4E-BP1 counteracts human mesenchymal stem cell senescence via maintaining mitochondrial homeostasis

Abstract

Although the mTOR-4E-BP1 signaling pathway is implicated in aging and aging-related disorders, the role of 4E-BP1 in regulating human stem cell homeostasis remains largely unknown. Here, we report that the expression of 4E-BP1 decreases along with the senescence of human mesenchymal stem cells (hMSCs). Genetic inactivation of 4E-BP1 in hMSCs compromises mitochondrial respiration, increases mitochondrial reactive oxygen species (ROS) production, and accelerates cellular senescence. Mechanistically, the absence of 4E-BP1 destabilizes proteins in mitochondrial respiration complexes, especially several key subunits of complex III including UQCRC2. Ectopic expression of 4E-BP1 attenuates mitochondrial abnormalities and alleviates cellular senescence in 4E-BP1- deficient hMSCs as well as in physiologically aged hMSCs. These findings together demonstrate that 4E-BP1 functions as a geroprotector to mitigate human stem cell senescence and maintain mitochondrial homeostasis, particularly for the mitochondrial respiration complex III, thus providing a new potential target to counteract human stem cell senescence.

Keywords

4E-BP1 / mitochondria / aging

References

[1] Andrade J, Potente M. Endothelial metabolism-more complex (III) than previously thought. Nat Metab 2019; 1: 14- 5.

[2] Balaban RS, Nemoto S, Finkel , T. . Mitochondria, oxidants, and aging. Cell 2005; 120: 483- 95.

[3] Bi S, Liu Z, Wu Z et al. SIRT7 antagonizes human stem cell aging as a heterochromatin stabilizer. Protein cell 2020; 11: 483- 504.

[4] Bianco P. “Mesenchymal” stem cells. Annu Rev Cell Dev Biol 2014; 30: 677- 704.

[5] Boyette LB, Tuan RS. Adult stem cells and diseases of aging. J Clin Med 2014; 3: 88- 134.

[6] Burgstaller S, Bischof H, Lukowski R et al. Investigating the K(+) sensitivity of cellular metabolism by extracellular flux analysis. STAR Protoc 2021; 2: 100876.

[7] Carvalho GB, Drago I, Hoxha S et al. The 4E-BP growth pathway regulates the effect of ambient temperature on metabolism and lifespan. Proc Natl Acad Sci USA 2017; 114: 9737- 42.

[8] Chen Q, Vazquez EJ, Moghaddas S et al. Production of reactive oxygen species by mitochondria: central role of complex III. J Biol Chem 2003; 278: 36027- 31.

[9] Choo AY, Yoon S-O, Kim SG et al. Rapamycin differentially inhibits S6Ks and 4E-BP1 to mediate cell-type-specific repression of mRNA translation. Proc Natl Acad Sci USA 2008; 105: 17414- 9.

[10] Chu Q, Liu F, He Y et al. mTORC2/RICTOR exerts differential levels of metabolic control in human embryonic, mesenchymal and neural stem cells. Protein Cell 2022; 13: 676- 82.

[11] Daum B, Walter A, Horst A et al. Age-dependent dissociation of ATP synthase dimers and loss of inner-membrane cristae in mitochondria. Proc Natl Acad Sci USA 2013; 110: 15301- 6.

[12] Demontis F, Perrimon N. FOXO/4E-BP signaling in Drosophila muscles regulates organism-wide proteostasis during aging. Cell 2010; 143: 813- 25.

[13] Diao Z, Ji Q, Wu Z et al. SIRT3 consolidates heterochromatin and counteracts senescence. Nucleic Acids Res 2021; 49: 4203- 19.

[14] Ermolaeva M, Neri F, Ori A et al. Cellular and epigenetic drivers of stem cell ageing. Nat Rev Mol Cell Biol 2018; 19: 594- 610.

[15] Feher J, Kovacs I, Artico M et al. Mitochondrial alterations of retinal pigment epithelium in age-related macular degeneration. Neurobiol Aging 2006; 27: 983- 93.

[16] Fehrer C, Lepperdinger G. Mesenchymal stem cell aging. Exp Gerontol 2005; 40: 926- 30.

[17] Fu L, Hu Y, Song M et al. Up-regulation of FOXD1 by YAP alleviates senescence and osteoarthritis. PLoS Biol 2019; 17: e3000201.

[18] Gaignard P, Eyer D, Lebigot E et al. UQCRC2 mutation in a patient with mitochondrial complex III deficiency causing recurrent liver failure, lactic acidosis and hypoglycemia. J Hum Genet 2017; 62: 729- 31.

[19] Gandin V, Sikström K, Alain T et al. Polysome fractionation and analysis of mammalian translatomes on a genome-wide scale. J Vis Exp 2014; 87: 51455.

