A large scale shRNA barcode screen identifies the circadian clock component ARNTL as putative regulator of the p53 tumor suppressor pathway.
BACKGROUND
The p53 tumor suppressor gene is mutated in about half of human cancers, but the p53 pathway is thought to be functionally inactivated in the vast majority of cancer. Understanding how tumor cells can become insensitive to p53 activation is therefore of major importance. Using an RNAi-based genetic screen, we have identified three novel genes that regulate p53 function.


RESULTS
We have screened the NKI shRNA library targeting 8,000 human genes to identify modulators of p53 function. Using the shRNA barcode technique we were able to quickly identify active shRNA vectors from a complex mixture. Validation of the screening results indicates that the shRNA barcode technique can reliable identify active shRNA vectors from a complex pool. Using this approach we have identified three genes, ARNTL, RBCK1 and TNIP1, previously unknown to regulate p53 function. Importantly, ARNTL (BMAL1) is an established component of the circadian regulatory network. The latter finding adds to recent observations that link circadian rhythm to the cell cycle and cancer. We show that cells having suppressed ARNTL are unable to arrest upon p53 activation associated with an inability to activate the p53 target gene p21(CIP1).


CONCLUSIONS
We identified three new regulators of the p53 pathway through a functional genetic screen. The identification of the circadian core component ARNTL strengthens the link between circadian rhythm and cancer.
Mullenders
Jasper
J
Division of Molecular Carcinogenesis, Centre for Biomedical Genetics and Cancer Genomics Centre, Netherlands Cancer Institute, Amsterdam, The Netherlands.
Fabius
Armida W M
AW
Madiredjo
Mandy
M
Bernards
René
R
Beijersbergen
Roderick L
RL
D056930:ARNTL Transcription Factors; D051792:Basic Helix-Loop-Helix Proteins / Q000235:genetics / Q000502:physiology*; D002455:Cell Division; D002460:Cell Line; D002940:Circadian Rhythm / Q000235:genetics; D050759:Cyclin-Dependent Kinase Inhibitor p21 / Q000096:biosynthesis / Q000235:genetics; D004268:DNA-Binding Proteins / Q000235:genetics / Q000502:physiology; D015723:Gene Library; D016158:Genes, p53; D005822:Genetic Vectors / Q000235:genetics; D006801:Humans; D055029:Inverted Repeat Sequences; D015345:Oligonucleotide Probes; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics* / Q000302:isolation & purification; D014157:Transcription Factors / Q000235:genetics / Q000502:physiology; D016159:Tumor Suppressor Protein p53 / Q000502:physiology*; D044767:Ubiquitin-Protein Ligases
D056930:ARNTL Transcription Factors; D051792:Basic Helix-Loop-Helix Proteins; D050759:Cyclin-Dependent Kinase Inhibitor p21; D004268:DNA-Binding Proteins; D015345:Oligonucleotide Probes; D034741:RNA, Small Interfering; D014157:Transcription Factors; D016159:Tumor Suppressor Protein p53; D044767:Ubiquitin-Protein Ligases; C538960:BMAL1 protein, human; C493956:CDKN1A protein, human; C117046:TNIP1 protein, human; C504507:RBCK1 protein, human