Project Summary Cellular stress-responsive mechanisms are essential for cells to adapt to various environmental and physiological conditions. The overall goal of the research program is to understand how cells regulate mRNA translation as part of their stress adaptation process. Of particular interest is a pathway referred to as the Integrated Stress Response (ISR), which is initiated by stress-activated eIF2a kinases that respond to several distinct types of stress. Abnormal regulation of ISR is associated with various metabolic and neurodegenerative diseases, including ribosomopathies caused by heterozygosity in ribosome subunit genes and Charcot Marie Tooth Disease caused by certain tRNA synthetase mutations. The ISR signaling mechanism is intriguing because this pathway induces stress-responsive gene transcription, and coincidentally, suppresses general mRNA translation. ISR inhibits mRNA translation at multiple levels, including the specific inhibition of translation initiation factors, eIF2a and eIF-4E. In addition, recent evidence indicates that ribosome stalling on mRNAs is associated with ISR. These observations raise a fundamental question regarding how stress-responsive transcripts overcome these multiple translational blocks, and in some cases, increase their translation as part of ISR signaling. To address this, we propose to use the facile genetic tools of Drosophila. The ISR regulatory mechanisms are conserved in this organism, and there are genetic mutations that serve as models for human diseases with abnormal ISR signaling. Our preliminary genetic screen in Drosophila has identified several poorly characterized translational regulators as factors required for ISR signaling. Building on these observations, we will test the central hypothesis that the newly identified factors specifically mediate the translation of stress-responsive transcripts, thereby helping those mRNAs to bypass translational blocks imposed by ISR. We will further determine the physiological significances of the newly identified translational regulatory mechanisms in the Drosophila models of ribosomopathies and Charcot Marie Tooth Disease. A successful realization of the proposed plan will advance our conceptual understanding of stress-responsive gene expression, and help develop new therapeutic strategies against diseases associated with ISR.