A chemical genetics approach to examine the functions of AAA proteins.
AAA Proteins
/ antagonists & inhibitors
ATPases Associated with Diverse Cellular Activities
/ genetics
Basic Helix-Loop-Helix Transcription Factors
/ genetics
Endosomal Sorting Complexes Required for Transport
/ genetics
Humans
Katanin
/ genetics
Microtubule-Associated Proteins
/ genetics
Microtubules
/ genetics
Protein Conformation
/ drug effects
Protein Domains
/ genetics
Pyridines
/ chemistry
Triazoles
/ chemistry
Journal
Nature structural & molecular biology
ISSN: 1545-9985
Titre abrégé: Nat Struct Mol Biol
Pays: United States
ID NLM: 101186374
Informations de publication
Date de publication:
04 2021
04 2021
Historique:
received:
18
11
2020
accepted:
18
02
2021
pubmed:
31
3
2021
medline:
29
6
2021
entrez:
30
3
2021
Statut:
ppublish
Résumé
The structural conservation across the AAA (ATPases associated with diverse cellular activities) protein family makes designing selective chemical inhibitors challenging. Here, we identify a triazolopyridine-based fragment that binds the AAA domain of human katanin, a microtubule-severing protein. We have developed a model for compound binding and designed ASPIR-1 (allele-specific, proximity-induced reactivity-based inhibitor-1), a cell-permeable compound that selectively inhibits katanin with an engineered cysteine mutation. Only in cells expressing mutant katanin does ASPIR-1 treatment increase the accumulation of CAMSAP2 at microtubule minus ends, confirming specific on-target cellular activity. Importantly, ASPIR-1 also selectively inhibits engineered cysteine mutants of human VPS4B and FIGL1-AAA proteins, involved in organelle dynamics and genome stability, respectively. Structural studies confirm our model for compound binding at the AAA ATPase site and the proximity-induced reactivity-based inhibition. Together, our findings suggest a chemical genetics approach to decipher AAA protein functions across essential cellular processes and to test hypotheses for developing therapeutics.
Identifiants
pubmed: 33782614
doi: 10.1038/s41594-021-00575-9
pii: 10.1038/s41594-021-00575-9
pmc: PMC8592256
mid: NIHMS1721747
doi:
Substances chimiques
Basic Helix-Loop-Helix Transcription Factors
0
CAMSAP2 protein, human
0
Endosomal Sorting Complexes Required for Transport
0
FIGLA protein, human
0
Microtubule-Associated Proteins
0
Pyridines
0
Triazoles
0
AAA Proteins
EC 3.6.4.-
ATPases Associated with Diverse Cellular Activities
EC 3.6.4.-
VPS4B protein, human
EC 3.6.4.6
Katanin
EC 5.6.1.1
pyridine
NH9L3PP67S
Types de publication
Journal Article
Research Support, N.I.H., Extramural
Research Support, Non-U.S. Gov't
Research Support, U.S. Gov't, Non-P.H.S.
Langues
eng
Sous-ensembles de citation
IM
Pagination
388-397Subventions
Organisme : NIGMS NIH HHS
ID : R35 GM130234
Pays : United States
Organisme : NCI NIH HHS
ID : T32 CA009673
Pays : United States
Organisme : NIGMS NIH HHS
ID : T32 GM115327
Pays : United States
Références
Puchades, C., Sandate, C. R. & Lander, G. C. The molecular principles governing the activity and functional diversity of AAA proteins. Nat. Rev. Mol. Cell Biol. 21, 43–58 (2020).
pubmed: 31754261
doi: 10.1038/s41580-019-0183-6
Seraphim, T. V. & Houry, W. A. AAA proteins. Curr. Biol. 30, R251–R257 (2020).
pubmed: 32208144
doi: 10.1016/j.cub.2020.01.044
Gates, S. N. & Martin, A. Stairway to translocation: AAA+ motor structures reveal the mechanisms of ATP‐dependent substrate translocation. Protein Sci. 29, 407–419 (2020).
pubmed: 31599052
doi: 10.1002/pro.3743
Wendler, P., Ciniawsky, S., Kock, M. & Kube, S. Structure and function of the AAA nucleotide binding pocket. Biochim. Biophys. Acta Mol. Cell Res. 1823, 2–14 (2012).
