Abstract
Transcription factor EB (TFEB) is a master regulator of lysosomal biogenesis and cellular clearance pathways. TFEB activity is tightly controlled by multiple post-translational mechanisms, but the exact molecular mechanism controlling its stability has remained elusive. Here, we identify the IκB kinase (IKK) complex as a key regulator of TFEB protein stability through a phosphorylation–ubiquitination cascade. A high-content kinase inhibitor screen reveals that IKK inhibition increases TFEB protein levels, and genetic ablation of IKK components increases TFEB stability, upregulates lysosomal genes, and enhances lysosomal biogenesis and degradative capacity. Mechanistically, we show that IKK phosphorylates TFEB on a cluster of serine residues (423SPFPSLS429), generating a phosphodegron recognized by the E3 ligase β-TrCP2, which in turn targets TFEB for proteasomal degradation via ubiquitination of adjacent lysine residues (K430 and K431). Mutation of either the phosphosites or the ubiquitination sites stabilizes TFEB without impairing its ability to translocate to the nucleus, activate target gene expression, or promote tau clearance in a cell model of tauopathy. These findings establish IKK–β-TrCP2 as a core regulatory axis controlling TFEB protein turnover and levels and reveal a mechanistically distinct layer of TFEB regulation that may be leveraged to enhance lysosomal function in disease contexts.
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Data availability
Raw mass spectrometry data have been deposited in the MassIVE publicly available repository under accession code: Accession: MSV000093106 (TFEB pS427 detection) ftp://massive-ftp.ucsd.edu/v06/MSV000093106/ Accession: MSV000095363 (TFEB phospho-peptide analysis following IL-1β treatment) ftp://massive-ftp.ucsd.edu/v08/MSV000095363/ Accession: MSV000096549 (TFEB ubiquitination analysis) ftp://massive-ftp.ucsd.edu/v07/MSV000096549/ All other data generated or analyzed in this study are included in the article and its Supplementary Information and the Source data file. Source data are provided in this paper.
References
Sardiello, M. et al. A gene network regulating lysosomal biogenesis and function. Science 325, 473–477 (2009).
Settembre, C. et al. TFEB links autophagy to lysosomal biogenesis. Science 332, 1429–1433 (2011).
Palmieri, M. et al. Characterization of the CLEAR network reveals an integrated control of cellular clearance pathways. Hum. Mol. Genet. 20, 3852–3866 (2011).
Song, W. et al. TFEB regulates lysosomal proteostasis. Hum. Mol. Genet. 22, 1994–2009 (2013).
Medina, D. L. et al. Transcriptional activation of lysosomal exocytosis promotes cellular clearance. Dev. Cell 21, 421–430 (2011).
Martina, J. A., Diab, H. I., Brady, O. A. & Puertollano, R. TFEB and TFE3 are novel components of the integrated stress response. EMBO J. 35, 479–495 (2016).
Ma, X. et al. Regulation of the transcription factor EB-PGC1alpha axis by beclin-1 controls mitochondrial quality and cardiomyocyte death under stress. Mol. Cell Biol. 35, 956–976 (2015).
Mansueto, G. et al. Transcription factor EB controls metabolic flexibility during exercise. Cell Metab. 25, 182–196 (2017).
Pena-Llopis, S. et al. Regulation of TFEB and V-ATPases by mTORC1. EMBO J. 30, 3242–3258 (2011).
Vega-Rubin-de-Celis, S., Pena-Llopis, S., Konda, M. & Brugarolas, J. Multistep regulation of TFEB by MTORC1. Autophagy 13, 464–472 (2017).
Settembre, C. et al. A lysosome-to-nucleus signalling mechanism senses and regulates the lysosome via mTOR and TFEB. EMBO J. 31, 1095–1108 (2012).
Martina, J. A., Chen, Y., Gucek, M. & Puertollano, R. MTORC1 functions as a transcriptional regulator of autophagy by preventing nuclear transport of TFEB. Autophagy 8, 903–914 (2012).
Roczniak-Ferguson, A. et al. The transcription factor TFEB links mTORC1 signaling to transcriptional control of lysosome homeostasis. Sci. Signal 5, ra42 (2012).
