1. A genome-wide CRISPR screen identifies DPM1 as a modifier of DPAGT1 deficiency and ER stress.
- Author
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Dalton, Hans M., Viswanatha, Raghuvir, Brathwaite Jr., Roderick, Zuno, Jae Sophia, Berman, Alexys R., Rushforth, Rebekah, Mohr, Stephanie E., Perrimon, Norbert, and Chow, Clement Y.
- Subjects
CRISPRS ,GENOME editing ,DROSOPHILA melanogaster ,PROTEIN folding ,FRUIT flies ,CONGENITAL disorders - Abstract
Partial loss-of-function mutations in glycosylation pathways underlie a set of rare diseases called Congenital Disorders of Glycosylation (CDGs). In particular, DPAGT1-CDG is caused by mutations in the gene encoding the first step in N-glycosylation, DPAGT1, and this disorder currently lacks effective therapies. To identify potential therapeutic targets for DPAGT1-CDG, we performed CRISPR knockout screens in Drosophila cells for genes associated with better survival and glycoprotein levels under DPAGT1 inhibition. We identified hundreds of candidate genes that may be of therapeutic benefit. Intriguingly, inhibition of the mannosyltransferase Dpm1, or its downstream glycosylation pathways, could rescue two in vivo models of DPAGT1 inhibition and ER stress, even though impairment of these pathways alone usually causes CDGs. While both in vivo models ostensibly cause cellular stress (through DPAGT1 inhibition or a misfolded protein), we found a novel difference in fructose metabolism that may indicate glycolysis as a modulator of DPAGT1-CDG. Our results provide new therapeutic targets for DPAGT1-CDG, include the unique finding of Dpm1-related pathways rescuing DPAGT1 inhibition, and reveal a novel interaction between fructose metabolism and ER stress. Author summary: Sugar modifications are important to proper protein folding and function. The biological pathways that control these sugar modifications are called glycosylation pathways. When these pathways are disrupted, such as through genetic mutation, it can cause rare developmental disorders called Congenital Disorders of Glycosylation (CDGs). Most CDGs lack any treatment options, and little is known about interacting genes or pathways that might improve these disorders. Here, we study DPAGT1-CDG which involves mutations in the enzyme-encoding gene, DPAGT1, by using the fruit fly model organism, Drosophila melanogaster. Using inhibitory drugs, we perform two cell-based screens with CRISPR technology to determine which genes are important under DPAGT1 inhibition. We then used a living fly, that models DPAGT1-CDG in the eye, in order to determine which of the top screened genes can improve its impaired eye development. Surprisingly, we found that inhibiting other CDG-causing genes, such as our top hit Dpm1, could improve our DPAGT1-CDG cell and fly models. In addition, loss of certain metabolism genes, and changes in diet, also improved the DPAGT1-CDG model. This study finds multiple new genes and pathways implicated in DPAGT1-CDG and lays the groundwork for new CDG therapies. [ABSTRACT FROM AUTHOR]
- Published
- 2022
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