Human TRIB2 is a repressor of FOXO that contributes to the malignant phenotype of melanoma cells.
FOXO transcription factors are evolutionarily conserved proteins that orchestrate gene expression programs known to control a variety of cellular processes such as cell cycle, apoptosis, DNA repair and protection from oxidative stress. As the abrogation of FOXO function is a key feature of many tumor cells, regulation of FOXO factors is receiving increasing attention in cancer research. In order to discover genes involved in the regulation of FOXO activity, we performed a large-scale RNA-mediated interference (RNAi) screen using cell-based reporter systems that monitor transcriptional activity and subcellular localization of FOXO. We identified genes previously implicated in phosphoinositide 3-kinase/Akt signaling events, which are known to be important for FOXO function. In addition, we discovered a previously unrecognized FOXO-repressor function of TRIB2, the mammalian homolog of the Drosophila gene tribbles. A cancer-profiling array revealed specific overexpression of TRIB2 in malignant melanoma, but not in other types of skin cancer. We provide experimental evidence that TRIB2 transcript levels correlate with the degree of cytoplasmic localization of FOXO3a. Moreover, we show that TRIB2 is important in the maintenance of the oncogenic properties of melanoma cells, as its silencing reduces cell proliferation, colony formation and wound healing. Tumor growth was also substantially reduced upon RNAi-mediated TRIB2 knockdown in an in vivo melanoma xenograft model. Our studies suggest that TRIB2 provides the melanoma cells with growth and survival advantages through the abrogation of FOXO function. Altogether, our results show the potential of large-scale cell-based RNAi screens to identify promising diagnostic markers and therapeutic targets.
Zanella
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F
Experimental Therapeutics Program, Centro Nacional de Investigaciones Oncologicas (CNIO), Madrid 28029, Spain.
Renner
O
O
García
B
B
Callejas
S
S
Dopazo
A
A
Peregrina
S
S
Carnero
A
A
Link
W
W
D000818:Animals; D017209:Apoptosis; D014408:Biomarkers, Tumor / Q000235:genetics / Q000378:metabolism; D015153:Blotting, Western; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D049109:Cell Proliferation; D002478:Cells, Cultured; D003114:Colony-Forming Units Assay; D003593:Cytoplasm / Q000378:metabolism; D000071161:Forkhead Box Protein O1; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors / Q000235:genetics / Q000378:metabolism*; D020869:Gene Expression Profiling; D006801:Humans; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics / Q000378:metabolism*; D008156:Luciferases / Q000378:metabolism; D008545:Melanoma / Q000235:genetics / Q000378:metabolism* / Q000473:pathology; D051379:Mice; D016513:Mice, SCID; D020411:Oligonucleotide Array Sequence Analysis; D010641:Phenotype; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D034741:RNA, Small Interfering / Q000494:pharmacology; D012097:Repressor Proteins / Q000235:genetics / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014945:Wound Healing
D014408:Biomarkers, Tumor; C085126:FOXO1 protein, human; C499517:FOXO3 protein, human; D000071161:Forkhead Box Protein O1; D000071316:Forkhead Box Protein O3; D051858:Forkhead Transcription Factors; D047908:Intracellular Signaling Peptides and Proteins; D012333:RNA, Messenger; D034741:RNA, Small Interfering; D012097:Repressor Proteins; D008156:Luciferases; D017871:Calcium-Calmodulin-Dependent Protein Kinases; C489044:TRIB2 protein, human