The overall goal of our studies is to understand the regulation, function, and disease association of the keratin intermediate filament (IF) cytoskeletal proteins in digestive organs. Keratins 8 and 18 (K8/K18) are the IFs of hepatocytes and their major function is cytoprotection from mechanical and nonmechanical stresses such as apoptosis. This function is consistent with the finding that specific K8 and K18 genetic variants predispose to liver disease progression. K8 and K18 are also the major constituents of Mallory body (MB) inclusions that are found in association with some liver diseases. Several transgenic animal studies demonstrated that K8 and a K8-greater-than-K18 protein ratio, coupled with a drug insult are all essential in order for MBs to form. MBs can be rapidly re-induced once initially formed but the mechanism of such rapid predisposition to reformation is unknown. Our proposal includes 4 aims to examine the pathogenesis of MBs. The first three aims test the hypothesis that keratin transamidation at specific amino acids is essential for MB formation. The 4th aim tests the hypothesis that chaperone function is altered during MB (re)formation. The proposed studies utilize cell culture and mouse MB models to help understand their pathogenesis and, ultimately, their importance in human disease. The 4 aims are: (i) Study MB formation in transglutaminase-2 (TG2) null and control mice using established in vivo models.
This aim i s based on our preliminary findings that TG2-null mice have a markedly blunted ability to form MBs. (ii) Identify keratin transamidation sites and their crosslinked partners in vivo/vitro and study their role in crosslinking and inclusion body formation in cell culture. This is based on the findings that K8/K18 are excellent TG2 substrates (K8>K18) in vitro, (iii) Generate transgenic mice that express transamidation-mutant keratins and test the effect of the mutations on MB formation and susceptibility to liver injury, (iv) Test the effect of MB-inducing injury on chaperone function and the potential role of chaperones as """"""""memory proteins"""""""" in association with MB re-accumulation. This is based on our recent findings that MB formation correlates with chaperone dysfunction. Our proposed studies are likely to generate important new biologic and clinically relevant information regarding the pathogenesis of MBs. We anticipate that completion of this project will shed light on the specific keratin amino acids that allow MB formation and address if MBs are bystanders, protectors or promoters of injury. Our findings should also shed light on whether chaperone dysfunction can provide 'molecular memory"""""""" and as such contribute to rapid MB reformation. Our proposal represents a direct approach to fully understand the pathogenesis and significance of MBs that were first described in 1911 by Dr. Frank Mallory, and may also impact on other inclusions that are found in association with several other neurological and neuromuscular human diseases whose pathogenesis is unknown.

Agency
National Institute of Health (NIH)
Institute
National Institute of Diabetes and Digestive and Kidney Diseases (NIDDK)
Type
Research Project (R01)
Project #
5R01DK052951-15
Application #
8113450
Study Section
Special Emphasis Panel (ZRG1-DIG-C (02))
Program Officer
Serrano, Jose
Project Start
1997-09-01
Project End
2012-06-30
Budget Start
2011-07-01
Budget End
2012-06-30
Support Year
15
Fiscal Year
2011
Total Cost
$338,892
Indirect Cost
Name
University of Michigan Ann Arbor
Department
Physiology
Type
Schools of Medicine
DUNS #
073133571
City
Ann Arbor
State
MI
Country
United States
Zip Code
48109
Omary, M Bishr (2017) Intermediate filament proteins of digestive organs: physiology and pathophysiology. Am J Physiol Gastrointest Liver Physiol 312:G628-G634
Zhang, Deqiang; Tong, Xin; VanDommelen, Kyle et al. (2017) Lipogenic transcription factor ChREBP mediates fructose-induced metabolic adaptations to prevent hepatotoxicity. J Clin Invest 127:2855-2867
Ajluni, Nevin; Meral, Rasimcan; Neidert, Adam H et al. (2017) Spectrum of disease associated with partial lipodystrophy: lessons from a trial cohort. Clin Endocrinol (Oxf) 86:698-707
Ku, Nam-On; Strnad, Pavel; Bantel, Heike et al. (2016) Keratins: Biomarkers and modulators of apoptotic and necrotic cell death in the liver. Hepatology 64:966-76
Snider, Natasha T; Portney, Daniel A; Willcockson, Helen H et al. (2016) Ethanol and Acetaminophen Synergistically Induce Hepatic Aggregation and TCH346-Insensitive Nuclear Translocation of GAPDH. PLoS One 11:e0160982
Elenbaas, Jared S; Maitra, Dhiman; Liu, Yang et al. (2016) A precursor-inducible zebrafish model of acute protoporphyria with hepatic protein aggregation and multiorganelle stress. FASEB J 30:1798-810
Weerasinghe, Sujith V W; Park, Min-Jung; Portney, Daniel A et al. (2016) Mouse genetic background contributes to hepatocyte susceptibility to Fas-mediated apoptosis. Mol Biol Cell 27:3005-3012
Gilbert, St├ęphane; Loranger, Anne; Omary, M Bishr et al. (2016) Keratin impact on PKC?- and ASMase-mediated regulation of hepatocyte lipid raft size - implication for FasR-associated apoptosis. J Cell Sci 129:3262-73
Snider, Natasha T; Omary, M Bishr (2016) Assays for Posttranslational Modifications of Intermediate Filament Proteins. Methods Enzymol 568:113-38
Sun, Jingyuan; Groppi, Vincent E; Gui, Honglian et al. (2016) High-Throughput Screening for Drugs that Modulate Intermediate Filament Proteins. Methods Enzymol 568:163-85

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