Palmieri, M. et al. mTORC1-independent TFEB activation via Akt inhibition promotes cellular clearance in neurodegenerative storage diseases. Nat. Commun. 8, 14338 (2017).
Palmieri, M., Pal, R. & Sardiello, M. AKT modulates the autophagy-lysosome pathway via TFEB. Cell Cycle 16, 1237–1238 (2017).
Paquette, M. et al. AMPK-dependent phosphorylation is required for transcriptional activation of TFEB and TFE3. Autophagy 17, 3957–3975 (2021).
Ma, T. et al. Low-dose metformin targets the lysosomal AMPK pathway through PEN2. Nature 603, 159–165 (2022).
Chen, M., Sun, T., Zhong, Y., Zhou, X. & Zhang, J. A highly sensitive fluorescent Akt biosensor reveals lysosome-selective regulation of lipid second messengers and kinase activity. ACS Cent. Sci. 7, 2009–2020 (2021).
Hyun, S. W. et al. The sialidase NEU1 directly interacts with the juxtamembranous segment of the cytoplasmic domain of mucin-1 to inhibit downstream PI3K-Akt signaling. J. Biol. Chem. 297, 101337 (2021).
Li, Y. et al. Protein kinase C controls lysosome biogenesis independently of mTORC1. Nat. Cell Biol. 18, 1065–1077 (2016).
Martina, J. A., Jeong, E. & Puertollano, R. p38 MAPK-dependent phosphorylation of TFEB promotes monocyte-to-macrophage differentiation. EMBO Rep. 24, e55472 (2023).
Hsu, C. L. et al. MAP4K3 mediates amino acid-dependent regulation of autophagy via phosphorylation of TFEB. Nat. Commun. 9, 942 (2018).
Yin, Q. et al. CDK4/6 regulate lysosome biogenesis through TFEB/TFE3. J. Cell Biol. 219, https://doi.org/10.1083/jcb.201911036 (2020).
Medina, D. L. et al. Lysosomal calcium signalling regulates autophagy through calcineurin and TFEB. Nat. Cell Biol. 17, 288–299 (2015).
Song, W., Wang, F., Lotfi, P., Sardiello, M. & Segatori, L. 2-Hydroxypropyl-beta-cyclodextrin promotes transcription factor EB-mediated activation of autophagy: implications for therapy. J. Biol. Chem. 289, 10211–10222 (2014).
Polito, V. A. et al. Selective clearance of aberrant tau proteins and rescue of neurotoxicity by transcription factor EB. EMBO Mol. Med. 6, 1142–1160 (2014).
Lotfi, P. et al. Trehalose reduces retinal degeneration, neuroinflammation and storage burden caused by a lysosomal hydrolase deficiency. Autophagy 14, 1419–1434 (2018).
Martini-Stoica, H. et al. TFEB enhances astroglial uptake of extracellular tau species and reduces tau spreading. J. Exp. Med. 215, 2355–2377 (2018).
Sardiello, M. Transcription factor EB: from master coordinator of lysosomal pathways to candidate therapeutic target in degenerative storage diseases. Ann. N. Y Acad. Sci. 1371, 3–14 (2016).
Sharma, J., di Ronza, A., Lotfi, P. & Sardiello, M. Lysosomes and brain health. Annu. Rev. Neurosci. 41, 255–276 (2018).
Xu, Y. et al. TFEB regulates lysosomal exocytosis of tau and its loss of function exacerbates tau pathology and spreading. Mol. Psychiatry 26, 5925–5939 (2021).
Lee, D. F. et al. IKK beta suppression of TSC1 links inflammation and tumor angiogenesis via the mTOR pathway. Cell 130, 440–455 (2007).
Dan, H. C. et al. Akt-dependent activation of mTORC1 complex involves phosphorylation of mTOR (mammalian target of rapamycin) by IkappaB kinase alpha (IKKalpha). J. Biol. Chem. 289, 25227–25240 (2014).
Dan, H. C. & Baldwin, A. S. Differential involvement of IkappaB kinases alpha and beta in cytokine- and insulin-induced mammalian target of rapamycin activation determined by Akt. J. Immunol. 180, 7582–7589 (2008).
