HeLa cell RNA (2‘-O-methyladenosine-N6-)-methyltransferase specific for the capped 5‘-end of messenger RNA.

PCIF1 Catalyzes m6Am mRNA Methylation to Regulate Gene Expression

“…If this first nucleotide is 2-O-methyladenosine (Am), it can be further methylated at its N6 position to generate m6Am (Figure 1A). While the methylase mediating Am methylation is known, the methyltransferase responsible for generating m6Am has not been identified (Keith et al, 1978;Mauer et al, 2017). As a consequence, the impact of m6Am on gene regulation is still largely unexplored.…”

The Mammalian Cap-Specific m6Am RNA Methyltransferase PCIF1 Regulates Transcript Levels in Mouse Tissues

“…Unlike the internal m 6 A marks that have been shown to mediate RNA decay by the action of YTH proteins (Ke et al, 2017;Wang et al, 2014), the cap-specific m 6 Am on the TSS adenosine is demonstrated to promote RNA stability by providing resistance to the action of the mRNA decapping enzyme DCP2 (Mauer et al, 2017). An activity responsible for cap-specific m 6 Am methylation was partially purified as ã 65-kDa protein from human HeLa cell extracts (Keith et al, 1978), and this was recently revealed to be Phosphorylated CTD-Interacting Factor 1 (PCIF1) (Akichika et al, 2019;Boulias et al, 2019;Sendinc et al, 2019;Sun et al, 2019). PCIF1 was originally identified as a factor interacting with the phosphorylated CTD of RNA Pol II by its N-terminal WW domain (Fan et al, 2003;Hirose et al, 2008) and was later shown to have an affinity for the Ser5-phosphorylated CTD (Akichika et al, 2019).…”

A comprehensive review of m6A/m6Am RNA methyltransferase structures

“…The co-crystallized cap-mimicking m 7 Gpp entity is sandwiched in between the HD and MTD and bound by residues from both the HD and the MTD (Figure 4B ,C). Activity studies suggest that CAPAM identifies the capped substrate by specifically recognizing the N7-methylation on m 7 G ( 46 , 47 , 54 , 97 ), but oddly, the structure reveals no residues that specifically recognize this modification (Figure 4B ,C). CAPAM does not methylate internal Am ( 46 , 97 ) which is likely a combination of cap-specific recognition where removal of this element abrogates MTase activity ( 52 ) and steric hindrance disallowing a mRNA with a sequence 5′ of the methylation site to be accommodated on CAPAM.…”

“…m 6 Am (Figure 1B ) has, since its discovery, been identified as an internal modification in mRNA ( 43 ) and snRNA ( 44 , 45 ) and is found at the 5′-end of most eukaryotic mRNA ( 2 , 46 ), where the modified Am ( 46 , 47 ) is positioned adjacent to the mRNA cap structure (reviewed in ( 48 )) (Figure 1A ). The cellular abundance of the N6-methylation at the mRNA cap is context dependent (reviewed in ( 49 )).…”

“…CAPAM prefers long substrates ( 47 , 52 ) and the large HD could accommodate such extended substrates. The enzyme shows no sequence specificity ( 46 , 47 ) and the recognition of these long substrates is therefore likely mainly via the RNA backbone rather than with nucleobases. The preference for longer substrates agrees with that capping of nascent mRNA occurs when these are at least 20 nts of length ( 99 ).…”