Regulatory features of transcription in isolated mitochondria from Artemia franciscana embryos

Abstract

Optimal conditions were developed for an in organello transcriptional run-on assay using mitochondria isolated from Artemia franciscana embryos to investigate potential regulatory features of RNA synthesis under conditions of anoxia-induced quiescence. Transcription is not dependent on oxidative phosphorylation for maximal activity when exogenous ATP is available. Bona fide transcription products, as assessed by hybridization with specific mitochondrial cDNAs from A. franciscana, are produced in an inhibitor-sensitive manner. Transcription rate measured at pH 7.9 is reduced 80% when pH is lowered to 6.3, a pH range that mimics the in vivo change seen on exposure of embryos to anoxia. The proton sensitivity of mitochondrial RNA synthesis may provide a mechanism to depress this significant energy expenditure during quiescence. The influence of nucleotide concentration on kinetics is complicated by an interdependence among nucleotide species. ATP inhibition observed at subsaturating UTP concentrations is relieved when UTP is at saturating, physiologically relevant levels. Taken together, these data suggest that local (versus nuclear mediated) control is important in dictating mitochondrial transcription during rapid modulations in gene expression, such as those observed under anoxia-induced quiescence.