Internucleosomal fragmentation and DNA synthesis in wheat seedlings

Internucleosomal fragmentation of nuclear DNA (nDNA) [(180 +/- 10 base pair s)(n), n greater than or equal to 1], characteristic of apoptosis, was obse rved in leaf and coleoptile zones of wheat (Triticum aestivum L.) seedlings , and labeled precursors were found to incorporate rapidly into the "heavy" (rho = 1.716 g/cm(3)) fraction of mitochondrial DNA (hmtDNA). When separat ed by electrophoresis in agarose, the fragments of hmtDNA differed by value s proportional to 195 +/- 10 base pairs and were similar in this aspect to oligonucleosomal fragments produced by apoptic degradation of nuclear DNA. The amount of hmtDNA in older (over 190 h) coleoptiles of wheat seedlings w as comparable to the amount of nDNA (rho = 1.700 g/cm(3)) and reached as hi gh as 50% of the total coleoptile DNA content. In younger (under 144 h) col eoptile, hmtDNA was not detected in CsCl gradients by UV absorption and cou ld be discerned only by its radioactivity. In contrast to nDNA, hmtDNA did not contain 5-methylcytosine and was produced due to unusually active DNA s ynthesis in the mitochondrial fraction. The known inhibitors of DNA synthes is, cycloheximide and ethidium bromide, did not hamper superproduction of h mtDNA; on the contrary, these inhibitors notably stimulated hmtDNA synthesi s. The signal for hmtDNA synthesis and accumulation arrived in the course o f the fifth cycle of synchronous DNA replication in nondividing leaf and co leoptile cells of 135-145-h-old seedlings and apparently did not migrate fu rther along the seedling, hmtDNA superproduction is a characteristic progra mmed event, which is synchronously developed in the senescing cells of inta ct plant organs. The emergence and accumulation of hmtDNA can be used as an indicator of senescence.