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Protein & Peptide Letters

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ISSN (Print): 0929-8665
ISSN (Online): 1875-5305

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Research Article

Analysis of the Interaction of UBE2Q1 with B4GALT1 and P53: Experimental and Molecular Modeling Study

Author(s): Hadi Ghasemi, Atefeh Seghatoleslam*, Mohammad Ali Fahmideh Kar, Laleh Mahbudi, Behrouz Gharesi Fard and Mahdi Jamshidi

Volume 30, Issue 8, 2023

Published on: 09 June, 2023

Page: [668 - 678] Pages: 11

DOI: 10.2174/0929866530666230517121827

Price: $65

Abstract

Background: UBE2Q1-dependent ubiquitination of key proteins including β 1,4- galactosyltransferase (GalT1), and P53 might play a pivotal role in cancer development.

Objective: The present study aimed to evaluate the molecular analysis of possible interactions between UBE2Q1 with B4GALT1 and P53 proteins.

Methods: We established SW1116 colorectal cancer cell line stably transfected with UBE2Q1. To verify the overexpression of UBE2Q1, we performed western blot and fluorescent microscopy analysis. Using the immunoprecipitation (IP) product of the over-expressed protein on the silver staining gel, we observed the potential interacting partners of UBE2Q1. The Molecular Operating Environment (MOE) software was also used to perform the molecular docking of the UBC domain of UBE2Q1 (2QGX) with B4GALT1 (2AGD), and P53 (tetramerization (1AIE) and DNA binding domains (1GZH)) proteins.

Results: Western blot and IP analysis detected a UBE2Q1-GFP band in transfected cells, while no band was detected for mock-transfected cells. Moreover, the overexpression of UBE2Q1 tagged with GFP was observed under fluorescent microscopy as well with about 60-70% shining. Silver staining of IP gel revealed several bands in colorectal cancer (CRC) with UBE2Q1 overexpression. Protein- Protein interaction (PPI) analysis also depicted a high affinity of the UBC domain of UBE2Q1 to the B4GALT1 and P53 (tetramerization and DNA binding domains). Molecular docking also revealed hot-spot regions for all poses.

Conclusion: Our data suggest that UBE2Q1 as an E2 enzyme of ubiquitination system can interact with B4GALT1 and P53, and may contribute to the accumulation of misfolded important proteins and colorectal tumor development.

Keywords: Ubiquitination, E2 enzyme, UBE2Q1, immunoprecipitation (IP), B4GALT1, P53, molecular docking.