Furuya, K. et al. Stabilization of p73 by nuclear IkappaB kinase-alpha mediates cisplatin-induced apoptosis. J. Biol. Chem. 282, 18365–18378 (2007).
Yamaguchi, T., Miki, Y. & Yoshida, K. Protein kinase C delta activates IkappaB-kinase alpha to induce the p53 tumor suppressor in response to oxidative stress. Cell Signal 19, 2088–2097 (2007).
Kwak, Y. T. et al. IkappaB kinase alpha regulates subcellular distribution and turnover of cyclin D1 by phosphorylation. J. Biol. Chem. 280, 33945–33952 (2005).
Sil, A. K., Maeda, S., Sano, Y., Roop, D. R. & Karin, M. IkappaB kinase-alpha acts in the epidermis to control skeletal and craniofacial morphogenesis. Nature 428, 660–664 (2004).
Anest, V., Cogswell, P. C. & Baldwin, A. S. Jr. IkappaB kinase alpha and p65/RelA contribute to optimal epidermal growth factor-induced c-fos gene expression independent of IkappaBalpha degradation. J. Biol. Chem. 279, 31183–31189 (2004).
Dong, W. et al. IKKalpha contributes to UVB-induced VEGF expression by regulating AP-1 transactivation. Nucleic Acids Res. 40, 2940–2955 (2012).
Fernandez-Majada, V. et al. Nuclear IKK activity leads to dysregulated notch-dependent gene expression in colorectal cancer. Proc. Natl. Acad. Sci. USA 104, 276–281 (2007).
Kanemori, Y., Uto, K. & Sagata, N. Beta-TrCP recognizes a previously undescribed nonphosphorylated destruction motif in Cdc25A and Cdc25B phosphatases. Proc. Natl. Acad. Sci. USA 102, 6279–6284 (2005).
Zandi, E., Rothwarf, D. M., Delhase, M., Hayakawa, M. & Karin, M. The IkappaB kinase complex (IKK) contains two kinase subunits, IKKalpha and IKKbeta, necessary for IkappaB phosphorylation and NF-kappaB activation. Cell 91, 243–252 (1997).
Solt, L. A. & May, M. J. The IkappaB kinase complex: master regulator of NF-kappaB signaling. Immunol. Res. 42, 3–18 (2008).
Israel, A. The IKK complex, a central regulator of NF-kappaB activation. Cold Spring Harb. Perspect. Biol. 2, a000158 (2010).
Gao, X. et al. Tsc tumour suppressor proteins antagonize amino-acid-TOR signalling. Nat. Cell Biol. 4, 699–704 (2002).
Fuchs, S. Y., Spiegelman, V. S. & Kumar, K. G. The many faces of beta-TrCP E3 ubiquitin ligases: reflections in the magic mirror of cancer. Oncogene 23, 2028–2036 (2004).
Magliozzi, R. et al. Control of epithelial cell migration and invasion by the IKKbeta- and CK1alpha-mediated degradation of RAPGEF2. Dev. Cell 27, 574–585 (2013).
Xia, Y. et al. Phosphorylation of p53 by IkappaB kinase 2 promotes its degradation by beta-TrCP. Proc. Natl. Acad. Sci. USA 106, 2629–2634 (2009).
Iwasaki, H. et al. The IkappaB kinase complex regulates the stability of cytokine-encoding mRNA induced by TLR-IL-1R by controlling degradation of regnase-1. Nat. Immunol. 12, 1167–1175 (2011).
Cui, D. et al. The cross talk of two family members of beta-TrCP in the regulation of cell autophagy and growth. Cell Death Differ. 27, 1119–1133 (2020).
Shi, G. et al. Rapalogs downmodulate intrinsic immunity and promote cell entry of SARS-CoV-2. J. Clin. Invest. 132, https://doi.org/10.1172/jci160766 (2022).
Zhang, X. et al. Rapamycin directly activates lysosomal mucolipin TRP channels independent of mTOR. PLoS Biol. 17, e3000252 (2019).
Zhen, Y., McGaha, T. L., Finkelman, F. D. & Shao, W. H. The Akt-mTORC1 pathway mediates Axl receptor tyrosine kinase-induced mesangial cell proliferation. J. Leukoc. Biol. 111, 563–571 (2022).
