structure-function analysis of rny1 in trna cleavage and growth inhibition结构分析的rny1 trna乳沟和抑制增长.pdf

Structure-Function Analysis of Rny1 in tRNA Cleavage and Growth Inhibition 1 2 Natalie Luhtala , Roy Parker * 1 Cancer Biology Graduate Interdisciplinary Program, University of Arizona, Tucson, Arizona, United States of America, 2 Howard Hughes Medical Institute and Department of Molecular and Cellular Biology, University of Arizona, Tucson, Arizona, United States of America Abstract T2 ribonucleases are conserved nucleases that affect a variety of processes in eukaryotic cells including the regulation of self-incompatibility by S-RNases in plants, modulation of host immune cell responses by viral and schistosome T2 enzymes, and neurological development and tumor progression in humans. These roles for RNaseT2’s can be due to catalytic or catalytic-independent functions of the molecule. Despite this broad importance, the features of RNaseT2 proteins that modulate catalytic and catalytic-independent functions are poorly understood. Herein, we analyze the features of Rny1 in Saccharomyces cerevisiae to determine the requirements for cleaving tRNA in vivo and for inhibiting cellular growth in a catalytic-independent manner. We demonstrate that catalytic-independent inhibition of growth is a combinatorial property of the protein and is affected by a fungal-specific C-terminal extension, the conserved catalytic core, and the presence of a signal peptide. Catalytic functions of Rny1 are independent of the C-terminal extension, are affected by many mutations in the catalytic core, and also require a signal peptide. Biochemical flotation assays reveal that in rny1D cells, some tRNA molecules associate with membranes suggesting that cleavage of tRNAs by Rny1 can involve either tRNA association with, or uptake into, membrane compartments. Citation: Luhtala N, Parker R (2012) Structure