved in both DNA damage and growth arrest responses.
- To explore the evolutionary conservation and regulatory mechanisms of these inducible genes.
Main Methods:
- Hybridization subtraction to isolate cDNA clones.
- Northern blot analysis to detect transcript induction.
- Analysis of gene expression in response to various growth arrest conditions (serum reduction, medium depletion, contact inhibition, hydroxyurea).
- Cross-species hybridization to assess evolutionary conservation.
- Examination of gene expression in HL60 cells undergoing terminal differentiation.
- Analysis of gadd gene expression in mice with chromosomal deletions.
Main Results:
- Five cDNA clones, designated gadd (growth arrest and DNA damage inducible), encode transcripts induced by both DNA-damaging agents and various growth cessation signals.
- Two gadd cDNA clones hybridized to transcripts in human cells induced by similar signals, indicating conserved regulation.
- gadd gene induction was not observed in HL60 cells undergoing terminal differentiation, suggesting signal specificity.
- The gadd genes appear to be coordinately regulated, with similar induction kinetics.
- Overexpression of gadd genes in mice with a specific chromosome 7 deletion suggests a trans-acting negative regulator.
Conclusions:
- The gadd genes represent a novel class of genes induced by both DNA damage and diverse growth arrest conditions.
- The conserved induction of gadd genes across mammalian species highlights their importance in cellular regulation.
- gadd gene regulation is specific to certain types of growth cessation signals.
- Evidence suggests a coordinated regulatory pathway for gadd genes, potentially involving a negative effector encoded on chromosome 7.
- The gadd genes may play a role in a novel regulatory pathway for negative control of mammalian cell growth.