Abstract
Lysosomes, as primary degradative organelles, are the endpoint of different converging pathways, including macroautophagy. To date, lysosome degradative function has been mainly studied in interphase cells, while their role during mitosis remains controversial. Mitosis dictates the faithful transmission of genetic material among generations, and perturbations of mitotic division lead to chromosomal instability, a hallmark of cancer. Heretofore, correct mitotic progression relies on the orchestrated degradation of mitotic factors, which was mainly attributed to ubiquitin-triggered proteasome-dependent degradation. Here, we show that mitotic transition also relies on lysosome-dependent degradation, as impairment of lysosomes increases mitotic timing and leads to mitotic errors, thus promoting chromosomal instability. Furthermore, we identified several putative lysosomal targets in mitotic cells. Among them, WAPL, a cohesin regulatory protein, emerged as a novel SQSTM1-interacting protein for targeted lysosomal degradation. Finally, we characterized an atypical nuclear phenotype, the toroidal nucleus, as a novel biomarker for genotoxic screenings. Our results establish lysosome-dependent degradation as an essential event to prevent chromosomal instability. Abbreviations: 3D: three-dimensional; APC/C: anaphase-promoting complex; ARL8B: ADP ribosylation factor like GTPase 8B; ATG: autophagy-related; BORC: BLOC-one-related complex; CDK: cyclin-dependent kinase; CENPE: centromere protein E; CIN: chromosomal instability; ConcA: concanamycin A; CQ: chloroquine; DAPI: 4,6-diamidino-2-penylinole; FTI: farnesyltransferase inhibitors; GFP: green fluorescent protein; H2B: histone 2B; KIF: kinesin family member; LAMP2: lysosomal associated membrane protein 2; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MEF: mouse embryonic fibroblast; MTOR: mechanistic target of rapamycin kinase; PDS5B: PDS5 cohesin associated factor B; SAC: spindle assembly checkpoint; PLEKHM2: pleckstrin homology and RUN domain containing M2; SQSTM1: sequestosome 1; TEM: transmission electron microscopy; ULK1: unc-51 like autophagy activating kinase 1; UPS: ubiquitin-proteasome system; v-ATPase: vacuolar-type H+-translocating ATPase; WAPL: WAPL cohesion release factor.
| Original language | English (US) |
|---|---|
| Pages (from-to) | 796-813 |
| Number of pages | 18 |
| Journal | Autophagy |
| Volume | 17 |
| Issue number | 3 |
| DOIs | |
| State | Published - Jun 23 2020 |
Bibliographical note
Funding Information:This work was supported by the Agència de Gestió d’Ajuts Universitaris i de Recerca [2017SGR1743]; H2020 Marie Skłodowska-Curie Actions [799000]; Instituto de Salud Carlos III [RD12/0036/0049]; Ministerio de Ciencia, Innovación y Universidades [IJCI-2015-24716]; Ministerio de Economía, Industria y Competitividad, Gobierno de España [SAF2017-85561-R]; Ministerio de Educación, Cultura y Deporte [FPU13/05400]; National Institutes of Health [RO1-HL125353]; Mayo Clinic/National Institutes of Health training grant [5-T32-CA217836-02]. We thank Dr. George Thomas, Dr. Sara Kozma, Dr. Antonio Gentilella and the rest of the members of the laboratory of Cancer Metabolism for scientific inputs and sharing reagents. We also would like to thank the undergraduate students Júlia Granell, Belen Hernandez and Laia Jordana for their punctual participation in the research project. We thank Dr. Neus Agell for providing H2B-GFP stable U2OS cells, Dr. Patricia Boya and Dr. Antonio Zorzano for MEF WT and atg5 -/- and Dr. Cristina Muñoz for siRNAs siATG5 and siSQSTM1. We are grateful to Dr. Juan S. Bonifacino for sharing KIF5A-GFP plasmid and to Dr. Fangwei Wang for WAPL-GFP plasmid. We also thank the technical facilities at CCITUB for FACS analysis and confocal microscopy. We acknowledge the proteomic facility