[20] Geng L, Liu Z, Zhang W et al. Chemical screen identifies a geroprotective role of quercetin in premature aging [J]. Protein Cell 2019; 10: 417- 35.

[21] Goodell MA, Rando TA. Stem cells and healthy aging. Science 2015; 350: 1199- 204.

[22] Hu H, Ji Q, Song M et al. ZKSCAN3 counteracts cellular senescence by stabilizing heterochromatin. Nucleic Acids Res 2020; 48: 6001- 18.

[23] Kang M-J, Vasudevan D, Kang K et al. 4E-BP is a target of the GCN2-ATF4 pathway during Drosophila development and aging. J Cell Biol 2017; 216: 115- 29.

[24] Kong M, Guo L, Xu W, He C, Jia X, Zhao Z, Gu Z. Aging-associated accumulation of mitochondrial DNA mutations in tumor origin. Life Med 2022. 10.1093/lifemedi/lnac014

[25] Le Bacquer O, Combe K, Patrac V et al. 4E-BP1 and 4E-BP2 double knockout mice are protected from aging-associated sarcopenia. J Cachexia Sarcopenia Muscle 2019; 10: 696- 709.

[26] Leng SX, Pawelec G. Single-cell immune atlas for human aging and frailty. Life Med 2022. 10.1093/lifemedi/lnac013

[27] Li W, Zou Z, Cai Y et al. Low-dose chloroquine treatment extends the lifespan of aged rats. Protein Cell 2022; 13: 454- 61.

[28] Liang C, Liu Z, Song M et al. Stabilization of heterochromatin by CLOCK promotes stem cell rejuvenation and cartilage regeneration. Cell Res 2021; 31: 187- 205.

[29] Llanos S, Garcia-Pedrero JM, Morgado-Palacin L et al. Stabilization of p21 by mTORC1/4E-BP1 predicts clinical outcome of head and neck cancers. Nat Commun 2016; 7: 10438.

[30] McCrimmon A, Domondon M, Sultanova RF et al. Comprehensive assessment of mitochondrial respiratory function in freshly isolated nephron segments. Am J Physiol Renal Physiol 2020; 318: F1237- 45.

[31] Miyake N, Yano S, Sakai C et al. Mitochondrial complex III deficiency caused by a homozygous UQCRC2 mutation presenting with neonatal-onset recurrent metabolic decompensation. Hum Mutat 2013; 34: 446- 52.

[32] Moskal N, Riccio V, Bashkurov M et al. ROCK inhibitors upregulate the neuroprotective Parkin-mediated mitophagy pathway. Nat Commun 2020; 11: 88.

[33] Musa J, Orth MF, Dallmayer M et al. Eukaryotic initiation factor 4E-binding protein 1 (4E-BP1): a master regulator of mRNA translation involved in tumorigenesis. Oncogene 2016; 35: 4675- 88.

[34] Neves J, Sousa-Victor P, Jasper H. Rejuvenating strategies for stem cell-based therapies in aging. Cell stem cell 2017; 20: 161- 75.

[35] Oh J, Lee YD, Wagers AJ. Stem cell aging: mechanisms, regulators and therapeutic opportunities. Nat Med 2014; 20: 870- 80.

[36] Panda AC, Martindale JL, Gorospe M. Polysome Fractionation to Analyze mRNA Distribution Profiles [J]. Bio Protoc, 2017, 73: e2126.

[37] Payne BAI, Chinnery PF. Mitochondrial dysfunction in aging: Much progress but many unresolved questions. Biochim Biophys Acta 2015; 1847: 1347- 53.

[38] Petroulakis E, Parsyan A, Dowling RJ et al. p53-dependent translational control of senescence and transformation via 4E-BPs. Cancer Cell 2009; 16: 439- 46.

[39] Protasoni M, Pérez-Pérez R, Lobo-Jarne T et al. Respiratory supercomplexes act as a platform for complex III-mediated maturation of human mitochondrial complexes I and IV. EMBO J 2020; 39: e102817.

[40] Qin X, Jiang B, Zhang Y. 4E-BP1, a multifactor regulated multifunctional protein. Cell Cycle 2016; 15: 781- 6.

[41] Ren R, Ocampo A, Liu G-H et al. Regulation of Stem Cell Aging by Metabolism and Epigenetics. Cell Metab 2017; 26: 460- 74.

[42] Schalm SS, Fingar DC, Sabatini DM et al. TOS motif-mediated raptor binding regulates 4E-BP1 multisite phosphorylation and function. Curr Biol 2003; 13: 797- 806.

[43] Schultz MB, Sinclair DA. When stem cells grow old: phenotypes and mechanisms of stem cell aging. Development 2016; 143: 3- 14.

[44] Shukla KK, Kwon W-S, Rahman MS et al. Nutlin-3a decreases male fertility via UQCRC2. PLoS ONE 2013; 8: e76959.