Park, D. W. et al. The JAK2-Akt-glycogen synthase kinase-3beta signaling pathway is involved in toll-like receptor 2-induced monocyte chemoattractant protein-1 regulation. Mol. Med Rep. 5, 1063–1067 (2012).
Hu, Y. et al. Abnormal morphogenesis but intact IKK activation in mice lacking the IKKalpha subunit of IkappaB kinase. Science 284, 316–320 (1999).
Li, Z. W. et al. The IKKbeta subunit of IkappaB kinase (IKK) is essential for nuclear factor kappaB activation and prevention of apoptosis. J. Exp. Med. 189, 1839–1845 (1999).
Makris, C. et al. Female mice heterozygous for IKK gamma/NEMO deficiencies develop a dermatopathy similar to the human X-linked disorder incontinentia pigmenti. Mol. Cell 5, 969–979 (2000).
Pastore, N. et al. TFEB and TFE3 cooperate in the regulation of the innate immune response in activated macrophages. Autophagy 12, 1240–1258 (2016).
Liu, F., Xia, Y., Parker, A. S. & Verma, I. M. IKK biology. Immunol. Rev. 246, 239–253 (2012).
Antonia, R. J., Hagan, R. S. & Baldwin, A. S. Expanding the view of IKK: new substrates and new biology. Trends Cell Biol. 31, 166–178 (2021).
Kinoshita, E., Kinoshita-Kikuta, E., Takiyama, K. & Koike, T. Phosphate-binding tag, a new tool to visualize phosphorylated proteins. Mol. Cell Proteom. 5, 749–757 (2006).
Hutti, J. E. et al. IkappaB kinase beta phosphorylates the K63 deubiquitinase A20 to cause feedback inhibition of the NF-kappaB pathway. Mol. Cell Biol. 27, 7451–7461 (2007).
Marinis, J. M. et al. IkappaB kinase alpha phosphorylation of TRAF4 downregulates innate immune signaling. Mol. Cell Biol. 32, 2479–2489 (2012).
Kong, A. T., Leprevost, F. V., Avtonomov, D. M., Mellacheruvu, D. & Nesvizhskii, A. I. MSFragger: ultrafast and comprehensive peptide identification in mass spectrometry-based proteomics. Nat. Methods 14, 513–520 (2017).
Sha, Y., Rao, L., Settembre, C., Ballabio, A. & Eissa, N. T. STUB1 regulates TFEB-induced autophagy-lysosome pathway. EMBO J. 36, 2544–2552 (2017).
Lear, T. B. et al. Inhibition of virally induced TFEB proteasomal degradation as a host-centric therapeutic approach for coronaviral infection. Sci Adv. 11, eadv4033 (2025).
Low, T. Y. et al. A systems-wide screen identifies substrates of the SCFbetaTrCP ubiquitin ligase. Sci. Signal 7, rs8 (2014).
Mavrommati, I. et al. beta-TrCP- and casein kinase II-mediated degradation of cyclin F controls timely mitotic progression. Cell Rep. 24, 3404–3412 (2018).
Tatarskiy, V. V. et al. Stability of the PHF10 subunit of PBAF signature module is regulated by phosphorylation: role of beta-TrCP. Sci. Rep. 7, 5645 (2017).
Chowdhry, S. et al. Nrf2 is controlled by two distinct beta-TrCP recognition motifs in its Neh6 domain, one of which can be modulated by GSK-3 activity. Oncogene 32, 3765–3781 (2013).
Melvin, A. T. et al. A comparative analysis of the ubiquitination kinetics of multiple degrons to identify an ideal targeting sequence for a proteasome reporter. PLoS ONE 8, e78082 (2013).
Winer, I. S., Bommer, G. T., Gonik, N. & Fearon, E. R. Lysine residues Lys-19 and Lys-49 of beta-catenin regulate its levels and function in T cell factor transcriptional activation and neoplastic transformation. J. Biol. Chem. 281, 26181–26187 (2006).
