THE GUANYLYLTRANSFERASE DOMAIN OF MAMMALIAN MESSENGER-RNA CAPPING ENZYME BINDS TO THE PHOSPHORYLATED CARBOXYL-TERMINAL DOMAIN OF RNA-POLYMERASE-II

We have conducted a biochemical and genetic analysis of mouse mRNA cap ping enzyme (Mce1), a bifunctional 597-amino acid protein with RNA tri phosphatase and RNA guanylyltransferase activities. The principal conc lusions are as follows: (i) the mammalian capping enzyme consists of a utonomous and nonoverlapping functional domains; (ii) the guanylyltran sferase domain Mce1(211-597) is catalytically active in vitro and func tional in vivo in yeast in lieu of the endogenous guanylyltransferase Ceg1; (iii) the guanylyltransferase domain per se binds to the phospho rylated RNA polymerase II carboxyl-terminal domain (CTD), whereas the triphosphatase domain, Mce1(1-210), does not bind to the CTD; and (iv) a mutation of the active site cysteine of the mouse triphosphatase el icits a strong growth-suppressive phenotype in yeast, conceivably by s equestering pre-mRNA ends in a nonproductive complex or by blocking ac cess of the endogenous yeast triphosphatase to RNA polymerase II. Thes e findings contribute to an emerging model of mRNA biogenesis wherein RNA processing enzymes are targeted to nascent polymerase II transcrip ts through contacts with the CTD, The phosphorylation-dependent intera ction between guanylyltransferase and the CTD is conserved from yeast to mammals.