Capping of mRNA occurs shortly after transcription initiation, preceding other mRNA processing events such as mRNA splicing and polyadenylation. To determine the mechanism of coupling between transcription and capping, we tested for a physical interaction between capping enzyme and the transcription machinery. Capping enzyme is not stably associated with basal transcription factors or the RNA polymerase II (Pol II) holoenzyme. However, capping enzyme can directly and specifically interact with the phosphorylated form of the RNA polymerase carboxy-terminal domain (CTD). This association occurs in the context of the transcription initiation complex and is blocked by the CTD-kinase inhibitor H8. Furthermore, conditional truncation mutants of the Pol II CTD are lethal when combined with a capping enzyme mutant. Our results provide in vitro and in vivo evidence that capping enzyme is recruited to the transcription complex via phosphorylation of the RNA polymerase CTD.

mRNA capping requires the sequential action of three enzymatic activities: RNA triphosphatase, guanylyl-transferase, and methyltransferase. Here we characterize a gene (CEL-1) believed to encode the C. elegans capping enzyme. CEL-1 has a C-terminal domain containing motifs found in yeast and vaccinia virus capping enzyme guanylyltransferases. The N-terminal domain of CEL-1 has RNA triphosphatase activity. Surprisingly, this domain does not resemble the vaccinia virus capping enzyme but does have significant sequence similarity to the protein tyrosine phosphatase (PTP) enzyme family. However, CEL-1 has no detectable PTP activity. The mechanism of the RNA triphosphatase is similar to that of PTPs: the active site contains a conserved nucleophilic cysteine required for activity. These results broaden the superfamily of PTP-like phosphatases to include enzymes with RNA substrates.

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