Enzymatic Characterization of In Vitro Activity of RNA Methyltransferase PCIF1 on DNA.

PCIF1 and FTO are a pair of human mRNA cap-specific modification enzymes that have opposing activities. PCIF1 adds a methyl group to the N6-position of 2' O -methyladenosine (A m ), generating N6, 2' O -dimethyladenosine (m 6 A m ), when A m is the cap-proximal nucleotide. FTO removes the N6-methyl group from m 6 A m . In addition, FTO has a demethylase activity on a broad spectrum of various RNA substrates, as well as on D NA N6-methyldeoxyadenosine (m 6 dA). While the existence of m 6 dA in mammalian D NA remains controversial, we show here that PCIF1 has significant methylation activity on single stranded DNA deoxyadenosine, double stranded RNA/DNA hybrids, and double stranded DNA, though with lower catalytic efficiency than that on its preferred RNA substrate. PCIF1 has activities in the order ssRNA > RNA/DNA hybrid > ssDNA > dsDNA. We discuss the implications of PCIF1 generation, and FTO removal, of DNA adenine methylation.