IN ORGANELLO FOOTPRINT ANALYSIS OF HUMAN MITOCHONDRIAL-DNA - HUMAN MITOCHONDRIAL TRANSCRIPTION FACTOR-A INTERACTIONS AT THE ORIGIN OF REPLICATION

Using in organello footprint analysis, we demonstrate that within huma n placental mitochondria there is a high level of protein-DNA binding at regularly phased intervals throughout a 500-bp region encompassing the D-loop DNA origins and two promoter regions. Comparison with in vi tro DNase I protection studies indicates that this protein-DNA interac tion is due to non-sequence-specific binding by human mitochondrial tr anscription factor A (h-mtTFA). Since h-mtTFA can bend and wrap DNA, l ike its yeast counterpart ABF2, a primary function of h-mtTFA appears to be specific packaging of the mitochondrial DNA control region in vi vo. Intervals of protein binding coincide with the spacing of the RNA start sites and prominent D-loop DNA 5' ends, suggesting a role for ph ased h-mtTFA binding in defining transcription and H-strand DNA replic ation origins. Significant protein-DNA interaction was also observed w ithin the human homolog of conserved sequence block 1, both in organel lo and in vitro, using purified h-mtTFA.