1 Sorbonne Université - Campus Pierre et Marie Curie, Laboratoire SU-UR6 “Enzymology of RNA” (Bât. B); Case Courrier 60 - 4, Place Jussieu, F-75252, Paris Cedex 05, France
2 Laboratory of Biochemistry and Molecular Biology, Institute of Biomedical Sciences, University of Abomey-Calavi, Cotonou, Benin
3 Sorbonne Université - Campus Pierre et Marie Curie, Institut de Biologie Paris Seine (IBPS) Plateforme d’interactions moléculaires, CNRS-FR3631; 7, Quai Saint Bernard, F-75252, Paris Cedex 05, France
4 Laboratory of Medicinal Organic Chemistry (MOCL), Faculty of Health Sciences, 01 BP: 188 Cotonou, Benin
5 Division of Chromatin Regulation, National Institute for Basic Biology, Nishigonaka 38, Myodaiji, Okazaki 444-8585, Japan
6 Sorbonne Université; CNRS Unit Biological Adaptation and Ageing, Photobiology Team, F-75252, Paris Cedex 05, France
We have recently demonstrated that the eukaryote-specific large subunit ribosomal protein
(rp) eL42 assists catalysis of peptide bond formation at the peptidyl transferase center of 80S
ribosomes in eukaryotic cells. Recently, several ribosomal proteins were shown to have extraribosomal
functions independent of protein biosynthesis. Such functions include regulation of
apoptosis, cell cycle arrest, cell proliferation, neoplastic transformation, cell migration and
invasion, and tumorigenesis through both Mdm2-p53-dependent and p53-independent
mechanisms. Our objective is to demonstrate that overexpression of eL42 in tumor may
incapacitate cell anti-tumor mechanism through interaction with the tumor suppressor protein
p53 and its partner Mdm2.
Co-immunoprecipitation technique and the binding assays on Biacore were used to
probe interactions between recombinant eL42, p53 and Mdm2 proteins in a so-called rp-p53-Mdm2 axis.
We demonstrate that the ribosomal protein eL42, the tumor suppressor protein p53 and the ubiquitin E3 ligase Mdm2 interact with each other in a ternary rp.eL42:p53:Mdm2 complex. Precisely, the interaction between eL42 and p53 is characterized by a strong binding affinity (KD value in the nanomolar range) that is likely to trigger the sequestration of p53 and the inhibition of its tumor suppressor activity. Furthermore, the p53:Mdm2 and eL42:Mdm2 complexes exhibit comparable binding affinities in the micromolar range compatible with Mdm2 being the enzyme which ubiquitinates both the p53 and eL42 substrates. Interestingly, pyridoxal 5'-phosphate (PLP), one of the active forms of vitamin B6, binds to eL42 and significantly inhibits the interaction between eL42 and p53, in accordance with the observation that vitamin B6 is associated with reduced risk of cancer.
Our study emphasized one more major mechanism of p53 downregulation involving its sequestration by eL42 upon the overexpression of this ribosomal protein. The mechanism described in the present report complemented the well-known p53 downregulation triggered by proteasomal degradation mediated through its ubiquitination by Mdm2.
Keywords: eL42 protein, Human 80S ribosomes, Tumor suppressor p53, Ubiquitin E3 ligase Mdm2, eL42:p53 and eL42:Mdm2 complexes, Cancer-pertinent rp.eL42-p53-Mdm2 pathway, Binding assays on Biacore, Co-immunoprecipitation.
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* Address correspondence to these authors at the Sorbonne Université; Campus Pierre et Marie Curie, Laboratoire “Enzymologie de l’ARN”, SU-UR6; Case courrier 60 - 4, Place Jussieu, F-75251, Paris, Cedex 05 France; E-mail: email@example.com; Sorbonne Université, CNRS Unit Biological Adaptation and Ageing, Photobiology Team, Paris, France; F-75252, Paris Cedex 05, France; E-mail: Nathalie.firstname.lastname@example.org