Culjkovic-Kraljacic B, Skrabanek L, Revuelta MV, Gasiorek J, Cowling VH, Cerchietti L, Borden KLB.

Proc Natl Acad Sci U S A. 2020 Oct 27;117(43):26773-26783. doi:10.1073/pnas.2002360117

The eukaryotic translation initiation factor eIF4E elevates steady-state m7G capping of coding and noncoding transcripts.

Some RNAs have more than half of their molecules non-capped. Increasing expression of eIF4E increases their capped proportion to similar levels as other genes. Enzymatic (cap quantification; CapQ) and quantitative cap immunoprecipitation (CapIP) methods. Uses the mouse IgG2aκ anti-7-methylguanosine (m7G)-Cap antibody of MBL (RN016M). The maker's site notes that “This antibody (Clone 150-15) reacts with 5′-terminal 7-methylguanosine (m7G) cap structure of RNA and partially cross-reacts with m7G within RNA”. Overexpression of eIF4E-Flag in RNA immunoprecipitation experiments increased enrichment of RNA export targets such as RNMT or RNGTT, but export-deficient S53A-eIF4E-Flag mutants did not. Overexpression also shifted RNA translation to heavier polysomal fractions and elevated m7G levels in the nuclear and cytoplasmic compartments. This elevation is reduced by RNMT knockdown and stronger on specific target, for instance from the capping machinery and the Myc pathway. A CapQ assay (sequencing an oligo-capping library and a control PNK library) confirmed the observations made by immunoprecipitation. In control conditions, the capping rate of the targets of eIF4E capping are lower than those of the non-targets.