In disease states, unfolded proteins accumulate in the endoplasmic reticulum because the folding capacity is exceeded, initiating a cellular stress response (the unfolded protein response or UPR). Our long-term interest is to understand the molecular mechanisms that allow cells to withstand stress and that contribute to pathologies during prolonged stress. Regulation of gene expression by transcriptional activators and repressors is a key feature of the stress response. During the previous grant period we found that the adaptive response to nutrient starvation increases expression of amino acid transporter genes, which can facilitate the recovery from stress. We also found that transcription of the arginine/lysine transporter gene, Cat-1, and of other genes involved in amino acid metabolism is attenuated during prolonged ER stress, mediated by the CCAAT/enhancer binding protein family transcription factor, C/EBP2. Regulated translation of the C/EBP2 mRNA produces both LAP, a transcriptional activator, and LIP a repressor. The LAP/LIP ratio plays a critical role in cell fate and metabolism. We found that the LAP/LIP ratios change during the UPR via mechanisms that involve proteasomal degradation of the proteins and translational control of the C/EBP2 mRNA;this provides the driving force behind this proposal. The regulation of transcription factor levels during ER stress via the proteasome pathway is a novel mechanism to modulate the cellular stress response. We hypothesize that the LAP/LIP ratio plays a role in controlling transcription of stress-response genes. We also hypothesize that the regulation of LIP levels promotes expression of prosurvival genes early in the stress response and restricts expression of proapoptotic genes during prolonged stress. In this proposal, we will study the mechanisms that regulate LIP synthesis and degradation during ER stress in cultured cells and in mice. Experiments using stress-inducing drugs and models of human disease will reveal the physiological significance of this regulation.
Our Specific Aims are: (i) Determine the mechanism for diminished LIP levels during the early (prosurvival) phase of ER stress (ii) Investigate the signaling pathways that regulate proteasome-mediated degradation of LIP during the prosurvival phase of ER stress. (iii) Determine the mechanisms that increase LIP levels during the late (proapoptotic) phase of ER stress. (iv) Determine the physiological significance of the LAP/LIP ratio during ER stress using MEFs defficient in C/EBP2 (v) Determine the effect of disruption of the C/EBP2 gene in animal models of ER stress-mediated disease. Our long term goal is to generate transgenic mice expressing only LAP and only LIP by knock-in mutations in the C/EBP2 gene, using the state of the art system of Bacterial Artificial Chromosomes. The knock-in mice will be a valuable tool to determining the functions of LAP and LIP in ER- stress mediated apoptosis and enable us to test the findings of Aims 1-5 in a physiological context with relevance to human disease. Cellular stress is important in a large number of diseases, such as diabetes, neurodegeneration, cancer and complications of obesity. A common feature of these diseases is the accumulation of damaged secretory proteins in the endoplasmic reticulum (ER), a vital organelle responsible for proper cellular function and metabolism. During the previous grant period we found that the adaptive response to nutrient starvation increases expression of amino acid transporter genes, which can facilitate the recovery from stress. PROJECT
This proposal will study C/EBP2, an important regulator of the stress response and how this regulator controls the balance between cellular survival and death. Our studies will generate new therapeutic targets for the many stress-mediated diseases and provide a novel mechanism that regulates the balance between survival and death during these diseases.
|Davuluri, Gangarao; Krokowski, Dawid; Guan, Bo-Jhih et al. (2016) Metabolic adaptation of skeletal muscle to hyperammonemia drives the beneficial effects of l-leucine in cirrhosis. J Hepatol 65:929-937|
|Hao, Yujun; Samuels, Yardena; Li, Qingling et al. (2016) Oncogenic PIK3CA mutations reprogram glutamine metabolism in colorectal cancer. Nat Commun 7:11971|
|Kenche, Harshavardhan; Ye, Zhi-Wei; Vedagiri, Kokilavani et al. (2016) Adverse Outcomes Associated with Cigarette Smoke Radicals Related to Damage to Protein-disulfide Isomerase. J Biol Chem 291:4763-78|
|Hsu, K-S; Guan, B-J; Cheng, X et al. (2016) Translational control of PML contributes to TNFÎ±-induced apoptosis of MCF7 breast cancer cells and decreased angiogenesis in HUVECs. Cell Death Differ 23:469-83|
|Saikia, Mridusmita; Hatzoglou, Maria (2015) The Many Virtues of tRNA-derived Stress-induced RNAs (tiRNAs): Discovering Novel Mechanisms of Stress Response and Effect on Human Health. J Biol Chem 290:29761-8|
|Han, Dong-Yun; Guan, Bo-Jhih; Wang, Ya-Juan et al. (2015) L-type Calcium Channel Blockers Enhance Trafficking and Function of Epilepsy-associated Î±1(D219N) Subunits of GABA(A) Receptors. ACS Chem Biol 10:2135-48|
|Komar, Anton A; Hatzoglou, Maria (2015) Exploring Internal Ribosome Entry Sites as Therapeutic Targets. Front Oncol 5:233|
|Majumder, Mithu; Mitchell, Daniel; Merkulov, Sergei et al. (2015) Residues required for phosphorylation of translation initiation factor eIF2Î± under diverse stress conditions are divergent between yeast and human. Int J Biochem Cell Biol 59:135-41|
|Schuster, Andrew T; Homer, Craig R; Kemp, Jacqueline R et al. (2015) Chromosome-associated protein D3 promotes bacterial clearance in human intestinal epithelial cells by repressing expression of amino acid transporters. Gastroenterology 148:1405-16.e3|
|Gao, Xing-Huang; Krokowski, Dawid; Guan, Bo-Jhih et al. (2015) Quantitative H2S-mediated protein sulfhydration reveals metabolic reprogramming during the integrated stress response. Elife 4:e10067|
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