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

Melissa A Roberts; Kirandeep K Deol; Alyssa J Mathiowetz; Mike Lange; Dara E Leto; Julian Stevenson; Sayed Hadi Hashemi; David W Morgens; Emilee Easter; Kartoosh Heydari; Mike A Nalls; Michael C Bassik; Martin Kampmann; Ron R Kopito; Faraz Faghri; James A Olzmann

doi:10.1016/j.devcel.2023.07.001

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

Dev Cell. 2023 Sep 25;58(18):1782-1800.e10. doi: 10.1016/j.devcel.2023.07.001. Epub 2023 Jul 25.

Authors

Melissa A Roberts¹, Kirandeep K Deol¹, Alyssa J Mathiowetz¹, Mike Lange¹, Dara E Leto², Julian Stevenson³, Sayed Hadi Hashemi⁴, David W Morgens⁵, Emilee Easter¹, Kartoosh Heydari⁶, Mike A Nalls⁷, Michael C Bassik⁸, Martin Kampmann⁹, Ron R Kopito², Faraz Faghri⁷, James A Olzmann¹⁰

Affiliations

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.

Abstract

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.

Keywords: CRISPR; ERAD; MARCH6; PLIN2; endoplasmic reticulum; genetic screen; lipid droplet; metabolism; perilipin; resource.

Publication types

Research Support, N.I.H., Intramural
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't

MeSH terms

CRISPR-Cas Systems* / genetics
Cell Line
Humans
Lipid Droplets* / metabolism
Lipid Metabolism / genetics
Perilipin-2 / genetics
Perilipin-2 / metabolism

Substances

Perilipin-2
PLIN2 protein, human

Abstract

Publication types

MeSH terms

Substances

Grants and funding