Wu, G. et al. Structure of a beta-TrCP1-Skp1-beta-catenin complex: destruction motif binding and lysine specificity of the SCF(beta-TrCP1) ubiquitin ligase. Mol. Cell 11, 1445–1456 (2003).
Lamming, D. W. & Bar-Peled, L. Lysosome: The metabolic signaling hub. Traffic 20, 27–38 (2019).
Luzio, J. P., Pryor, P. R. & Bright, N. A. Lysosomes: fusion and function. Nat. Rev. Mol. Cell Biol. 8, 622–632 (2007).
Saftig, P. & Klumperman, J. Lysosome biogenesis and lysosomal membrane proteins: trafficking meets function. Nat. Rev. Mol. Cell Biol. 10, 623–635 (2009).
Bellettato, C. M. & Scarpa, M. Pathophysiology of neuropathic lysosomal storage disorders. J. Inherit. Metab. Dis. 33, 347–362 (2010).
Meikle, P. J., Hopwood, J. J., Clague, A. E. & Carey, W. F. Prevalence of lysosomal storage disorders. JAMA 281, 249–254 (1999).
Hoffmann, B. & Mayatepek, E. Neurological manifestations in lysosomal storage disorders - from pathology to first therapeutic possibilities. Neuropediatrics 36, 285–289 (2005).
Cortes, C. J. & La Spada, A. R. TFEB dysregulation as a driver of autophagy dysfunction in neurodegenerative disease: Molecular mechanisms, cellular processes, and emerging therapeutic opportunities. Neurobiol. Dis. 122, 83–93 (2019).
Napolitano, G. & Ballabio, A. TFEB at a glance. J. Cell Sci. 129, 2475–2481 (2016).
Raben, N. & Puertollano, R. TFEB and TFE3: Linking Lysosomes to Cellular Adaptation to Stress. Annu. Rev. Cell Dev. Biol. 32, 255–278 (2016).
Liu, G. Y. & Sabatini, D. M. mTOR at the nexus of nutrition, growth, ageing and disease. Nat. Rev. Mol. Cell Biol. 21, 183–203 (2020).
Odle, R. I. et al. An mTORC1-to-CDK1 Switch Maintains Autophagy Suppression during Mitosis. Mol. Cell 77, 228–240.e227 (2020).
Nixon, R. A. The aging lysosome: An essential catalyst for late-onset neurodegenerative diseases. Biochim. Biophys. Acta Proteins Proteom. 1868, 140443 (2020).
Boland, B. & Platt, F. M. Bridging the age spectrum of neurodegenerative storage diseases. Best. Pr. Res. Clin. Endocrinol. Metab. 29, 127–143 (2015).
Xiao, Q. et al. Neuronal-targeted TFEB accelerates lysosomal degradation of APP, reducing Abeta generation and amyloid plaque pathogenesis. J. Neurosci. 35, 12137–12151 (2015).
Decressac, M. et al. TFEB-mediated autophagy rescues midbrain dopamine neurons from alpha-synuclein toxicity. Proc. Natl. Acad. Sci. USA 110, E1817–E1826 (2013).
Torra, A. et al. Overexpression of TFEB drives a pleiotropic neurotrophic effect and prevents Parkinson’s disease-related neurodegeneration. Mol. Ther. 26, 1552–1567 (2018).
Arotcarena, M. L. et al. Transcription factor EB overexpression prevents neurodegeneration in experimental synucleinopathies. JCI Insight 4, e129719 (2019).
Puertollano, R., Ferguson, S. M., Brugarolas, J. & Ballabio, A. The complex relationship between TFEB transcription factor phosphorylation and subcellular localization. EMBO J. 37, https://doi.org/10.15252/embj.201798804 (2018).
Takla, M., Keshri, S. & Rubinsztein, D. C. The post-translational regulation of transcription factor EB (TFEB) in health and disease. EMBO Rep. 24, e57574 (2023).
Dodeller, F., Gottar, M., Huesken, D., Iourgenko, V. & Cenni, B. The lysosomal transmembrane protein 9B regulates the activity of inflammatory signaling pathways. J. Biol. Chem. 283, 21487–21494 (2008).
