Excessive EFHD1-dependent ER-mitochondrial contacts drive a maladaptive antiviral response in metabolic liver disease
David R. Eberhardt, Emma C. Rekate, Yasmin B. Masini, Hannah E. Duron, David Mollinedo, Adrian M. Velarde, Devorah Stucki, Tara R. Price, Sandra H.J. Lee, Enrique Balderas, Neeraj K. Rai, Ashley R. Bratt, Anthony M. Balynas, Chris J. Stubben, Ryan Bia, Sudipa Maity, Nicolas Hartel, Xue Yin, Andrea Corbin, Anshu Kumari, Dung M. Nguyen, Daisuke Shimura, Vu D. Nguyen, Vishaka Vinod, Kamrul H. Chowdhury, Francisco Verdeguer, Joel Zvick, Patrice N. Mimche, Sihem Boudina, Stavros G. Drakos, Ademuyiwa S. Aromolaran, Sarah Franklin, Vivek Garg, Robin M. Shaw, William L. Holland, Scott A. Summers, Marcus G. Pezzolesi, Jared Rutter, Kimberley J. Evason, Dipayan Chaudhuri
David R. Eberhardt, Emma C. Rekate, Yasmin B. Masini, Hannah E. Duron, David Mollinedo, Adrian M. Velarde, Devorah Stucki, Tara R. Price, Sandra H.J. Lee, Enrique Balderas, Neeraj K. Rai, Ashley R. Bratt, Anthony M. Balynas, Chris J. Stubben, Ryan Bia, Sudipa Maity, Nicolas Hartel, Xue Yin, Andrea Corbin, Anshu Kumari, Dung M. Nguyen, Daisuke Shimura, Vu D. Nguyen, Vishaka Vinod, Kamrul H. Chowdhury, Francisco Verdeguer, Joel Zvick, Patrice N. Mimche, Sihem Boudina, Stavros G. Drakos, Ademuyiwa S. Aromolaran, Sarah Franklin, Vivek Garg, Robin M. Shaw, William L. Holland, Scott A. Summers, Marcus G. Pezzolesi, Jared Rutter, Kimberley J. Evason, Dipayan Chaudhuri
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Research Article Cell biology Hepatology Metabolism

Excessive EFHD1-dependent ER-mitochondrial contacts drive a maladaptive antiviral response in metabolic liver disease

Abstract

Metabolic dysfunction–associated steatohepatitis (MASH) involves hepatocyte damage that cannot be explained solely by lipid accumulation. Here, to discover injury-specific pathways, we focused on a gene of uncertain function, EF-hand domain family member D1 (EFHD1), identified in human genome-wide association studies of liver injury but not liver fat. We showed that EFHD1, a Ca2+-dependent actin cross-linker, stabilizes endoplasmic reticulum–mitochondria contact sites (ERMCS) by detecting spatiotemporal coincidence of interorganellar proximity and ER Ca2+ release. During MASH, EFHD1 upregulation drives pathological mitochondrial fragmentation via excessive contact persistence. This structural failure promotes mitochondrial double-stranded RNA escape and activation of a maladaptive, antiviral, protein kinase RNA-activated–associated stress response, a causal relationship also supported by Mendelian randomization in humans. Consequently, inhibiting EFHD1 in human and mouse models blunts hepatocyte damage. These findings identify EFHD1 as a Ca2+-dependent ERMCS stabilizer, reveal a hepatocyte-intrinsic injury pathway, and suggest EFHD1 inhibition as a therapeutic strategy.

Authors

David R. Eberhardt, Emma C. Rekate, Yasmin B. Masini, Hannah E. Duron, David Mollinedo, Adrian M. Velarde, Devorah Stucki, Tara R. Price, Sandra H.J. Lee, Enrique Balderas, Neeraj K. Rai, Ashley R. Bratt, Anthony M. Balynas, Chris J. Stubben, Ryan Bia, Sudipa Maity, Nicolas Hartel, Xue Yin, Andrea Corbin, Anshu Kumari, Dung M. Nguyen, Daisuke Shimura, Vu D. Nguyen, Vishaka Vinod, Kamrul H. Chowdhury, Francisco Verdeguer, Joel Zvick, Patrice N. Mimche, Sihem Boudina, Stavros G. Drakos, Ademuyiwa S. Aromolaran, Sarah Franklin, Vivek Garg, Robin M. Shaw, William L. Holland, Scott A. Summers, Marcus G. Pezzolesi, Jared Rutter, Kimberley J. Evason, Dipayan Chaudhuri

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Figure 5

Excessive ERMCS stabilization triggers pathological mt-dsRNA release and a PKR-dependent ISR in MASH.

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Excessive ERMCS stabilization triggers pathological mt-dsRNA release and...
(A and B) Western blot (A) and summary (B). EIF2α, eukaryotic translation initiation factor 2-α; p-EIF2α, phosphorylated EIF2α; PKR, protein kinase RNA-activated; p-PKR, phosphorylated PKR; PERK, PKR-like endoplasmic reticulum kinase/eukaryotic translation initiation factor 2-alpha kinase 3 (EIF2AK3); p-PERK, phosphorylated PERK. (C and D) MASH diet Western blot (C) and band quantification (D). (E and F) Representative images (E) and quantification summary (F) of dsRNA immunohistochemistry in human liver. (G) Isolated mouse hepatocyte immunohistochemistry. (H) Representative high-contrast immunohistochemistry of human livers. Arrowheads, PKR in dense nonhepatocyte nuclei; arrows, dsRNA- and PKR-stained cytoplasmic signal. (I) Representative images, isolated mouse hepatocytes. (J–M) Quantitative RT-PCR following immunoprecipitation of dsRNA (J and K) or PKR (L and M) from isolated mouse hepatocytes. (N) Representative immunocytochemistry of isolated hepatocytes from MASH diet mice. Because less dsRNA was evident in Efhd1–/– hepatocytes, image acquisition settings for these were adjusted to maximize dsRNA signal. Bottom, inset at higher magnification. (O) Average dsRNA-DRP1 distance per hepatocyte. (P) Penalized and robust inverse variance-weighted MR analyses support a causal relationship between human PKR (EIF2AK2) and liver disease, using AST as the exposure. *, 2 distributions with non-overlapping 95% confidence intervals. t test: B and D. Mann-Whitney U test: B, J–M, and O. Linear mixed-effect model: F. N values, figure panel in parentheses. Chow: 3–6(B); MASH: 5–6(B). WT: 6(D); Efhd1–/–: 6–7(D). Stage 0, 1, 2–3: 6(F); Stage 4: 5(F). Chow: 8(J), 5(L); MASH: 6(J), 5(L). WT: 6(K), 5(M); Efhd1–/–: 8(K), 7(M). WT: 49(O); Efhd1–/–: 19(O). Bar: mean ± SEM.