Parallel CRISPR-Cas9 screens identify mechanisms of PLIN2 and lipid droplet regulation.

Roberts MA, Deol KK, Mathiowetz AJ, Lange M, Leto DE, Stevenson J, Hashemi SH, Morgens DW, Easter E, Heydari K

Dev Cell 2023

1. Dev Cell. 2023 Sep 25;58(18):1782-1800.e10. doi: 10.1016/j.devcel.2023.07.001. Epub 2023 Jul 25. Parallel CRISPR-Cas9 screens identify mechanisms of PLIN2 and lipid droplet regulation. Roberts MA(1), Deol KK(1), Mathiowetz AJ(1), Lange M(1), Leto DE(2), Stevenson J(3), Hashemi SH(4), Morgens DW(5), Easter E(1), Heydari K(6), Nalls MA(7), Bassik MC(8), Kampmann M(9), Kopito RR(2), Faghri F(7), Olzmann JA(10). Author information: (1)Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA; Department of Nutritional Sciences and Toxicology, University of California, Berkeley, Berkeley, CA 94720, USA. (2)Department of Biology, Stanford University, Stanford, CA 94305, USA. (3)Department of Nutritional Sciences and Toxicology, University of California, Berkeley, Berkeley, CA 94720, USA. (4)Department of Computer Science, University of Illinois at Urbana-Champaign, Urbana, IL 61820, USA. (5)Department of Plant and Microbial Biology, University of California, Berkeley, Berkeley, CA 94720, USA. (6)Cancer Research Laboratory FACS Core Facility, University of California, Berkeley, Berkeley, CA 94720, USA. (7)Data Tecnica International, LLC, Washington, DC, USA; Center for Alzheimer's and Related Dementias, National Institutes of Health, Bethesda, MD 20892, USA; Laboratory of Neurogenetics, National Institute on Aging, National Institutes of Health, Bethesda, MD 20892, USA. (8)Department of Genetics, Stanford University, Stanford, CA 94305, USA. (9)Institute for Neurodegenerative Diseases, Department of Biochemistry and Biophysics, University of California, San Francisco, San Francisco, CA 94158, USA. (10)Department of Molecular and Cell Biology, University of California, Berkeley, Berkeley, CA 94720, USA; Department of Nutritional Sciences and Toxicology, University of California, Berkeley, Berkeley, CA 94720, USA; Chan Zuckerberg Biohub, San Francisco, CA 94158, USA. Electronic address: olzmann@berkeley.edu. Despite the key roles of perilipin-2 (PLIN2) in governing lipid droplet (LD) metabolism, the mechanisms that regulate PLIN2 levels remain incompletely understood. Here, we leverage a set of genome-edited human PLIN2 reporter cell lines in a series of CRISPR-Cas9 loss-of-function screens, identifying genetic modifiers that influence PLIN2 expression and post-translational stability under different metabolic conditions and in different cell types. These regulators include canonical genes that control lipid metabolism as well as genes involved in ubiquitination, transcription, and mitochondrial function. We further demonstrate a role for the E3 ligase MARCH6 in regulating triacylglycerol biosynthesis, thereby influencing LD abundance and PLIN2 stability. Finally, our CRISPR screens and several published screens provide the foundation for CRISPRlipid (http://crisprlipid.org), an online data commons for lipid-related functional genomics data. Our study identifies mechanisms of PLIN2 and LD regulation and provides an extensive resource for the exploration of LD biology and lipid metabolism. Copyright © 2023 The Authors. Published by Elsevier Inc. All rights reserved. DOI: 10.1016/j.devcel.2023.07.001 PMCID: PMC10530302 PMID: 37494933 [Indexed for MEDLINE] Conflict of interest statement: Declaration of interests J.A.O. is a member of the scientific advisory board for Vicinitas Therapeutics and has patent applications related to ferroptosis. M.K. is an inventor on a US patent related to CRISPRi and CRISPRa screening; serves on the scientific advisory boards of Engine Biosciences, Casma Therapeutics, Cajal Neuroscience and Alector; and is a consultant to Modulo Bio and Recursion Therapeutics. F.F. and M.A.N.’s participation in this project was part of a competitive contract awarded to Data Tecnica International LLC by the National Institutes of Health to support open science research. M.A.N. also currently serves on the scientific advisory board for Clover Therapeutics and is an advisor to Neuron23 Inc. M.C.B. has outside interest in DEM Biopharma.