C-Raf antagonizes apoptosis induced by IFN-alpha in human lung cancer cells by phosphorylation and increase of the intracellular content of elongation factor 1A.
Interferon alpha (IFNalpha) induces both apoptosis and a counteracting epidermal growth factor Erk-dependent survival response in cancer cells. In this report, IFNalpha increased eukaryotic elongation factor 1A (eEF-1A) protein expression by inhibition of eEF-1A degradation via a proteasome-dependent pathway. The reduction of the expression level of eEF-1A by RNA interference enhanced the apoptosis induced by IFNalpha on the same cells. Moreover, IFNalpha induced the phosphorylation of both serine and threonine in eEF-1A. These effects were paralleled by an increased co-immunoprecipitation and colocalization of eEF-1A with C-Raf. The suppression of C-Raf kinase activity with the inhibitor BAY 43-9006 completely antagonized the increase of both eEF-1A phosphorylation and expression and of C-Raf/eEF-1A colocalization induced by IFNalpha and enhanced apoptosis and eEF-1A ubiquitination. Cell transfection with the mutated K48R ubiquitin increased EF-1A expression and desensitized tumor cells to the modulating effects of IFNalpha. The dynamic simulation of 3Dstructure of eEF-1A identified putative serine and threonine phosphorylation sites. In conclusion, the interaction between eEF-1A and C-Raf increases eEF-1A stability and induces a survival activity.
Lamberti
A
A
Department of Biochemistry and Medical Biotechnology, Federico II University of Naples, Naples, Italy.
Longo
O
O
Marra
M
M
Tagliaferri
P
P
Bismuto
E
E
Fiengo
A
A
Viscomi
C
C
Budillon
A
A
Rapp
U R
UR
Wang
E
E
Venuta
S
S
Abbruzzese
A
A
Arcari
P
P
Caraglia
M
M
D017209:Apoptosis / Q000187:drug effects*; D045744:Cell Line, Tumor; D006801:Humans; D047468:Immunoprecipitation; D016898:Interferon-alpha / Q000494:pharmacology*; D008175:Lung Neoplasms / Q000378:metabolism / Q000473:pathology*; D015513:Oncogene Proteins / Q000378:metabolism*; D020648:Peptide Elongation Factor 1 / Q000378:metabolism*; D010766:Phosphorylation / Q000187:drug effects; D010768:Phosphoserine / Q000378:metabolism; D010769:Phosphothreonine / Q000378:metabolism; D046988:Proteasome Endopeptidase Complex / Q000378:metabolism; D011485:Protein Binding / Q000187:drug effects; D011499:Protein Processing, Post-Translational / Q000187:drug effects; D021381:Protein Transport / Q000187:drug effects; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000378:metabolism; D025801:Ubiquitin / Q000378:metabolism
C094814:EEF1A1 protein, human; D016898:Interferon-alpha; D015513:Oncogene Proteins; D020648:Peptide Elongation Factor 1; D034741:RNA, Small Interfering; D025801:Ubiquitin; D010769:Phosphothreonine; D010768:Phosphoserine; D019908:Proto-Oncogene Proteins c-raf; D046988:Proteasome Endopeptidase Complex