The cis acting sequences responsible for the differential decay of the unstable MFA2 and stable PGK1 transcripts in yeast include the context of the translational start codon.

A general pathway of mRNA turnover has been described for yeast in which the 3' poly(A) tail is first deadenylated to an oligo(A) length, leading to decapping and subsequent 5'-3' exonucleolytic decay. The unstable MFA2 mRNA and the stable PGK1 mRNAs both decay through this pathway, albeit at different rates of deadenylation and decapping. To determine the regions of the mRNAs that are responsible for these differences, we examined the decay of chimeric mRNAs derived from the 5' untranslated, coding, and 3' untranslated regions of these two mRNAs.

Isolation and characterization of Dcp1p, the yeast mRNA decapping enzyme.

A major mechanism of mRNA decay occurs by the process of deadenylation, decapping and 5' --> 3' exonucleolytic degradation. Recently, the product of the DCP1 gene has been shown to be required for decapping mRNAs in vivo and co-purifies with decapping activity in vitro. We have purified Dcp1p to homogeneity and shown that it is sufficient for decapping, thereby indicating that Dcp1p is the decapping enzyme.

An essential component of the decapping enzyme required for normal rates of mRNA turnover.

A major pathway of messenger RNA degradation in eukaryotic cells is initiated by shortening of the poly(A) tail, which, at least in yeast, triggers a decapping reaction, thereby exposing the mRNA to 5' --> 3' degradation. Decapping is the key step in this decay pathway because the transcript body is rapidly degraded following decapping. Accordingly, decapping is the site of numerous controls, including inhibition of decapping by the poly(A) tail and modulation of mRNA decapping rate by specific sequences.

mRNA decapping activities and their biological roles.

The 5' cap structure of eukaryotic mRNAs is significant for a variety of cellular events and also serves to protect mRNAs from premature degradation. Analysis of mRNA decay in Saccharomyces cerevisiae has shown that removal of the 5' cap structure is a key step in the turnover of many yeast mRNAs, and that this decapping is carried out by Dcp1p. In addition to the yeast decapping enzyme, other activities that can cleave the 5' cap structure have been described.

Phylogenetic comparative chemical footprint analysis of the interaction between ribonuclease P RNA and tRNA.

Ribonuclease P RNA is the catalytic moiety of the ribonucleoprotein enzyme that endonucleolytically cleaves precursor sequences from the 5' ends of pre-tRNAs. The bacterial RNase P RNA-tRNA complex was examined with a footprinting approach, utilizing chemical modification to determine RNase P RNA nucleotides that potentially contact tRNA. RNase P RNA was modified with dimethylsulfate or kethoxal in the presence or absence of tRNA, and sites of modification were detected by primer extension.