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SAMBA, a Plant-Specific Anaphase-Promoting Complex/Cyclosome Regulator Is Involved in Early Development and A-Type Cyclin Stabilization Nubia B

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SAMBA, a plant-specific anaphase-promoting complex/cyclosome regulator is involved in early development and A-type cyclin stabilization Nubia B. Eloy, Nathalie Gonzalez, Jelle van Leene, Katrien Maleux, Hannes Vanhaeren, Liesbeth de Milde, Stijn Dhondt, Leen Vercruysse, Erwin Witters, Raphaël Mercier, et al. To cite this version: Nubia B. Eloy, Nathalie Gonzalez, Jelle van Leene, Katrien Maleux, Hannes Vanhaeren, et al.. SAMBA, a plant-specific anaphase-promoting complex/cyclosome regulator is involved in early de- velopment and A-type cyclin stabilization. Proceedings of the National Academy of Sciences of the United States of America , National Academy of Sciences, 2012, 109 (34), pp.13853 - 13858. 10.1073/pnas.1211418109. hal-01190736 HAL Id: hal-01190736 https://hal.archives-ouvertes.fr/hal-01190736 Submitted on 29 May 2020 HAL is a multi-disciplinary open access L’archive ouverte pluridisciplinaire HAL, est archive for the deposit and dissemination of sci- destinée au dépôt et à la diffusion de documents entific research documents, whether they are pub- scientifiques de niveau recherche, publiés ou non, lished or not. The documents may come from émanant des établissements d’enseignement et de teaching and research institutions in France or recherche français ou étrangers, des laboratoires abroad, or from public or private research centers. publics ou privés. SAMBA, a plant-specific anaphase-promoting complex/ cyclosome regulator is involved in early development and A-type cyclin stabilization Nubia B. Eloya,b, Nathalie Gonzaleza,b, Jelle Van Leenea,b, Katrien Maleuxa,b, Hannes Vanhaerena,b, Liesbeth De Mildea,b, Stijn Dhondta,b, Leen Vercruyssea,b, Erwin Wittersc,d,e, Raphaël Mercierf, Laurence Cromerf, Gerrit T. S. Beemsterd, Han Remautg, Marc C. E. Van Montagub,1, Geert De Jaegera,b, Paulo C. G. Ferreirah, and Dirk Inzéa,b,1 aDepartment of Plant Systems Biology, VIB, and bDepartment of Plant Biotechnology and Bioinformatics, Ghent University, 9052 Ghent, Belgium; cCenter for Proteome Analysis and Mass Spectrometry and dDepartment of Biology, University of Antwerp, 2020 Antwerp, Belgium; eFlemish Institute for Technological Research (VITO), 2400 Mol, Belgium; fInstitut Jean-Pierre Bourgin, Unité Mixte de Recherche 1318, Institut National de la Recherche Agronomique– AgroParisTech, 78026 Versailles, France; gLaboratory of Structural and Molecular Microbiology, VIB–Vrije Universiteit Brussel, 1050 Brussels, Belgium; and hInstituto de Bioquímica Médica, Centro de Ciências da Saúde, Universidade Federal do Rio de Janeiro, CEP 21941-590, Rio de Janeiro, Brazil Contributed by Marc C. E. Van Montagu, July 11, 2012 (sent for review February 23, 2012) The anaphase-promoting complex/cyclosome (APC/C) is a large determine substrate specificity (21). Recently, ULTRAVIOLET- multiprotein E3 ubiquitin ligase involved in ubiquitin-dependent B-INSENSITIVE4 (UVI4) and UVI4-like/OMISSION OF proteolysis of key cell cycle regulatory proteins, including the SECOND DIVISION1/GIGAS CELL1 (UVI4/OSD1/GIG1) destruction of mitotic cyclins at the metaphase-to-anaphase transi- were identified as plant-specific inhibitors of the APC/C that are tion. Despite its importance, the role of the APC/C in plant cells and required for proper mitotic progression in Arabidopsis (22, 23). the regulation of its activity during cell division remain poorly In Arabidopsis, the genes encoding the subunits APC2, APC3a, understood. Here, we describe the identification of a plant-specific APC3b, APC4, APC6, APC8,andAPC10 have been investigated negative regulator of the APC/C complex, designated SAMBA. In functionally. In all cases, except for APC8, the analysis of the Arabidopsis thaliana, SAMBA is expressed during embryogenesis mutants revealed female gametophytic defects, probably as a con- – APC8 and early plant development and plays a key role in organ size