Cytosolic HMGB1 controls the cellular autophagy/apoptosis checkpoint during inflammation
Xiaorong Zhu, … , David L. Boone, Eugene B. Chang
Xiaorong Zhu, … , David L. Boone, Eugene B. Chang
Published February 2, 2015
Citation Information: J Clin Invest. 2015;125(3):1098-1110. https://doi.org/10.1172/JCI76344.
View: Text | PDF
Research Article Gastroenterology

Cytosolic HMGB1 controls the cellular autophagy/apoptosis checkpoint during inflammation

Abstract

The intracellular protein HMGB1 is released from cells and acts as a damage-associated molecular pattern molecule during many diseases, including inflammatory bowel disease (IBD); however, the intracellular function of HMGB1 during inflammation is poorly understood. Here, we demonstrated that cytosolic HMGB1 regulates apoptosis by protecting the autophagy proteins beclin 1 and ATG5 from calpain-mediated cleavage during inflammation. Colitis in mice with an intestinal epithelial cell–specific Hmgb1 deletion and patients with IBD were both characterized by increased calpain activation, beclin 1 and ATG5 cleavage, and intestinal epithelial cell (IEC) death compared with controls. In vitro cleavage assays and studies of enteroids verified that HMGB1 protects beclin 1 and ATG5 from calpain-mediated cleavage events that generate proapoptotic protein fragments. Together, our results indicate that HMGB1 is essential for mitigating the extent and severity of inflammation-associated cellular injury by controlling the switch between the proautophagic and proapoptotic functions of beclin 1 and ATG5 during inflammation. Moreover, these studies demonstrate that HMGB1 is pivotal for reducing tissue injury in IBD and other complex inflammatory disorders.

Authors

Xiaorong Zhu, Jeannette S. Messer, Yunwei Wang, Fanfei Lin, Candace M. Cham, Jonathan Chang, Timothy R. Billiar, Michael T. Lotze, David L. Boone, Eugene B. Chang

×

Figure 5

Loss of cell-intrinsic HMGB1 functions leads to IEC death.

Options: View larger image (or click on image) Download as PowerPoint
Loss of cell-intrinsic HMGB1 functions leads to IEC death. IEC progenito...
IEC progenitors were isolated from the small intestines of Hmgb1fl/fl (n = 4) and Vil-Cre Hmgb1fl/fl (n = 4) mice and grown in culture until intestinal enteroids formed. The cells were then treated with 10 μg/ml L-18 MDP and lysed in 1% Triton lysis buffer. (A) Immunoblot for HMGB1 in lysates from cells treated with MDP for 4 hours. (B) Immunoblot for beclin 1 in lysates from cells treated with MDP for 4 hours. (C) Immunoblot for active caspase 3 in lysates from cells treated with MDP for 4 hours. (D) Immunoblot for active caspase 3 in lysates from cells treated with MDP for 4 hours in the presence of DMSO (vehicle control) or 1 μg/ml calpeptin. (E) Immunoblot for LC3B in lysates treated with MDP for 4 hours in the presence or absence of 100 nM bafilomycin A1. (F) Immunoblot for beclin 1 in lysates from cells treated with MDP at the indicated time points. (G) Immunoblot for ATG5 in lysates from cells treated with MDP at the indicated time points. (H) Immunoblot for cleaved caspase 3 in lysates from cells treated with MDP at the indicated time points. (I) Calpain activity in enteroid lysates at the indicated times after MDP treatment. Data were analyzed by 2-way ANOVA with Bonferroni’s multiple comparisons test as well as the 2-tailed Student’s t test for between-genotype comparisons within a treatment group **P < 0.01; ****P < 0.001.