Export of messenger RNA transcripts (mRNAs) from the nucleus to the cytoplasm is an inherent property of eukaryotic gene expression, however, the mechanism is poorly understood. Recently, the Wente laboratory discovered a physical interaction between the conserved mRNA export factor, Gle1 and the nuclear poly(A)- binding protein, Nab2. The goal of this proposal is to characterize the Gle1:Nab2 complex as a critical, terminal step in mRNA export using the budding yeast S. cerevisiae as a model system. Results gained from this proposal will delineate the first functional role of Gle1 and will be a profound advancement for the field of gene expression.
In aim I, will use classic biochemistry coupled with cutting-edge mass spectrometry to determine the nature of the Gle1:Nab2 complex in yeast as well as the stabiility of this complex in mutant backgrounds. Additionally, I will employ techniques in yeast genetics and cellular microscopy to characterize any newly identified, novel mRNA export factors.
In aim II, I will develop an in vitro assembly system to determine if the Gle1:Nab2 complex associates with RNA and if RNA-binding affinity is altered by addition of other members of the Gle1Nab2 complex, this provides a powerful, in vitro complement to aim I.