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[1]
Kraft, E.; Stone, S.L.; Ma, L.; Su, N.; Gao, Y.; Lau, O.S.; Deng, X.W.; Callis, J. Genome analysis and functional characterization of the E2 and RING-type E3 ligase ubiquitination enzymes of Arabidopsis. Plant Physiol., 2005, 139(4), 1597-1611.
[http://dx.doi.org/10.1104/pp.105.067983] [PMID: 16339806]
[2]
Ye, Y.; Rape, M. Building ubiquitin chains: E2 enzymes at work. Nat. Rev. Mol. Cell Biol., 2009, 10(11), 755-764.
[http://dx.doi.org/10.1038/nrm2780] [PMID: 19851334]
[3]
Suzuki, T.; Fujii, A.; Ochi, H.; Nakamura, H. Ubiquitination and downregulation of ErbB2 and estrogen receptor-alpha by kinase inhibitor MP-412 in human breast cancer cells. J. Cell. Biochem., 2011, 112(9), 2279-2286.
[http://dx.doi.org/10.1002/jcb.23147] [PMID: 21503962]
[4]
Ghasemi, H.; Amini, M.A.; Pegah, A.; Azizi, E.; Tayebinia, H.; Khanverdilou, S.; Mousavibahar, S.H.; Alizamir, A. Overexpression of reactive oxygen species modulator 1 is associated with advanced grades of bladder cancer. Mol. Biol. Rep., 2020, 47(9), 6497-6505.
[http://dx.doi.org/10.1007/s11033-020-05702-1] [PMID: 32770525]
[5]
Wijk, S.J.L.; Timmers, H.T.M. The family of ubiquitin-conjugating enzymes (E2s): Deciding between life and death of proteins. FASEB J., 2010, 24(4), 981-993.
[http://dx.doi.org/10.1096/fj.09-136259] [PMID: 19940261]
[6]
Stewart, M.D.; Ritterhoff, T.; Klevit, R.E.; Brzovic, P.S. E2 enzymes: More than just middle men. Cell Res., 2016, 26(4), 423-440.
[http://dx.doi.org/10.1038/cr.2016.35] [PMID: 27002219]
[7]
Gundogdu, M.; Walden, H. Structural basis of generic versus specific E2-RING E3 interactions in protein ubiquitination. Protein Sci., 2019, 28(10), 1758-1770.
[http://dx.doi.org/10.1002/pro.3690]
[8]
Millan, I.C.; Squillace, A.L.; Gava, L.M.; Ramos, C.H. The stability of wild-type and deletion mutants of human C-terminus Hsp70-interacting protein (CHIP). Protein Pept. Lett., 2013, 20(5), 524-529.
[http://dx.doi.org/10.2174/0929866511320050005] [PMID: 22670667]
[9]
Geng, J.; Klionsky, D.J. The Atg8 and Atg12 ubiquitin-like conjugation systems in macroautophagy. EMBO Rep., 2008, 9(9), 859-864.
[http://dx.doi.org/10.1038/embor.2008.163] [PMID: 18704115]
[10]
Slobodkin, M.R.; Elazar, Z.; Elazar, Z. The Atg8 family: Multifunctional ubiquitin-like key regulators of autophagy. Essays Biochem., 2013, 55(1), 51-64.
[PMID: 24070471]
[11]
Pruneda, J.N.; Smith, F.D.; Daurie, A.; Swaney, D.L.; Villén, J.; Scott, J.D.; Stadnyk, A.W.; Le Trong, I.; Stenkamp, R.E.; Klevit, R.E.; Rohde, J.R.; Brzovic, P.S. E2~Ub conjugates regulate the kinase activity of Shigella effector OspG during pathogenesis. EMBO J., 2014, 33(5), 437-449.
[PMID: 24446487]
[12]
Shafiee, S.M.; Seghatoleslam, A.; Nikseresht, M.; Hosseini, S.V.; Alizadeh-Naeeni, M.; Safaei, A.; Owji, A.A. UBE2Q1 expression in human colorectal tumors and cell lines. Mol. Biol. Rep., 2013, 40(12), 7045-7051.
[http://dx.doi.org/10.1007/s11033-013-2824-8] [PMID: 24197692]
[13]
Shafiee, S.M.; Rasti, M.; Seghatoleslam, A.; Azimi, T.; Owji, A.A. UBE2Q1 in a human breast carcinoma cell line: overexpression and interaction with p53. Asian Pac. J. Cancer Prev., 2015, 16(9), 3723-3727.