[45] Silva DF, Selfridge JE, Lu J et al. Bioenergetic flux, mitochondrial mass and mitochondrial morphology dynamics in AD and MCI cybrid cell lines. Hum Mol Genet 2013; 22: 3931- 46.

[46] Song G, Ma Y, Gao X et al. CRISPR/Cas9-mediated Genetic Correction Reverses Spinocerebellar Ataxia 3 Disease-associated Phenotypes in Differentiated Cerebellar Neurons. Life Med 2022. 10.1093/lifemedi/lnac020

[47] Spinelli JB, Haigis MC. The multifaceted contributions of mitochondria to cellular metabolism. Nat Cell Biol 2018; 20: 745- 54.

[48] Sun Y, Li Q, Kirkland JL. Targeting senescent cells for a healthier longevity: the roadmap for an era of global aging. Life Med 2022. 10.1093/lifemedi/lnac030.

[49] Szeto HH, Liu S. Cardiolipin-targeted peptides rejuvenate mitochondrial function, remodel mitochondria, and promote tissue regeneration during aging. Arch Biochem Biophys 2018; 660: 137- 48.

[50] Thoreen CC, Chantranupong L, Keys HR et al. A unifying model for mTORC1-mediated regulation of mRNA translation. Nature 2012; 485: 109- 13.

[51] Tsai S-Y, Rodriguez AA, Dastidar SG et al. Increased 4E-BP1 expression protects against diet-induced obesity and insulin resistance in male mice. Cell Rep 2016a; 16: 1903- 14.

[52] Tsai SY, Rodriguez AA, Dastidar SG et al. Increased 4E-BP1 expression protects against diet-induced obesity and insulin resistance in male mice. Cell Rep 2016b; 16: 1903- 14.

[53] Tucker EJ, Wanschers BFJ, Szklarczyk R et al. Mutations in the UQCC1-interacting protein, UQCC2, cause human complex III deficiency associated with perturbed cytochrome b protein expression. PLoS Genet 2013; 9: e1004034.

[54] Vakifahmetoglu-Norberg , H , Ouchida AT, Norberg , E. . The role of mitochondria in metabolism and cell death. Biochem Biophys Res Commun 2017; 482: 426- 31.

[55] Wang Y, Liu Y, Chen E et al. The role of mitochondrial dysfunction in mesenchymal stem cell senescence. Cell Tissue Res 2020; 382: 457- 62.

[56] Wang L, Liu J, Liu H et al. The secret of youth - how is systemic rejuvenation achieved at the single cell level? Life Med 2022. 10.1093/lifemedi/lnac018

[57] Weichhart T. mTOR as regulator of lifespan, aging, and cellular senescence: a mini-review. Gerontology 2018; 64: 127- 34.

[58] Wu Z, Zhang W, Song M et al. Differential stem cell aging kinetics in Hutchinson-Gilford progeria syndrome and Werner syndrome[J]. Protein Cell 2018; 9: 333- 50.

[59] Xu X, Duan S, Yi F et al. Mitochondrial regulation in pluripotent stem cells. Cell Metab 2013; 18: 325- 32.

[60] Yang Y, Li X, Zhang T et al. RIP kinases and necroptosis in aging and aging-related diseases. Life Med 2022. 10.1093/lifemedi/lnac003.

[61] Yao CH, Wang R, Wang Y et al. Mitochondrial fusion supports increased oxidative phosphorylation during cell proliferation. Elife 2019; 8: e41351.

[62] Zhang S, Wu Z, Shi Y et al. FTO stabilizes MIS12 and counteracts senescence. Protein Cell 2022; 13: 954- 60.

[63] Zhao D, Chen S. Failures at every level: breakdown of the epigenetic machinery of aging. Life Med 2022. 10.1093/lifemedi/lnac016

[64] Zhao H, Ji Q, Wu Z et al. Destabilizing heterochromatin by APOE mediates senescence. Nat Aging 2022; 2: 303- 16.

[65] Zhou Y, Zhou B, Pache L, Chang M, Khodabakhshi AH, Tanaseichuk O, Benner C, Chanda SK. Metascape provides a biologist-oriented resource for the analysis of systems-level datasets. Nat Commun. 2019 Apr 3; 101: 1523. 10.1038/s41467-019-09234-6.

[66] Zid BM, Rogers AN, Katewa SD et al. 4E-BP extends lifespan upon dietary restriction by enhancing mitochondrial activity in Drosophila. Cell 2009; 139: 149- 60.

[67] Ziegler DV, Wiley CD, Velarde MC. Mitochondrial effectors of cellular senescence: beyond the free radical theory of aging. Aging Cell 2015; 14: 1- 7.

[68] Zong W-X, Rabinowitz JD, White E. Mitochondria and Cancer. Mol Cell 2016; 61: 667- 76.

RIGHTS & PERMISSIONS

The Author(s) 2022. Published by Oxford University Press on behalf of Higher Education Press.