doi: 10.1016/j.bbamcr.2011.06.014
McCullough, J., Frost, A. & Sundquist, W. I. Structures, functions, and dynamics of ESCRT-III/Vps4 membrane remodeling and fission complexes. Annu. Rev. Cell Dev. Biol. 34, 85–109 (2018).
pubmed: 30095293
pmcid: 6241870
doi: 10.1146/annurev-cellbio-100616-060600
Roll-Mecak, A. & McNally, F. J. Microtubule-severing enzymes. Curr. Opin. Cell Biol. 22, 96–103 (2010).
pubmed: 19963362
doi: 10.1016/j.ceb.2009.11.001
Bleichert, F., Botchan, M. R. & Berger, J. M. Mechanisms for initiating cellular DNA replication. Science 355, eaah6317 (2017).
pubmed: 28209641
doi: 10.1126/science.aah6317
Assimon, V. A. et al. CB-6644 is a selective inhibitor of the RUVBL1/2 complex with anticancer activity. ACS Chem. Biol. 14, 236–244 (2019).
pubmed: 30640450
doi: 10.1021/acschembio.8b00904
Anderson, D. J. et al. Targeting the AAA ATPase p97 as an approach to treat cancer through disruption of protein homeostasis. Cancer Cell 28, 653–665 (2015).
pubmed: 26555175
pmcid: 4941640
doi: 10.1016/j.ccell.2015.10.002
Bishop, A. C., Buzko, O. & Shokat, K. M. Magic bullets for protein kinases. Trends Cell Biol. 11, 167–172 (2001).
pubmed: 11306297
doi: 10.1016/S0962-8924(01)01928-6
Islam, K. The bump-and-hole tactic: expanding the scope of chemical genetics. Cell Chem. Biol. 25, 1171–1184 (2018).
pubmed: 30078633
pmcid: 6195450
doi: 10.1016/j.chembiol.2018.07.001
Zhou, H.-J. et al. Discovery of a first-in-class, potent, selective and orally bioavailable inhibitor of the p97 AAA ATPase (CB-5083). J. Med. Chem. 58, 9480–9497 (2015).
pubmed: 26565666
doi: 10.1021/acs.jmedchem.5b01346
Cupido, T., Pisa, R., Kelley, M. E. & Kapoor, T. M. Designing a chemical inhibitor for the AAA protein spastin using active site mutations. Nat. Chem. Biol. 15, 444–452 (2019).
pubmed: 30778202
pmcid: 6558985
doi: 10.1038/s41589-019-0225-6
Pisa, R., Cupido, T., Steinman, J. B., Jones, N. H. & Kapoor, T. M. Analyzing resistance to design selective chemical inhibitors for AAA proteins. Cell Chem. Biol. 26, 1263–1273 (2019).
pubmed: 31257183
pmcid: 6754270
doi: 10.1016/j.chembiol.2019.06.001
Pisa, R. & Kapoor, T. M. Chemical strategies to overcome resistance against targeted anticancer therapeutics. Nat. Chem. Biol. 16, 817–825 (2020).
pubmed: 32694636
doi: 10.1038/s41589-020-0596-8
Chou, T.-F. & Deshaies, R. J. Development of p97 AAA ATPase inhibitors. Autophagy 7, 1091–1092 (2011).
pubmed: 21606684
pmcid: 3210319
doi: 10.4161/auto.7.9.16489
Pisa, R., Cupido, T. & Kapoor, T. M. Designing allele-specific inhibitors of spastin, a microtubule-severing AAA protein. J. Am. Chem. Soc. 141, 5602–5606 (2019).
pubmed: 30875216
pmcid: 6637947
doi: 10.1021/jacs.8b13257
Tang, W. K., Odzorig, T., Jin, W. & Xia, D. Structural basis of p97 inhibition by the site-selective anticancer compound CB-5083. Mol. Pharmacol. 95, 286–293 (2019).
pubmed: 30591537
pmcid: 6355941
doi: 10.1124/mol.118.114256
Niesen, F. H., Berglund, H. & Vedadi, M. The use of differential scanning fluorimetry to detect ligand interactions that promote protein stability. Nat. Protoc. 2, 2212–2221 (2007).