[http://dx.doi.org/10.7314/APJCP.2015.16.9.3723] [PMID: 25987028]
[14]
Voutsadakis, I.A. Ubiquitin- and ubiquitin-like proteins-conjugating enzymes (E2s) in breast cancer. Mol. Biol. Rep., 2013, 40(2), 2019-2034.
[http://dx.doi.org/10.1007/s11033-012-2261-0] [PMID: 23187732]
[15]
Chang, R.; Wei, L.; Lu, Y.; Cui, X.; Lu, C.; Liu, L.; Jiang, D.; Xiong, Y.; Wang, G.; Wan, C.; Qian, H. Upregulated expression of ubiquitin-conjugating enzyme E2Q1 (UBE2Q1) is associated with enhanced cell proliferation and poor prognosis in human hapatocellular carcinoma. J. Mol. Histol., 2015, 46(1), 45-56.
[http://dx.doi.org/10.1007/s10735-014-9596-x] [PMID: 25311764]
[16]
Seghatoleslam, A.; Nikseresht, M.; Shafiee, S.M.; Monabati, A.; Namavari, M.M.; Talei, A.; Safaei, A.; Owji, A.A. Expression of the novel human gene, UBE2Q1, in breast tumors. Mol. Biol. Rep., 2012, 39(5), 5135-5141.
[http://dx.doi.org/10.1007/s11033-011-1309-x] [PMID: 22167327]
[17]
Zhang, B.; Deng, C.; Wang, L.; Zhou, F.; Zhang, S.; Kang, W.; Zhan, P.; Chen, J.; Shen, S.; Guo, H.; Zhang, M.; Wang, Y.; Zhang, F.; Zhang, W.; Xiao, J.; Kong, B.; Friess, H.; Zhuge, Y.; Yan, H.; Zou, X. Upregulation of UBE2Q1 via gene copy number gain in hepatocellular carcinoma promotes cancer progression through β-catenin-EGFR-PI3K-Akt-mTOR signaling pathway. Mol. Carcinog., 2018, 57(2), 201-215.
[http://dx.doi.org/10.1002/mc.22747] [PMID: 29027712]
[18]
Topno, R.; Singh, I.; Kumar, M.; Agarwal, P. Integrated bioinformatic analysis identifies UBE2Q1 as a potential prognostic marker for high grade serous ovarian cancer. BMC Cancer, 2021, 21(1), 220.
[http://dx.doi.org/10.1186/s12885-021-07928-z] [PMID: 33663405]
[19]
Wassler, M.J.; Shur, B.D.; Zhou, W.; Geng, Y.J. Characterization of a novel ubiquitin-conjugating enzyme that regulates β1,4-galactosyltransferase-1 in embryonic stem cells. Stem Cells, 2008, 26(8), 2006-2018.
[http://dx.doi.org/10.1634/stemcells.2007-1080] [PMID: 18511602]
[20]
Harrus, D.; Khoder-Agha, F.; Peltoniemi, M.; Hassinen, A.; Ruddock, L.; Kellokumpu, S.; Glumoff, T. The dimeric structure of wild-type human glycosyltransferase B4GalT1. PLoS One, 2018, 13(10), e0205571.
[http://dx.doi.org/10.1371/journal.pone.0205571] [PMID: 30352055]
[21]
Han, K.; Jeng, E.E.; Hess, G.T.; Morgens, D.W.; Li, A.; Bassik, M.C. Synergistic drug combinations for cancer identified in a CRISPR screen for pairwise genetic interactions. Nat. Biotechnol., 2017, 35(5), 463-474.
[http://dx.doi.org/10.1038/nbt.3834] [PMID: 28319085]
[22]
Inoue, T.; Geyer, R.K.; Yu, Z.K.; Maki, C.G. Downregulation of MDM2 stabilizes p53 by inhibiting p53 ubiquitination in response to specific alkylating agents. FEBS Lett., 2001, 490(3), 196-201.
[http://dx.doi.org/10.1016/S0014-5793(01)02123-8] [PMID: 11223035]
[23]
D’Anselmi, F.; Cucina, A.; Cavallaro, G.; Bizzarri, M.; Cavallaro, R.; Fuso, A.; Cavallaro, A.; Scarpa, S. S-adenosylmethionine inhibits ubiquitin-proteasome system in vitro and on rat vascular smooth muscle cells. Protein Pept. Lett., 2008, 15(1), 58-62.
