RT Journal
A1 Blumberg, Amit
A1 Rice, Edward J.
A1 Kundaje, Anshul
A1 Danko, Charles G.
A1 Mishmar, Dan
T1 Initiation of mtDNA transcription is followed by pausing, and diverges across human cell types and during evolution
JF Genome Research 
JO Genome Research 
YR 2017 
FD March 01 
VO 27 
IS 3 
SP 362 
OP 373 
DO 10.1101/gr.209924.116 
UL http://genome.cshlp.org/content/27/3/362.abstract 
AB Mitochondrial DNA (mtDNA) genes are long known to be cotranscribed in polycistrones, yet it remains impossible to study nascent mtDNA transcripts quantitatively in vivo using existing tools. To this end, we used deep sequencing (GRO-seq and PRO-seq) and analyzed nascent mtDNA-encoded RNA transcripts in diverse human cell lines and metazoan organisms. Surprisingly, accurate detection of human mtDNA transcription initiation sites (TISs) in the heavy and light strands revealed a novel conserved transcription pausing site near the light-strand TIS. This pausing site correlated with the presence of a bacterial pausing sequence motif, with reduced SNP density, and with a DNase footprinting signal in all tested cells. Its location within conserved sequence block 3 (CSBIII), just upstream of the known transcription–replication transition point, suggests involvement in such transition. Analysis of nonhuman organisms enabled de novo mtDNA sequence assembly, as well as detection of previously unknown mtDNA TIS, pausing, and transcription termination sites with unprecedented accuracy. Whereas mammals (Pan troglodytes, Macaca mulatta, Rattus norvegicus, and Mus musculus) showed a human-like mtDNA transcription pattern, the invertebrate pattern (Drosophila melanogaster and Caenorhabditis elegans) profoundly diverged. Our approach paves the path toward in vivo, quantitative, reference sequence-free analysis of mtDNA transcription in all eukaryotes.