The control region is the main non-coding region of vertebrate mitochondrial (mt) DNA, and contains the major regulatory elements controlling replication and transcription of this genome. In many vertebrates, the control region appears to be the fastest evolving locus in the mt genome, and portions of this locus are often used in population and phylogenetics studies. The control region has most extensively been studied in mammals, where it is partitioned into three domains: the two peripheral, highly variable domains I and III, and a central conserved domain II. To better understand the organization and evolutionary dynamics of this locus in vertebrates, we have performed a comparative analysis of control regions from divergent sharks with the mammalian control region. Our analysis reveals an overall similarity in the structure of the control region between sharks and mammals, and identifies several shared conserved elements (TAS, CSBs1-3). Like mammals, variability in shark control regions occurs mainly in peripheral domains I and III, with the central domain II exhibiting significant conservation among species. Domain II from sixteen globally distributed blue sharks reveals no sequence variation. The overall similarity in control region structure between sharks and mammals suggests strong functional constraints control evolution of this locus in vertebrates.