The first step in RNA processing is the addition of an inverted GMP nucleotide, called a 'cap', onto the 5' end of the first transcribed nucleotide. In the nucleus this is bound by CBP80, which interacts with the processing machinery to coordinate intron splicing and addition of the poly(A) tail at the 3' end. The newly-processed mRNA is exported through the nuclear pore complex cap-end first, where CBP80 is replaced by eIF4E to begin the process of its translation into protein. The cap is removed as one of the first steps in the overall process of mRNA decay, and it was generally thought that loss of the cap results in rapid degradation by an exonuclease acting on the unprotected end.
In the dogma of molecular biology cap addition only occurs in the nucleus and its loss in the cytoplasm is irreversible. There are numerous reasons why this made sense, the most compelling of which is the concentration of the responsible protein (capping enzyme( in the nucleus and the biochemistry of cap addition, which requires a substrate with 2 phosphate groups, not the single phosphate that is left after the cap is removed. I will present work from my lab describing a new mechanism by which the cap can be restored onto cytoplasmic mRNAs after it has been removed by decapping or endonuclease cleavage. This work began with re-examination of results published in 1992 and never followed up describing a cap or cap-like structure on decay products of ÃŸ-globin mRNA in patients with ÃŸ-thalassemia (Cooley's anemia), a fatal disorder of hemoglobin production that is caused by inheriting two copies of this gene with a premature termination codon. I will describe how we validated those results, some of the basic biochemistry behind the re-capping process, and the identification and properties of a cytoplasmic complex that contains the enzymes that are responsible for mRNA re-capping. The loss of the cap is one of the key steps by which microRNAs repress translation and silence gene expression, and my talk will cover the cycle by which cytoplasmic re-capping may function in re-activating these silenced mRNAs. I will also touch on the possible links between cytoplasmic capping and the activation of neuronal or maternal mRNAs that must be kept in a silenced state until their translation is required. Although at this point it is highly speculative, cytoplasmic capping may also expand the proteome by enabling the translation of different forms of a protein from mRNAs that have lost the cap and sequences from their 5' ends, and the challenges the complexity of this process presents for bioinformatics, molecular and cell biology.