Glowacka, W. K., Alberts, P., Ouchida, R., Wang, J. Y. & Rotin, D. LAPTM5 protein is a positive regulator of proinflammatory signaling pathways in macrophages. J. Biol. Chem. 287, 27691–27702 (2012).
Zein, L. et al. Linear ubiquitination at damaged lysosomes induces local NFKB activation and controls cell survival. Autophagy 21, 1075–1095 (2025).
Hayashi, K., Taura, M. & Iwasaki, A. The interaction between IKKalpha and LC3 promotes type I interferon production through the TLR9-containing LAPosome. Sci. Signal 11, https://doi.org/10.1126/scisignal.aan4144 (2018).
Liu, S. & Chen, Z. J. Expanding role of ubiquitination in NF-kappaB signaling. Cell Res. 21, 6–21 (2011).
Schnoder, L. et al. Deficiency of IKKbeta in neurons ameliorates Alzheimer’s disease pathology in APP- and tau-transgenic mice. FASEB J. 37, e22778 (2023).
Gu, Z. et al. TFEB in Alzheimer’s disease: From molecular mechanisms to therapeutic implications. Neurobiol. Dis. 173, 105855 (2022).
Cadwell, K. Crosstalk between autophagy and inflammatory signalling pathways: balancing defence and homeostasis. Nat. Rev. Immunol. 16, 661–675 (2016).
Harris, J. et al. Autophagy controls IL-1beta secretion by targeting pro-IL-1beta for degradation. J. Biol. Chem. 286, 9587–9597 (2011).
Nakahira, K. et al. Autophagy proteins regulate innate immune responses by inhibiting the release of mitochondrial DNA mediated by the NALP3 inflammasome. Nat. Immunol. 12, 222–230 (2011).
Liu, K. et al. SKP2 attenuates NF-kappaB signaling by mediating IKKbeta degradation through autophagy. J. Mol. Cell Biol. 10, 205–215 (2018).
Francois, A. et al. Impairment of autophagy in the central nervous system during lipopolysaccharide-induced inflammatory stress in mice. Mol. Brain 7, 56 (2014).
Zhou, M. et al. Boosting mTOR-dependent autophagy via upstream TLR4-MyD88-MAPK signalling and downstream NF-kappaB pathway quenches intestinal inflammation and oxidative stress injury. EBioMedicine 35, 345–360 (2018).
Xia, L. et al. Impaired autophagy increases susceptibility to endotoxin-induced chronic pancreatitis. Cell Death Dis. 11, 889 (2020).
Owen, K. A., Meyer, C. B., Bouton, A. H. & Casanova, J. E. Activation of focal adhesion kinase by Salmonella suppresses autophagy via an Akt/mTOR signaling pathway and promotes bacterial survival in macrophages. PLoS Pathog. 10, e1004159 (2014).
Shi, C. S. et al. Activation of autophagy by inflammatory signals limits IL-1beta production by targeting ubiquitinated inflammasomes for destruction. Nat. Immunol. 13, 255–263 (2012).
Prieto, P. et al. Activation of autophagy in macrophages by pro-resolving lipid mediators. Autophagy 11, 1729–1744 (2015).
Irazoqui, J. E. Key Roles of MiT transcription factors in innate immunity and inflammation. Trends Immunol. 41, 157–171 (2020).
El-Houjeiri, L. et al. The Transcription factors TFEB and TFE3 link the FLCN-AMPK signaling axis to innate immune response and pathogen resistance. Cell Rep. 26, 3613–3628 e3616 (2019).
Visvikis, O. et al. Innate host defense requires TFEB-mediated transcription of cytoprotective and antimicrobial genes. Immunity 40, 896–909 (2014).
Hayama, Y. et al. Lysosomal protein lamtor1 controls innate immune responses via nuclear translocation of transcription factor EB. J. Immunol. 200, 3790–3800 (2018).
Song, W. et al. Endothelial TFEB (transcription factor EB) restrains IKK (IkappaB Kinase)-p65 pathway to attenuate vascular inflammation in diabetic db/db mice. Arterioscler Thromb. Vasc. Biol. 39, 719–730 (2019).
Tsunemi, T. et al. PGC-1alpha rescues Huntington’s disease proteotoxicity by preventing oxidative stress and promoting TFEB function. Sci. Transl. Med. 4, 142ra197 (2012).