at IDIBELL for their help with the mass spectrometry experiment. We are grateful to Dr. Thomas Neufeld, Dr. Andrew Stephens and Dr. Terje Johansen for their constructive comments on the manuscript. We thank Dr. Carles Pons for his computational support during the revision. This study was supported by grants to A.T. from Ministerio de Economía, Industria y Competitividad (SAF2017-85561-R), which is part of Agencia Estatal de Investigación (Co-funded by European Regional Development Fund. ERDF, a way to build Europe), by joint grants to the Laboratory of Cancer Metabolism from Instituto de Salud Carlos III-Red Temática de Investigación Cooperativa en Cáncer (RD12/0036/0049), and from Generalitat de Catalunya- Suport als Grups de Recerca de Catalunya (2017SGR1743). E.A. was supported by Ministerio de Educación, Cultura y Deporte (FPU13/05400) and SAF2017-85561-R. C.D. was supported by Mayo Clinic/NIH training grant 5-T32-CA217836-02 and NIH grant RO1-HL125353 (through Edward Hinchcliffe). C.M. was supported by Juan de la Cierva fellowship (IJCI-2015-24716) from Ministerio de Ciencia, Innovación y Universidades and by European Union’s Horizon 2020 research and innovation program under the Marie Sklodowska-Curie grant agreement (M-Lysosomes, 799000). The authors declare no competing financial interests. We thank CERCA Program/Generalitat de Catalunya for institutional support to IDIBELL.
Funding Information:
We thank Dr. George Thomas, Dr. Sara Kozma, Dr. Antonio Gentilella and the rest of the members of the laboratory of Cancer Metabolism for scientific inputs and sharing reagents. We also would like to thank the undergraduate students Júlia Granell, Belen Hernandez and Laia Jordana for their punctual participation in the research project. We thank Dr. Neus Agell for providing H2B-GFP stable U2OS cells, Dr. Patricia Boya and Dr. Antonio Zorzano for MEF WT and atg5 and Dr. Cristina Muñoz for siRNAs siATG5 and siSQSTM1. We are grateful to Dr. Juan S. Bonifacino for sharing KIF5A-GFP plasmid and to Dr. Fangwei Wang for WAPL-GFP plasmid. We also thank the technical facilities at CCITUB for FACS analysis and confocal microscopy. We acknowledge the proteomic facility at IDIBELL for their help with the mass spectrometry experiment. We are grateful to Dr. Thomas Neufeld, Dr. Andrew Stephens and Dr. Terje Johansen for their constructive comments on the manuscript. We thank Dr. Carles Pons for his computational support during the revision. This study was supported by grants to A.T. from Ministerio de Economía, Industria y Competitividad (SAF2017-85561-R), which is part of Agencia Estatal de Investigación (Co-funded by European Regional Development Fund. ERDF, a way to build Europe), by joint grants to the Laboratory of Cancer Metabolism from Instituto de Salud Carlos III-Red Temática de Investigación Cooperativa en Cáncer (RD12/0036/0049), and from Generalitat de Catalunya- Suport als Grups de Recerca de Catalunya (2017SGR1743). E.A. was supported by Ministerio de Educación, Cultura y Deporte (FPU13/05400) and SAF2017-85561-R. C.D. was supported by Mayo Clinic/NIH training grant 5-T32-CA217836-02 and NIH grant RO1-HL125353 (through Edward Hinchcliffe). C.M. was supported by Juan de la Cierva fellowship (IJCI-2015-24716) from Ministerio de Ciencia, Innovación y Universidades and by European Union’s Horizon 2020 research and innovation program under the Marie Sklodowska-Curie grant agreement (M-Lysosomes, 799000). The authors declare no competing financial interests. We thank CERCA Program/Generalitat de Catalunya for institutional support to IDIBELL. -/-
Publisher Copyright:
© 2020 The Author(s). Published by Informa UK Limited, trading as Taylor & Francis Group.
Keywords
- Chromosomal instability
- chromosomes segregation
- lysosome
- mitosis
- selective autophagy
- toroidal nucleus