pubmed: 17853878
doi: 10.1038/nprot.2007.321
Lagoutte, R., Patouret, R. & Winssinger, N. Covalent inhibitors: an opportunity for rational target selectivity. Curr. Opin. Chem. Biol. 39, 54–63 (2017).
pubmed: 28609675
doi: 10.1016/j.cbpa.2017.05.008
Garske, A. L., Peters, U., Cortesi, A. T., Perez, J. L. & Shokat, K. M. Chemical genetic strategy for targeting protein kinases based on covalent complementarity. Proc. Natl Acad. Sci. USA 108, 15046–15052 (2011).
pubmed: 21852571
doi: 10.1073/pnas.1111239108
Magnaghi, P. et al. Covalent and allosteric inhibitors of the ATPase VCP/p97 induce cancer cell death. Nat. Chem. Biol. 9, 548–556 (2013).
pubmed: 23892893
doi: 10.1038/nchembio.1313
Kuo, T.-C. et al. Purine-type compounds induce microtubule fragmentation and lung cancer cell death through interaction with katanin. J. Med. Chem. 59, 8521–8534 (2016).
pubmed: 27536893
doi: 10.1021/acs.jmedchem.6b00797
Cheung, K. et al. Proteomic analysis of the mammalian Katanin family of microtubule-severing enzymes defines Katanin p80 subunit B-like 1 (KATNBL1) as a regulator of mammalian Katanin microtubule severing. Mol. Cell. Proteomics 15, 1658–1669 (2016).
pubmed: 26929214
pmcid: 4858946
doi: 10.1074/mcp.M115.056465
Hendershott, M. C. & Vale, R. D. Regulation of microtubule minus-end dynamics by CAMSAPs and patronin. Proc. Natl Acad. Sci. USA 111, 5860–5865 (2014).
pubmed: 24706919
doi: 10.1073/pnas.1404133111
Jiang, K. et al. Structural basis of formation of the microtubule minus-end-regulating CAMSAP–katanin complex. Structure 26, 375–382 (2018).
pubmed: 29395789
doi: 10.1016/j.str.2017.12.017
Jiang, K. et al. Microtubule minus-end stabilization by polymerization-driven CAMSAP deposition. Dev. Cell 28, 295–309 (2014).
pubmed: 24486153
doi: 10.1016/j.devcel.2014.01.001
Kumar, R., Duhamel, M., Coutant, E., Ben-Nahia, E. & Mercier, R. Antagonism between BRCA2 and FIGL1 regulates homologous recombination. Nucleic Acids Res. 47, 5170–5180 (2019).
pubmed: 30941419
pmcid: 6547764
doi: 10.1093/nar/gkz225
Pöhler, R. et al. A non-competitive inhibitor of VCP/p97 and VPS4 reveals conserved allosteric circuits in type I and II AAA ATPases. Angew. Chem. Int. Ed. 57, 1576–1580 (2018).
doi: 10.1002/anie.201711429
Scott, A. et al. Structural and mechanistic studies of VPS4 proteins. EMBO J. 24, 3658–3669 (2005).
pubmed: 16193069
pmcid: 1276703
doi: 10.1038/sj.emboj.7600818
Azmi, I. et al. Recycling of ESCRTs by the AAA-ATPase Vps4 is regulated by a conserved VSL region in Vta1. J. Cell Biol. 172, 705–717 (2006).
pubmed: 16505166
pmcid: 2063703
doi: 10.1083/jcb.200508166
Yang, D. & Hurley, J. H. Structural role of the Vps4-Vta1 interface in ESCRT-III recycling. Structure 18, 976–984 (2010).
pubmed: 20696398
pmcid: 3124813
doi: 10.1016/j.str.2010.04.014
Strelow, J. M. A perspective on the kinetics of covalent and irreversible inhibition. SLAS Discov. 22, 3–20 (2017).
pubmed: 27703080
doi: 10.1177/1087057116671509
Bradshaw, J. M. et al. Prolonged and tunable residence time using reversible covalent kinase inhibitors. Nat. Chem. Biol. 11, 525–531 (2015).
pubmed: 26006010
pmcid: 4472506
doi: 10.1038/nchembio.1817
Knott, G. J. & Doudna, J. A. CRISPR-Cas guides the future of genetic engineering. Science 361, 866–869 (2018).
