RRM
2009 Rustbelt RNA Meeting
RRM
|
|
|
|
|
Poster abstracts
Abstract:
In eukaryotes, tRNA is transcribed in the nucleus and functions in the cytoplasm. In the yeast Saccharomyces cerevisiae pre-tRNA splicing occurs on cytoplasmic surface of mitochondria and tRNA subcellular movement is not strictly unidirectional from the nucleus to the cytoplasm. However, much remains to be learned about tRNA nuclear-cytoplasmic dynamics. We are using genome-wide analyses and in vivo screening to identify novel gene products involved in tRNA processing and subcellular movement. We are employing Northern analysis to investigate the processing patterns of pre-tRNA from the yeast genome-wide deletion collection, screening for candidates that reproducibly evidence higher molar yield of intron-containing tRNAs relative to mature tRNAs. Such mutants are expected to either have defects in the pre-tRNA processing machinery and/or pre-tRNA subcellular distributions. So far, we identified mutations of mitochondrial and ribosomal protein-encoding genes. In a second approach we are investigating the β-importin family member, Exportin-5 (yeast Msn5), that has been implicated in tRNA subcellular dynamics. Exportin-5/Msn5 functions in nuclear export of micro-RNAs in metazoans and in nuclear export of particular phosphorylated nucleus/cytoplasmic shuttling proteins in yeast. Our data show that Msn5 is unable to export tRNAs in the primary round of export if the tRNAs are encoded by intron-containing tRNA, supporting the model that Msn5 functions solely in the re-export to the cytoplasm. To study the role of Msn5 in tRNA nuclear re-export, we are attempting to identify its binding partner in vivo. One possible binding partner is Tef1/2. TEF1 and TEF2 encode identical eukaryotic translation elongation factor 1 alpha. Tef1/2 bind aminoacyl-tRNAs and function in delivering aa-tRNA to ribosomes. Our studies show that in wild-type strains, Tef1-GFP is excluded from the nucleus, while in msn5Δ cells Tef1-GFP is localized in both cytoplasm and nucleus. This result suggests that Msn5 is responsible for exporting Tef1 from the nucleus and may be involved in the tRNA re-export process. To further investigate the interaction of Tef1 and Msn5, I will determine whether a Msn5•Tef1/2•tRNA•Ran-GTP complex assembles for tRNA re-export.
References:
1. Shaleen, H.H. and Hopper, A.K. (2005) Proc. Natl. Acad. Sci. U.S.A. 102, 11290-11295
2. Takano, A., Endo, T., Yoshihisa T. (2005) Science 309, 140-142
3. Yoshihisa, T., Yunoki-Esaki, K., Ohshima, C., Tanaka, N., Endo, T. (2003) Mol. Biol. Cell 14, 3266-3279
Keywords: tRNA processing
