MNK-driven eIF4E phosphorylation regulates the fibrogenic transformation of mesenchymal cells and chronic lung allograft dysfunction
Natalie M. Walker, … , Amanda L. Garner, Vibha N. Lama
Natalie M. Walker, … , Amanda L. Garner, Vibha N. Lama
Published August 15, 2024
Citation Information: J Clin Invest. 2024;134(16):e168393. https://doi.org/10.1172/JCI168393.
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MNK-driven eIF4E phosphorylation regulates the fibrogenic transformation of mesenchymal cells and chronic lung allograft dysfunction

Abstract

Tissue fibrosis remains unamenable to meaningful therapeutic interventions and is the primary cause of chronic graft failure after organ transplantation. Eukaryotic translation initiation factor (eIF4E), a key translational regulator, serves as convergent target of multiple upstream profibrotic signaling pathways that contribute to mesenchymal cell (MC) activation. Here, we investigate the role of MAP kinase–interacting serine/threonine kinase–induced (MNK-induced) direct phosphorylation of eIF4E at serine 209 (Ser209) in maintaining fibrotic transformation of MCs and determine the contribution of the MNK/eIF4E pathway to the pathogenesis of chronic lung allograft dysfunction (CLAD). MCs from patients with CLAD demonstrated constitutively higher eIF4E phosphorylation at Ser209, and eIF4E phospho-Ser209 was found to be critical in regulating key fibrogenic protein autotaxin, leading to sustained β-catenin activation and profibrotic functions of CLAD MCs. MNK1 signaling was upregulated in CLAD MCs, and genetic or pharmacologic targeting of MNK1 activity inhibited eIF4E phospho-Ser209 and profibrotic functions of CLAD MCs in vitro. Treatment with an MNK1/2 inhibitor (eFT-508) abrogated allograft fibrosis in an orthotopic murine lung-transplant model. Together these studies identify what we believe is a previously unrecognized MNK/eIF4E/ATX/β-catenin signaling pathway of fibrotic transformation of MCs and present the first evidence, to our knowledge, for the utility of MNK inhibitors in fibrosis.

Authors

Natalie M. Walker, Yuta Ibuki, A. Patrick McLinden, Keizo Misumi, Dylan C. Mitchell, Gabriel G. Kleer, Alison M. Lock, Ragini Vittal, Nahum Sonenberg, Amanda L. Garner, Vibha N. Lama

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Figure 1

Phosphorylation of eIF4E (Ser209) is MNK1 dependent but mTORC-independent in fibrotic MCs derived from lung-transplant patients.

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Phosphorylation of eIF4E (Ser209) is MNK1 dependent but mTORC-independen...
(A) Protein expression of phosphorylated and total forms of eIF4E and MNK1 was measured by Western blot analysis in MCs derived from normal (non-Fib-MCs) or fibrotic (Fib-MCs) human-lung allografts. Densitometry analyses of phospho-eIF4E to total eIF4E are shown with Fib-MCs for RAS (circles) and BOS (triangles). (B) Lentiviral infections were performed using empty pLenti-LoxEV vector or vector expressing active MNK1 (T344D) in nonfibrotic MCs or kinase-dead MNK1 (D191A) in Fib-MCs. Western blotting and corresponding densitometry analyses were performed for phospho-eIF4E and total eIF4E. (C) Fibrotic MCs treated with eFT-508 (10 μM, 24 hours) were subjected to m7GDP cap pulldown assay followed by Western blotting analyses. (D) Fib-MCs were transfected with RPTOR-specific or scrambled siRNA, and protein lysates were analyzed by Western blotting. Representative immunoblots and corresponding densitometry are shown for phospho-eIF4E and total eIF4E. (E and F) Fib-MCs were treated with ATP-competitive mTORC inhibitors (AZD8055: 250 nM; rapamycin: 250 nM) for 24 hours and analyzed for phospho-eIF4E and total eIF4E by Western blotting and densitometry. (G) Fib-MCs were transfected with MNK1-specific or scrambled siRNA. Protein lysates were immunoblotted for mTORC1/2 substrates. Representative immunoblots and corresponding densitometry are shown for phosphorylated and total forms of 4E-BP1, p70S6K1, and AKT. Data are represented as means ± SEM. **P < 0.01; ***P < 0.001; ****P < 0.0001, unpaired t test.