Embryonic alcohol exposure disrupts the ubiquitin-proteasome system
Olivia Weeks, … , Trista E. North, Wolfram Goessling
Olivia Weeks, … , Trista E. North, Wolfram Goessling
Published December 8, 2022
Citation Information: JCI Insight. 2022;7(23):e156914. https://doi.org/10.1172/jci.insight.156914.
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Embryonic alcohol exposure disrupts the ubiquitin-proteasome system

Abstract

Ethanol (EtOH) is a commonly encountered teratogen that can disrupt organ development and lead to fetal alcohol spectrum disorders (FASDs); many mechanisms of developmental toxicity are unknown. Here, we used transcriptomic analysis in an established zebrafish model of embryonic alcohol exposure (EAE) to identify the ubiquitin-proteasome system (UPS) as a critical target of EtOH during development. Surprisingly, EAE alters 20S, 19S, and 11S proteasome gene expression and increases ubiquitylated protein load. EtOH and its metabolite acetaldehyde decrease proteasomal peptidase activity in a cell type–specific manner. Proteasome 20S subunit β 1 (psmb1hi2939Tg) and proteasome 26S subunit, ATPase 6 (psmc6hi3593Tg), genetic KOs define the developmental impact of decreased proteasome function. Importantly, loss of psmb1 or psmc6 results in widespread developmental abnormalities resembling EAE phenotypes, including growth restriction, abnormal craniofacial structure, neurodevelopmental defects, and failed hepatopancreas maturation. Furthermore, pharmacologic inhibition of chymotrypsin-like proteasome activity potentiates the teratogenic effects of EAE on craniofacial structure, the nervous system, and the endoderm. Our studies identify the proteasome as a target of EtOH exposure and signify that UPS disruptions contribute to craniofacial, neurological, and endodermal phenotypes in FASDs.

Authors

Olivia Weeks, Bess M. Miller, Brian J. Pepe-Mooney, Isaac M. Oderberg, Scott H. Freeburg, Colton J. Smith, Trista E. North, Wolfram Goessling

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Figure 6

psmb1 and psmc6 are required for cell survival in the developing brain, spinal cord, and pharyngeal arches.

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psmb1 and psmc6 are required for cell survival in the developing brain,...
(A) Confocal image analysis of acridine orange–stained (AO-stained) and TUNEL-stained psmb1hi2939 (50 hpf, 72 hpf, 96 hpf) mutants reveals increased apoptosis in the brain (top and middle rows, blue and white arrows) and spinal cord (bottom row, pink arrow). psmb1–/– have increased labeling in the pharyngeal arch area (middle row, green arrow). (B and C) ImageJ (NIH) quantification of fluorescent punctae in the brain and spinal cord of mutants following AO staining (****P ≤ 0.0001, unpaired 2-tailed t test; for +/+ vs. +/–, P > 0.05). For columns left to right, n = 10, 5, 6, 2, 8, 5 (B) and n = 9, 5, 8, 2, 9, 7 (C). (D–G) Confocal imaging and quantification of AO-stained larvae (75 hpf). Exposure to BTZ (2 μM, 12–75 hpf) alone resulted in significantly increased staining in the brain. Cotreatment with EtOH and BTZ increased the number of positive dots in the brain, spinal cord, and pharyngeal arch area relative to EtOH and BTZ alone. *P < 0.05, **P < 0.01, ***P < 0.001, ****P ≤ 0.0001, 2-way ANOVA with Tukey’s multiple-comparison test. From left, column sample n = 11, 9, 13, 9 (E); n = 12, 13, 12, 11 (F); and n = 12, 10, 13, 6 (G). Scale bars: 100 μm. Data represent mean ± SD.