MITOCHONDRIAL POLYNUCLEOTIDE DEGRADATION AS AN EARLY STAGE MARKER OF MAMMALIAN STRESS RESPONSE

D.R. Crawford
Center for Immunology and Microbial Disease, The Albany Medical College, Albany, New York 12208

Oxidative stress affects biological cells in many ways including the modulation of gene expression. We have found that a calcium-dependent down-regulation of mitochondrial RNA occurs when HA-1 hamster fibroblasts are exposed to hydrogen peroxide. This effect is specific to mitochondrial transcripts as 21 cytosolic mRNAs exhibited little or no down-regulation. Subsequent analysis determined that this down-regulation also occurs with mitochondrial genomic DNA and is the apparent result of DNA degradation.  Early stage degradation of genomic DNA, assessed within the first five hours of peroxide exposure, was specific to mitochondria; nuclear genomic DNA was not degraded under the same treatment conditions.  As with mitochondrial RNA, mitochondrial DNA degradation was calcium dependent. Degradation of both mitochondrial polynucleotides occurred at similar time points and was specific to transcripts derived from mitochondrial-encoded genes. Mitochondrial polynucleotide degradations occurred under conditions where extensive growth-arrest and moderate apoptosis were observed, and were accompanied by significant induction of the growth arrest mRNAs gadd45, gadd153 and adapt15/gadd7 as well as morphological nuclear fragmentation, cell shrinkage, and internucleosomal DNA fragmentation. Conversely, minimal, if any, degradation of cytosolic glyceraldehyde-3-phosphate dehydrogenase or actin mRNAs was observed.  Similar results were obtained for HA-1 cells treated with the protein kinase inhibitor staurosporine, and for HT-2 mouse T-lymphocytes induced to undergo apoptosis by interleukin-2 withdrawal. Mitochondrial 16S rRNA degradation was an early event that was discernable well before chromatin condensation in hydrogen peroxide-treated HA-1 cells. Combined, these results indicate that there is a general degradation of mitochondrial- but not cytosolic or nuclear-polynucleotides during early stage stress response in mammalian cells, and that these changes are associated with growth arrest and apoptosis. 

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