Shimizu, K. et al. The SCFbeta-TRCP E3 ubiquitin ligase complex targets Lipin1 for ubiquitination and degradation to promote hepatic lipogenesis. Sci. Signal 10, https://doi.org/10.1126/scisignal.aah4117 (2017).
Seo, E. et al. Multiple isoforms of beta-TrCP display differential activities in the regulation of Wnt signaling. Cell Signal 21, 43–51 (2009).
Davis, M. et al. Pseudosubstrate regulation of the SCF(beta-TrCP) ubiquitin ligase by hnRNP-U. Genes Dev. 16, 439–451 (2002).
Chang, K. T., Guo, J., di Ronza, A. & Sardiello, M. Aminode: identification of evolutionary constraints in the human proteome. Sci. Rep. 8, 1357 (2018).
Durand, J. K. & Baldwin, A. S. Targeting IKK and NF-kappaB for therapy. Adv. Protein Chem. Struct. Biol. 107, 77–115 (2017).
DiMaggio, P. A. Jr., Young, N. L., Baliban, R. C., Garcia, B. A. & Floudas, C. A. A mixed integer linear optimization framework for the identification and quantification of targeted post-translational modifications of highly modified proteins using multiplexed electron transfer dissociation tandem mass spectrometry. Mol. Cell Proteom. 8, 2527–2543 (2009).
Holt, M. V., Wang, T. & Young, N. L. High-throughput quantitative top-down proteomics: histone H4. J. Am. Soc. Mass Spectrom. 30, 2548–2560 (2019).
Wisniewski, J. R., Zougman, A., Nagaraj, N. & Mann, M. Universal sample preparation method for proteome analysis. Nat. Methods 6, 359–362 (2009).
Chen, Z. W., Fuchs, K., Sieghart, W., Townsend, R. R. & Evers, A. S. Deep amino acid sequencing of native brain GABAA receptors using high-resolution mass spectrometry. Mol. Cell Proteomics 11, M111 011445 (2012).
Meier, F. et al. Online parallel accumulation-serial fragmentation (PASEF) with a novel trapped ion mobility mass spectrometer. Mol. Cell Proteom. 17, 2534–2545 (2018).
Perkins, D. N., Pappin, D. J., Creasy, D. M. & Cottrell, J. S. Probability-based protein identification by searching sequence databases using mass spectrometry data. Electrophoresis 20, 3551–3567 (1999).
Acknowledgements
We thank Mike Prinsen for assistance with running the high-throughput screen. This work was supported by NIH grants P01 AG066606 and RM1 NS132962, and grants from the Beyond Batten Disease Foundation (to M.S.). We thank Petra Erdmann-Gilmore, Dr. Yiling Mi and Rose Connors for their expert technical assistance in running the proteomic experiments at the Washington University Proteomics Shared Resource (WU-PSR), directed by R. Reid Townsend, MD, PhD, with Drs. Qiang Zhang, PhD, and Robert Sprung, PhD, as Co-Directors. The WU-PSR is supported in part by the WU Institute of Clinical and Translational Sciences (NCATS UL1 TR000448), the Mass Spectrometry Research Resource (NIGMS P41 GM103422; R24GM136766) and the Siteman Comprehensive Cancer Center Support Grant (NCI P30 CA091842).
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Y.X. and M.S. conceived the project and designed the experiments. M.N.Y. and N.L.Y. designed mass-spectrometry experiments. Y.X. and M.X.G.I designed high-throughput screening experiments. Y.X., J.S., W.X., M.N.Y., K.F.P., M.X.G.I., B.G. and Q.W. performed experiments and analyzed the data under the supervision of H.Z., N.L.Y. and M.S. A.J. performed bioinformatic analyses. Y.X. and M.S. wrote the manuscript with input from all authors.
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Xiong, Y., Sharma, J., Young, M.N. et al. TFEB degradation is regulated by an IKK/β-TrCP2 phosphorylation-ubiquitination cascade. Nat Commun (2026). https://doi.org/10.1038/s41467-026-71001-1
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DOI: https://doi.org/10.1038/s41467-026-71001-1