pubmed: 30166482
pmcid: 6455913
doi: 10.1126/science.aat5011
Dunleavy, J. E. M. et al. Katanin-like 2 (KATNAL2) functions in multiple aspects of haploid male germ cell development in the mouse. PLoS Genet. 13, e1007078 (2017).
pubmed: 29136647
pmcid: 5705150
doi: 10.1371/journal.pgen.1007078
Neale, B. M. et al. Patterns and rates of exonic de novo mutations in autism spectrum disorders. Nature 485, 242–245 (2012).
pubmed: 22495311
pmcid: 3613847
doi: 10.1038/nature11011
Zhang, C. et al. A second-site suppressor strategy for chemical genetic analysis of diverse protein kinases. Nat. Methods 2, 435–441 (2005).
pubmed: 15908922
doi: 10.1038/nmeth764
Ma, J. et al. FIGNL1 is overexpressed in small cell lung cancer patients and enhances NCI-H446 cell resistance to cisplatin and etoposide. Oncol. Rep. 37, 1935–1942 (2017).
pubmed: 28260065
pmcid: 5367342
doi: 10.3892/or.2017.5483
McDonald, E. R. III et al. Project DRIVE: a compendium of cancer dependencies and synthetic lethal relationships uncovered by large-scale, deep RNAi screening. Cell 170, 577–592 (2017).
pubmed: 28753431
doi: 10.1016/j.cell.2017.07.005
Shin, S. H. et al. Synthetic lethality by targeting the RUVBL1/2–TTT complex in mTORC1-hyperactive cancer cells. Sci. Adv. 6, eaay9131 (2020).
pubmed: 32789167
pmcid: 7399646
doi: 10.1126/sciadv.aay9131
Marks, D. H. et al. Mad2 overexpression uncovers a critical role for TRIP13 in mitotic exit. Cell Rep. 19, 1832–1845 (2017).
pubmed: 28564602
pmcid: 5526606
doi: 10.1016/j.celrep.2017.05.021
Uphoff, C. C. & Drexler, H. G. Detection of mycoplasma contaminations. Methods Mol. Biol. 946, 1–13 (2013).
pubmed: 23179822
doi: 10.1007/978-1-62703-128-8_1
Miller, M. S. et al. Getting the most out of your crystals: data collection at the new high-flux, Microfocus MX beamlines at NSLS-II. Molecules 24, 496 (2019).
pmcid: 6384729
doi: 10.3390/molecules24030496
Kabsch, W. XDS. Acta Crystallogr. D Biol. Crystallogr. 66, 125–132 (2010).
pubmed: 20124692
pmcid: 2815665
doi: 10.1107/S0907444909047337
Adams, P. D. et al. PHENIX: a comprehensive Python-based system for macromolecular structure solution. Acta Crystallogr. D Biol. Crystallogr. 66, 213–221 (2010).
pubmed: 20124702
pmcid: 2815670
doi: 10.1107/S0907444909052925
McCoy, A. J. et al. Phaser crystallographic software. J. Appl. Crystallogr. 40, 658–674 (2007).
pubmed: 19461840
pmcid: 2483472
doi: 10.1107/S0021889807021206
Emsley, P., Lohkamp, B., Scott, W. G. & Cowtan, K. Features and development of Coot. Acta Crystallogr. D Biol. Crystallogr. 66, 486–501 (2010).
pubmed: 20383002
pmcid: 2852313
doi: 10.1107/S0907444910007493
Afonine, P. V. et al. Towards automated crystallographic structure refinement with phenix.refine. Acta Crystallogr. D Biol. Crystallogr. 68, 352–367 (2012).
pubmed: 22505256
pmcid: 3322595
doi: 10.1107/S0907444912001308
Moriarty, N. W., Grosse-Kunstleve, R. W. & Adams, P. D. Electronic Ligand Builder and Optimization Workbench (eLBOW): a tool for ligand coordinate and restraint generation. Acta Crystallogr. D Biol. Crystallogr. 65, 1074–1080 (2009).
pubmed: 19770504
pmcid: 2748967
doi: 10.1107/S0907444909029436
Chen, V. B. et al. MolProbity: all-atom structure validation for macromolecular crystallography. Acta Crystallogr. D Biol. Crystallogr. 66, 12–21 (2010).
pubmed: 20057044
doi: 10.1107/S0907444909042073