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Chronic ethanol feeding affects proteasome-interacting proteins.

ABSTRACT: Studies on alcoholic liver injury mechanisms show a significant inhibition of the proteasome activity. To investigate this phenomenon, we isolated proteasome complexes from the liver of rats fed ethanol chronically, and from the liver of their pair-fed controls, using a non-denaturing multiple centrifugations procedure to preserve proteasome-interacting proteins (PIPs). ICAT and MS/MS spectral counting, further confirmed by Western blot, showed that the levels of several PIPs were significantly decreased in the isolated ethanol proteasome fractions. This was the case of PA28alpha/beta proteasome activator subunits, and of three proteasome-associated deubiquitinases, Rpn11, ubiquitin C-terminal hydrolase 14, and ubiquitin carboxyl-terminal hydrolase L5. Interestingly, Rpn13 C-terminal end was missing in the ethanol proteasome fraction, which probably altered the linking of ubiquitin carboxyl-terminal hydrolase L5 to the proteasome. 20S proteasome and most 19S subunits were however not changed but Ecm29, a protein known to stabilize the interactions between the 20S and its activators, was decreased in the isolated ethanol proteasome fractions. It is proposed that ethanol metabolism causes proteasome inhibition by several mechanisms, including by altering PIPs and proteasome regulatory complexes binding to the proteasome.

SUBMITTER: Bousquet-Dubouch MP

PROVIDER: S-EPMC2766596 | biostudies-literature | 2009 Jul

REPOSITORIES: biostudies-literature

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Chronic ethanol feeding affects proteasome-interacting proteins.

Bousquet-Dubouch Marie-Pierre MP Nguen Sheila S Bouyssié David D Burlet-Schiltz Odile O French Samuel W SW Monsarrat Bernard B Bardag-Gorce Fawzia F

Proteomics 20090701 13

Studies on alcoholic liver injury mechanisms show a significant inhibition of the proteasome activity. To investigate this phenomenon, we isolated proteasome complexes from the liver of rats fed ethanol chronically, and from the liver of their pair-fed controls, using a non-denaturing multiple centrifugations procedure to preserve proteasome-interacting proteins (PIPs). ICAT and MS/MS spectral counting, further confirmed by Western blot, showed that the levels of several PIPs were significantly ...[more]

PMID: 19609968

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Project description:Bone loss in response to alcohol intake has previously been hypothesized to be mediated by excessive production of reactive oxygen species via NADPH oxidase (Nox) enzymes. Nox4 is one of several Nox enzymes expressed in bone. We investigated the role of Nox4 in the chondro-osteoblastic lineage of the long bones in mice during normal chow feeding and during chronic ethanol feeding for 90 days. We generated mice with a genotype (PrxCre +/- Nox4 fl/fl) allowing conditional knockout of Nox4 in the limb bud mesenchyme. Adult mice had 95% knockdown of Nox4 expression in the femoral shafts. For mice on regular chow, only whole-body Nox4 knockout mice had clearly increased cortical thickness and bone mineral density in the tibiae. When chronically fed a liquid diet with and without ethanol, conditional Nox4 knockout mice had slightly reduced dimensions of the cortical and trabecular regions of the tibiae (P < 0.1). The ethanol diet caused a significant reduction in cortical bone area and cortical thickness relative to a control diet without ethanol (P < 0.05). The ethanol diet further reduced gene expression of Frizzled related protein (Frzb), myosin heavy chain 3, and several genes encoding collagen and other major structural bone proteins (P < 0.05), whereas the Nox4 genotype had no effects on these genes. In conclusion, Nox4 expression from both mesenchymal and nonmesenchymal cell lineages appears to exert subtle effects on bone. However, chronic ethanol feeding reduces cortical bone mass and cortical gene expression of major structural bone proteins in a Nox4-independent manner. SIGNIFICANCE STATEMENT: Excessive alcohol intake contributes to osteopenia and osteoporosis, with oxidative stress caused by the activity of NADPH oxidases hypothesized to be a mediator. We tested the role of NADPH oxidase (Nox) 4 in osteoblast precursors in the long bones of mice with a conditional Nox4 knockout model. We found that Nox4 exerted effects independent of alcohol intake, and ethanol effects on bone were Nox4-independent.

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Chronic ethanol feeding affects proteasome-interacting proteins.

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Chronic ethanol feeding affects proteasome-interacting proteins.

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