Precise sequence assignment of replication origin in the control region of chick mitochondrial DNA relative to 5' and 3' D-loop ends, secondary structure, DNA synthesis, and protein binding.

The data reported identify for the first time the sequence of an avian mitochondrial heavy-strand replication origin, OH, located only about 12 nucleotides (nt) downstream from the conserved sequence block CSB-1, as well as the sequence of premature synthesis arrest of the 781 (+/-1) nt D-loop strand, only 6-7 nt downstream from a TAS-like (termination-associated) element. Both sites are associated with putative cruciform secondary structures. A major sequence-specific DNA-binding/cleavage site of a potential regulatory protein, the approximately 36-kDa aMDP1 (shown previously to stimulate mtDNA synthesis), is located about 90 nt upstream of OH. Correlated in vivo analysis of avian genome-length mtDNA replication provides missing evidence on the functional equivalence of D-loop origin with nascent initiation, and on the direction, asymmetry and temporal aspects of a full round of replication. The importance of the results to understanding the regulation of linked replication/transcription and the unusual sequence evolution of avian mtDNA is