sequence of the inability to degrade mitotic cyclins (24 28). control. Samba mutants produced larger seeds, leaves, and roots, was shown to be involved in male gametogenesis (29). Moreover, APC6 APC10 which resulted from enlarged root and shoot apical meristems, reduced expression levels of (10) or (10, 28) ex- and, additionally, they had a reduced fertility attributable to a ham- hibited several developmental abnormalities, including defects in SAMBA vascular development, whereas an APC4 loss-of-function mutant pered male gametogenesis. Inactivation of stabilized A2- APC3b CDC27b type cyclins during early development. Our data suggest that was defective in embryogenesis (26). ( ) has a role SAMBA regulates cell proliferation during early development by in the maintenance of cell division in meristems during post- APC10 PLANT BIOLOGY targeting CYCLIN A2 for APC/C-mediated proteolysis. embryonic development (30), and overexpression of en- hances cell division and accelerates the degradation of the mitotic fi CYCB1;1, leading to increased leaf sizes (28). lant organ size is determined by the total cell number and nal Here, we functionally analyzed an APC/C regulator of Arabi- cell size, resulting from cell division and cell expansion, re- dopsis samba P fi , designated SAMBA. The mutants have an enlarged spectively. In most, but not all, cases, the nal organ size correlates meristem size and show growth-related phenotypes, including the with cell number, rendering cell division the main driver that formation of large seeds, leaves, and roots; additionally, their controls growth (1). fertility is reduced because of a defect in male gametogenesis. A In all eukaryotes, unidirectional cell cycle progression requires biochemical analysis revealed that loss of function of SAMBA the coordinated destruction of essential cell cycle regulatory stabilizes CYCA2;3. We conclude that SAMBA is a plant-specific proteins by ubiquitin-dependent proteolysis pathways (2–5). Specific E3 ubiquitin ligases mediate the recognition of target negative regulator of APC/C involved in the degradation of A- proteins (6–8) that are subsequently polyubiquitinated and sub- type cyclins. jected to proteolysis by the 26S proteasome. One of the most Results complex ubiquitin ligases involved in cell cycle control is the anaphase-promoting complex/cyclosome (APC/C), of which SAMBA Is a Plant-Specific APC/C Regulator. The SAMBA protein, fi fi the composition can vary from 11 to 13 subunits depending on the encoded by AT1G32310, had been identi ed by tandem-af nity organism. The APC/C complex plays essential roles in mitosis, purification (TAP) associated with the core APC/C and, more meiosis, and postmitotically differentiated cells (3, 9–12). specifically, with the subunits APC3b, APC7, and APC10 in The APC/C triggers the metaphase-to-anaphase transition and protein complexes purified from Arabidopsis cell suspension the exit from mitosis by mediating the degradation of proteins cultures (31). In a TAP experiment on Arabidopsis cell cultures such as securin and mitotic cyclins (13). In plants, the A- and B- with SAMBA as bait, 12 interacting proteins were identified, type cyclins are subjected to proteolysis by APC/C through rec- including the subunits APC1, APC2, APC3b, APC4, APC5, ognition of specific amino acid motifs, the destruction (D) and APC6, APC7, APC8, and APC10, except APC11, and the reg- KEN boxes (6, 14, 15). Plant A-type cyclins are produced and ulators CCS52A2, UVI4, and UVI4-like/OSD1/GIG1 (31). degraded earlier in the cell cycle than B-type cyclins and have distinct and nonredundant functions in the progression of cell division (16). Based on their primary structures, the plant A-type Author contributions: N.B.E., M.C.E.V.M., and D.I. designed research; N.B.E., N.G., J.V.L., cyclins are classified into A1, A2, and A3 groups (17). The K.M., H.V., L.D.M., L.V., and L.C. performed research; N.B.E., N.G., S.D., E.W., R.M., G.T.S.B., transcriptional regulation of the CYCLIN A2 (CYCA2) group H.R., G.D.J., and P.C.G.F. analyzed data; and N.B.E. and D.I. wrote the paper. coordinates cell proliferation during plant development (18, The authors declare no conflict of interest. 