Generation of strains for conditional degradation of ER proteins ATF-6 and PEK-1 in Caenorhabditis elegans

• Grant number: 25/03832-4
• Support Opportunities: Scholarships abroad - Research Internship - Doctorate
• Start date: May 01, 2025
• End date: April 30, 2026
• Field of knowledge: Biological Sciences - Biochemistry - Metabolism and Bioenergetics
• Principal Investigator: Fernanda Marques da Cunha
• Grantee: Carolina Gomes de Assis
• Supervisor: Daphne Selvaggia Cabianca
• Host Institution: Escola Paulista de Medicina (EPM). Universidade Federal de São Paulo (UNIFESP). Campus São Paulo. São Paulo , SP, Brazil
• Institution abroad: German Research Center for Environmental Health, Germany
• Associated to the scholarship: 21/05350-6 - The study of the impact of LIPL-5 deficiency on the endoplasmic reticulum UPR and its relationship with the mitochondrial activity of Caenorhabiditis elegans, BP.DR

Abstract

The physical and functional interaction between mitochondria and Endoplasmic Reticulum (ER) is crucial for cellular response to stress, and profoundly impacts the activity of both organelles. During stressful situations, organisms activate adaptive responses, such as the unfolded protein response (UPR), in the attempt to restore homeostasis. Mitochondrial activity of Caenorhabditis elegans changes in response to food deprivation, a nutritional stress. We hypothesized that the UPRer response might be involved in the food deprivation-induced changes in mitochondrial phenotype. To address this question we need animals with impaired activation of each of the UPRer pathways. Working with deletion mutants is straightforward but can mask possible differences since compensatory mechanisms may be activated to overcome the functional absence of important proteins, especially during development. Thus we decided to try a different approach. The Auxin-Inducible Degron (AID) system provides reversible, spatiotemporal control for the knockdown of target proteins, avoiding compensatory mechanisms to be activated. Thus, the aim of this project is to generate C. elegans strains that will allow the depletion of the ER proteins PEK-1 and ATF-6 using the AID system. During the period covered by this proposal, we will design the AID tag knock-in strategy, generate AID-tagged C. elegans via CRISPR/Cas9, introduce Arabidopsis thaliana TIR1 and validate Auxin-Induced Protein Degradation for two strains, one whose target is PEK-1 and the other ATF-6.