[http://dx.doi.org/10.2174/092986608783330396] [PMID: 18221015]
[24]
Joerger, A.C.; Fersht, A.R. The tumor suppressor p53: From structures to drug discovery. Cold Spring Harb. Perspect. Biol., 2010, 2(6), a000919.
[http://dx.doi.org/10.1101/cshperspect.a000919] [PMID: 20516128]
[25]
Brooks, C.L.; Gu, W. p53 regulation by ubiquitin. FEBS Lett., 2011, 585(18), 2803-2809.
[http://dx.doi.org/10.1016/j.febslet.2011.05.022] [PMID: 21624367]
[26]
Ranjan, K.; Pathak, C. Expression of cFLIPL determines the basal interaction of Bcl-2 with beclin-1 and regulates p53 dependent ubiquitination of beclin-1 during autophagic stress. J. Cell. Biochem., 2016, 117(8), 1757-1768.
[http://dx.doi.org/10.1002/jcb.25474] [PMID: 26682748]
[27]
Moll, U.M.; Petrenko, O. The MDM2-p53 interaction. Mol. Cancer Res., 2003, 1(14), 1001-1008.
[PMID: 14707283]
[28]
Saville, M.K.; Sparks, A.; Xirodimas, D.P.; Wardrop, J.; Stevenson, L.F.; Bourdon, J.C.; Woods, Y.L.; Lane, D.P. Regulation of p53 by the ubiquitin-conjugating enzymes UbcH5B/C in vivo. J. Biol. Chem., 2004, 279(40), 42169-42181.
[http://dx.doi.org/10.1074/jbc.M403362200] [PMID: 15280377]
[29]
Tanzadehpanah, H.; Mahaki, H.; Moradi, M.; Afshar, S.; Moghadam, N.H.; Salehzadeh, S.; Najafi, R.; Amini, R.; Saidijam, M. The use of molecular docking and spectroscopic methods for investigation of the interaction between regorafenib with human serum albumin (Hsa) and calf thymus DNA (CT-DNA) in the presence of different site markers. Protein Pept. Lett., 2021, 28(3), 290-303.
[http://dx.doi.org/10.2174/0929866527666200921164536] [PMID: 32957871]
[30]
Ullah, K.; Zubia, E.; Narayan, M.; Yang, J.; Xu, G. Diverse roles of the E2/E3 hybrid enzyme UBE 2O in the regulation of protein ubiquitination, cellular functions, and disease onset. FEBS J., 2019, 286(11), 2018-2034.
[http://dx.doi.org/10.1111/febs.14708] [PMID: 30468556]
[31]
Liu, W.; Tang, X.; Qi, X.; Fu, X.; Ghimire, S.; Ma, R.; Li, S.; Zhang, N.; Si, H. The ubiquitin conjugating enzyme: An important ubiquitin transfer platform in ubiquitin-proteasome system. Int. J. Mol. Sci., 2020, 21(8), 2894-2910.
[http://dx.doi.org/10.3390/ijms21082894] [PMID: 32326224]
[32]
Dove, K.K.; Stieglitz, B.; Duncan, E.D.; Rittinger, K.; Klevit, R.E. Molecular insights into RBR E3 ligase ubiquitin transfer mechanisms. EMBO Rep., 2016, 17(8), 1221-1235.
[http://dx.doi.org/10.15252/embr.201642641] [PMID: 27312108]
[33]
Machida, Y.J.; Machida, Y.; Chen, Y.; Gurtan, A.M.; Kupfer, G.M.; D’Andrea, A.D.; Dutta, A. UBE2T is the E2 in the Fanconi anemia pathway and undergoes negative autoregulation. Mol. Cell, 2006, 23(4), 589-596.
[http://dx.doi.org/10.1016/j.molcel.2006.06.024] [PMID: 16916645]
[34]
Hoege, C.; Pfander, B.; Moldovan, G.L.; Pyrowolakis, G.; Jentsch, S. RAD6-dependent DNA repair is linked to modification of PCNA by ubiquitin and SUMO. Nature, 2002, 419(6903), 135-141.
[http://dx.doi.org/10.1038/nature00991] [PMID: 12226657]
[35]
Pickart, C.M.; Rose, I.A. Functional heterogeneity of ubiquitin carrier proteins. J. Biol. Chem., 1985, 260(3), 1573-1581.
[http://dx.doi.org/10.1016/S0021-9258(18)89632-6] [PMID: 2981864]
[36]