19) and CYCA2;3 overexpression enhances cell division (20). 1To whom correspondence may be addressed. E-mail: [email protected] or The APC/C is regulated partly by two activating proteins [email protected]. CELL DIVISION CYCLE 20 (CDC20) and CDC20 HOMOL- This article contains supporting information online at www.pnas.org/lookup/suppl/doi:10. OGY1/CELL CYCLE SWITCH 52 (CDH1/CCS52), that also 1073/pnas.1211418109/-/DCSupplemental. www.pnas.org/cgi/doi/10.1073/pnas.1211418109 PNAS | August 21, 2012 | vol. 109 | no. 34 | 13853–13858 To confirm that SAMBA forms a complex with APC/C in SAMBA expression was very weak in all tissues, except during seed planta, TAP was carried out with 6-d-old SAMBA-expressing development. To study the expression pattern of SAMBA, a 1.8-kb seedlings C-terminally fused to the GS tag (see SI Materials and fragment upstream of the ATG codon of the SAMBA gene was Methods) and proteins were identified by mass spectrometry fused to a β-glucuronidase (GUS)-GFP tandem reporter cassette (MS). In agreement with previous results, SAMBA interacted and introduced into Arabidopsis plants. SAMBA expression was with all APC/C subunits in planta, except APC11, which is dif- high during embryogenesis (Fig. 1 B and C) but decreased grad- ficult to identify with the selected analytical approach because of ually when seedlings germinated (Fig. 1 D and F). At 3 d after its small size (Table S1). Also, the interaction with the activator stratification (DAS) (Fig. 1D), GUS staining was still well visible in CCS52A2 was found, but not with CCS52B, UVI4, and UVI4- all tissues; at 5 DAS, it diminished and became patchy in root like/OSD1/GIG1. tissues (Fig. 1E); finally, at 8 DAS, it was present only in the Although TAP allowed the detection of the entire APC/C hypocotyls (Fig. 1F); and, at later stages of development, the ex- complex, it did not provide insight into the direct interactions pression was only observed in pollen grains (Fig.
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  • Regulation of Cyclin-Dependent Kinase Activity During Mitotic Exit and Maintenance of Genome Stability by P21, P27, and P107

    Regulation of Cyclin-Dependent Kinase Activity During Mitotic Exit and Maintenance of Genome Stability by P21, P27, and P107

    Regulation of cyclin-dependent kinase activity during mitotic exit and maintenance of genome stability by p21, p27, and p107 Taku Chibazakura*†, Seth G. McGrew‡§, Jonathan A. Cooper§, Hirofumi Yoshikawa*, and James M. Roberts‡§ *Deparment of Bioscience, Tokyo University of Agriculture, 1-1-1 Sakuragaoka, Setagaya-ku, Tokyo 156-8502, Japan; and ‡Howard Hughes Medical Institute and §Division of Basic Sciences, Fred Hutchinson Cancer Research Center, Seattle, WA 98019 Communicated by Robert N. Eisenman, Fred Hutchinson Cancer Research Center, Seattle, WA, February 4, 2004 (received for review October 28, 2003) To study the regulation of cyclin-dependent kinase (CDK) activity bind to and inactivate mitotic cyclin–CDK complexes (15, 16). during mitotic exit in mammalian cells, we constructed murine cell These CKIs accumulate and persist during mid-M-to-G1 phase ͞ lines that constitutively express a stabilized mutant of cyclin A until they are phosphorylated by Sic1 Rum1-resistant G1 cyclin- (cyclin A47). Even though cyclin A47 was expressed throughout CDKs, which initiates their ubiquitin-dependent degradation at mitosis and in G1 cells, its associated CDK activity was inactivated the G1-to-S phase transition (17–19). Thus, Sic1 and Rum1 after the transition from metaphase to anaphase. Cyclin A47 constitute a switch that controls the transition from a state of low associated with both p21 and p27 during mitotic exit, implicating CDK activity to that of high CDK activity, thereby regulating these proteins in CDK inactivation. However, cyclin A47 was fully mitotic exit and S phase entry. This parallels the activity of ؊/؊ ؊/؊ inhibited during the M-to-G1 transition in p21 p27 fibro- APC-Cdh1, and indeed these two pathways constitute redundant blasts.