The 5'-untranslated region of p16INK4a melanoma tumor suppressor acts as a cellular IRES, controlling mRNA translation under hypoxia through YBX1 binding.
CDKN2A/p16INK4a is an essential tumor suppressor gene that controls cell cycle progression and replicative senescence. It is also the main melanoma susceptibility gene. Here we report that p16INK4a 5'UTR mRNA acts as a cellular Internal Ribosome Entry Site (IRES). The potential for p16INK4a 5'UTR to drive cap-independent translation was evaluated by dual-luciferase assays using bicistronic and monocistronic vectors. Results of reporters' relative activities coupled to control analyses for actual bicistronic mRNA transcription, indicated that the wild type p16INK4a 5'UTR could stimulate cap-independent translation. Notably, hypoxic stress and the treatment with mTOR inhibitors enhanced the translation-stimulating property of p16INK4a 5'UTR. RNA immunoprecipitation performed in melanoma-derived SK-Mel-28 and in a patient-derived lymphoblastoid cell line indicated that YBX1 can bind the wild type p16INK4a mRNA increasing its translation efficiency, particularly during hypoxic stress. Modulation of YBX1 expression further supported its involvement in cap-independent translation of the wild type p16INK4a but not a c.-42T>A variant. RNA SHAPE assays revealed local flexibility changes for the c.-42T>A variant at the predicted YBX1 binding site region. Our results indicate that p16INK4a 5'UTR contains a cellular IRES that can enhance mRNA translation efficiency, in part through YBX1.
Laboratory of Transcriptional Networks, Centre for Integrative Biology, CIBIO, University of Trento, Trento, Italy.
Alessandra
A
Bisio
Laboratory of Genomic Screening, Centre for Integrative Biology, CIBIO, University of Trento, Trento, Italy.
Elisa
E
Latorre
Laboratory of Genetics of Rare Hereditary Cancers, DiMI, University of Genoa, Italy and IRCCS AOU San Martino-IST, Genoa, Italy.
Virginia
V
Andreotti
Service de Génétique, Institut Gustave Roussy, Villejuif, France.
Brigitte
B
Bressac-de Paillerets
Section of Epidemiology and Biostatistics, Leeds Institute of Cancer and Pathology, University of Leeds, Leeds, UK.
Mark
M
Harland
Laboratory of Genetics of Rare Hereditary Cancers, DiMI, University of Genoa, Italy and IRCCS AOU San Martino-IST, Genoa, Italy.
Giovanna Bianchi
GB
Scarra
Laboratory of Genetics of Rare Hereditary Cancers, DiMI, University of Genoa, Italy and IRCCS AOU San Martino-IST, Genoa, Italy.
Paola
P
Ghiorzo
Department of Pharmaceutical Sciences, University of California, Irvine, CA, USA.
Robert C
RC
Spitale
Laboratory of Genomic Screening, Centre for Integrative Biology, CIBIO, University of Trento, Trento, Italy.
Alessandro
A
Provenzani
Laboratory of Transcriptional Networks, Centre for Integrative Biology, CIBIO, University of Trento, Trento, Italy.
Alberto
A
Inga
D020121:5' Untranslated Regions / Q000235:genetics*; D015153:Blotting, Western; D015687:Cell Hypoxia; D045744:Cell Line, Tumor; D019941:Cyclin-Dependent Kinase Inhibitor p16 / Q000235:genetics* / Q000378:metabolism; D015972:Gene Expression Regulation, Neoplastic / Q000187:drug effects; D045325:HCT116 Cells; D006801:Humans; D000067556:Internal Ribosome Entry Sites / Q000235:genetics*; D061986:MCF-7 Cells; D008545:Melanoma / Q000235:genetics / Q000378:metabolism / Q000473:pathology; D009154:Mutation; D009287:Naphthyridines / Q000494:pharmacology; D011485:Protein Binding; D014176:Protein Biosynthesis; D034622:RNA Interference; D012333:RNA, Messenger / Q000235:genetics*; D020123:Sirolimus / Q000494:pharmacology; D058570:TOR Serine-Threonine Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism; D051840:Y-Box-Binding Protein 1 / Q000235:genetics* / Q000378:metabolism
C553294:1-(4-(4-propionylpiperazin-1-yl)-3-(trifluoromethyl)phenyl)-9-(quinolin-3-yl)benzo(h)(1,6)naphthyridin-2(1H)-one; D020121:5' Untranslated Regions; D019941:Cyclin-Dependent Kinase Inhibitor p16; D000067556:Internal Ribosome Entry Sites; D009287:Naphthyridines; D012333:RNA, Messenger; D051840:Y-Box-Binding Protein 1; C070856:YBX1 protein, human; D058570:TOR Serine-Threonine Kinases; D020123:Sirolimus
IRES; YBX1; hypoxia; melanoma; p16INK4a