“Toxic memory” via chaperone modification is a potential mechanism for rapid mallory-denk body reinduction

Abstract

The cytoplasmic hepatocyte inclusions, Mallory-Denk bodies (MDBs), are characteristic of several liver disorders, including alcoholic and nonalcoholic steatohepatitis. In mice, MDBs can be induced by long-term feeding with 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) for 3 to 4 months or rapidly reformed in DDC-induced then recovered mice by DDC refeeding or exposure to a wide range of toxins for only 5 to 7 days. The molecular basis for such a rapid reinduction of MDBs is unknown. We hypothesized that protein changes retained after DDC priming contribute to the rapid MDB reappearance and associate with MDB formation in general terms. Two-dimensional differential-in-gel-electrophoresis coupled with mass spectrometry were used to characterize protein changes in livers from the various treatment groups. The alterations were assessed by real-time reverse-transcription polymerase chain reaction and confirmed by immunoblotting. DDC treatment led to pronounced charged isoform changes in several chaperone families, including Hsp25, 60, 70, GRP58, GRP75, and GRP78, which lasted at least for 1 month after discontinuation of DDC feeding, whereas changes in other proteins normalized during recovery. DDC feeding also resulted in altered expression of Hsp72, GRP75, and Hsp25 and in functional impairment of Hsp60 and Hsp70 as determined using a protein complex formation and release assay. The priming toward rapid MDB reinduction lasts for at least 3 months after DDC discontinuation, but becomes weaker after prolonged recovery. MDB reinduction parallels the rapid increase in p62 and Hsp25 levels as well as keratin 8 cross-linking that is normally associated with MDB formation. Conclusion: Persistent posttranslational modifications in chaperone proteins, coupled with protein cross-linking and altered chaperone expression and function likely contribute to the "toxic memory" of DDC-primed mice. We hypothesize that similar changes are important contributors to inclusion body formation in several diseases. (HEPATOLOGY 2008;48:931-942.)

Toward unraveling the complexity of simple epithelial keratins in human disease

“…Last, proteasome inhibition promotes, but does not directly cause, mouse MDB formation in vivo (101). Such inhibition can mimic proteasome overutilization when coupled with a decrease in chaperone levels and function (102) and is consistent with the observation that MDB formation is paralleled by increased protein misfolding and β-sheet formation (103,104).…”

“…However, there is a toxic memory that leads to rapid MDB reformation in mice that have recovered after DDC or griseofulvin feeding (termed primed mice) when they are exposed for 5-7 days to any one of a variety of hepatotoxins that normally do not induce MDBs (13). One likely cause of this memory is a long-term loss of chaperone function that requires a long recovery period, possibly as a result of slow turnover of modified and functionally altered stress proteins (102).…”

Costaining for keratins 8/18 plus ubiquitin improves detection of hepatocyte injury in nonalcoholic fatty liver disease

“…These non-ballooned, but dysmorphic, hepatocytes typically localize near each other and in close proximity to KBH in zone 3 or zone 1, suggesting that all of these aberrant morphologies reflect similar types of cellular stress. Given that ubiquitinated K8 and K18 are the major constituents of Mallory-Denk bodies (14), the fact that Mallory body formation has been mechanistically linked with defective protein turnover and ER stress (15, 16), and other evidence that ballooned hepatocytes exhibit markers of the ER stress response (17), ER stress is the most likely culprit for the loss of K8/18 and accumulation of ubiquitinated proteins in non-ballooned hepatocytes in patients with NAFLD. Further support for this concept derives from our observation that the numbers of KBH, KH, and Ub in NAFLD patients increase in parallel with HOMA-IR scores, because a cause-effect relationship between ER stress and insulin resistance has already been established.…”

The ubiquitin ligase CHIP/STUB1 targets mutant keratins for degradation

“…Such protein aggregates closely resemble those found in vimentin-induced cataracts, in chronic liver disorders, AD, PD, and several other neurodegenerative disorders Bornheim et al, 2008;Strnad et al, 2008]. Here, we demonstrate for the first time that the stress-induced Hsp70 and Hsp90 were upregulated in neonatal K5 À/À mice and, upon proteasome inhibition, in a cell culture model of EBS at the RNA and protein level.…”