Williams, C.; van den Berg, M.; Sprenger, R.R.; Distel, B. A conserved cysteine is essential for Pex4p-dependent ubiquitination of the peroxisomal import receptor Pex5p. J. Biol. Chem., 2007, 282(31), 22534-22543.
[http://dx.doi.org/10.1074/jbc.M702038200] [PMID: 17550898]
[37]
Léon, S.; Subramani, S. A conserved cysteine residue of Pichia pastoris Pex20p is essential for its recycling from the peroxisome to the cytosol. J. Biol. Chem., 2007, 282(10), 7424-7430.
[http://dx.doi.org/10.1074/jbc.M611627200] [PMID: 17209040]
[38]
Grzmil, P.; Altmann, M.E.; Adham, I.M.; Engel, U.; Jarry, H.; Schweyer, S.; Wolf, S.; Mänz, J.; Engel, W. Embryo implantation failure and other reproductive defects in Ube2q1-deficient female mice. Reproduction, 2013, 145(1), 45-56.
[http://dx.doi.org/10.1530/REP-12-0054] [PMID: 23108111]
[39]
Madden, E.; Logue, S.E.; Healy, S.J.; Manie, S.; Samali, A. The role of the unfolded protein response in cancer progression: From oncogenesis to chemoresistance. Biol. Cell, 2019, 111(1), 1-17.
[http://dx.doi.org/10.1111/boc.201800050] [PMID: 30302777]
[40]
Ghasemi, H.; Mousavibahar, S.H.; Hashemnia, M.; Karimi, J.; Khodadadi, I.; Tavilani, H. Transitional cell carcinoma matrix stiffness regulates the osteopontin and YAP expression in recurrent patients. Mol. Biol. Rep., 2021, 48(5), 4253-4262.
[http://dx.doi.org/10.1007/s11033-021-06440-8] [PMID: 34086159]
[41]
Poeta, M.L.; Massi, E.; Parrella, P.; Pellegrini, P.; De Robertis, M.; Copetti, M.; Rabitti, C.; Perrone, G.; Muda, A.O.; Molinari, F.; Zanellato, E.; Crippa, S.; Caputo, D.; Caricato, M.; Frattini, M.; Coppola, R.; Fazio, V.M. Aberrant promoter methylation of beta-1,4 galactosyltransferase 1 as potential cancer-specific biomarker of colorectal tumors. Genes Chromosomes Cancer, 2012, 51(12), 1133-1143.
[http://dx.doi.org/10.1002/gcc.21998] [PMID: 22927297]
[42]
Harms, K.L.; Chen, X. The functional domains in p53 family proteins exhibit both common and distinct properties. Cell Death Differ., 2006, 13(6), 890-897.
[http://dx.doi.org/10.1038/sj.cdd.4401904] [PMID: 16543939]
[43]
Tanwar, K.; Pati, U. Inhibition of apoptosis via CHIP-mediated proteasomal degradation of TAp73α. J. Cell. Biochem., 2019, 120(7), 11091-11103.
[http://dx.doi.org/10.1002/jcb.28386] [PMID: 30714204]
[44]
Fu, T.; Min, H.; Xu, Y.; Chen, J.; Li, G. Molecular dynamic simulation insights into the normal state and restoration of p53 function. Int. J. Mol. Sci., 2012, 13(8), 9709-9740.
[http://dx.doi.org/10.3390/ijms13089709] [PMID: 22949826]
[45]
Kamada, R.; Nomura, T.; Anderson, C.W.; Sakaguchi, K. Cancer-associated p53 tetramerization domain mutants: Quantitative analysis reveals a low threshold for tumor suppressor inactivation. J. Biol. Chem., 2011, 286(1), 252-258.
[http://dx.doi.org/10.1074/jbc.M110.174698] [PMID: 20978130]
[46]
Rippin, T.M.; Freund, S.M.V.; Veprintsev, D.B.; Fersht, A.R. Recognition of DNA by p53 core domain and location of intermolecular contacts of cooperative binding. J. Mol. Biol., 2002, 319(2), 351-358.
[http://dx.doi.org/10.1016/S0022-2836(02)00326-1] [PMID: 12051912]

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