The role of the cap and poly(A) tail in mRNA metabolism
Most eukaryotic mRNAs possess a 7-methyl-guanosine at their 5' ends (5' cap) and a poly(A) tail at their 3' ends. These structures play important roles in gene expression, for example, it is believed that an mRNA must possess a poly(A) tail and possibly a 5' cap to leave the nucleus and both of these structures play important roles in translation and mRNA stability. Much of what we know about the roles of the 5' cap and 3' poly(A) tail results from experiments using cell extracts, exogenous mRNA electroporated into cells, or analyses performed with very unhealthy mutant cells, all of which may not imitate in vivo conditions. To circumvent these problems, I have made reporter constructs that are transcribed in a wild type yeast cell and then cleaved, resulting in an unadenylated or uncapped cleavage product. I have demonstrated that 5' unadenylated cleavage products can be translated but are rapidly targeted to the cytoplasmic exosome for degradation. In accordance with a 5' cap being required for translation and mRNS stability, uncapped 3' cleavage products are unstable and are not translated. This approach was then used to test the proposed requirement of the poly(A) tail in recognition of premature termination codons by the nonsense-mediated mRNA decay pathway. Contrary to what was believed, unadenylated nonsense mRNAs are still recognized as nonsense. This indicates that the distance between a premature termination codon and the poly(A) tail is not required for nonsense-mediated decay in yeast. Another set of studies has used reporter mRNAs that contain cleavage signals for the nuclear endonuclease, Rnt1p. These constructs have been used to study the requirement of the 5' cap and poly(A) tail for mRNA export. Using these reporters, I have demonstrated that the 5' cap and poly(A) tail are not required for export. This work has provided new insights into many aspects of gene expression. Furthermore, the constructs that I have described should be useful for more detailed studies of the roles of the 5' cap and poly(A) tail in all aspects of mRNA metabolism.
Meaux, Stacie Ann, "The role of the cap and poly(A) tail in mRNA metabolism" (2008). Texas Medical Center Dissertations (via ProQuest). AAI3305170.