Transcriptional role of cyclin D1 in development revealed by a genetic-proteomic screen.
Cyclin D1 belongs to the core cell cycle machinery, and it is frequently overexpressed in human cancers. The full repertoire of cyclin D1 functions in normal development and oncogenesis is unclear at present. Here we developed Flag- and haemagglutinin-tagged cyclin D1 knock-in mouse strains that allowed a high-throughput mass spectrometry approach to search for cyclin D1-binding proteins in different mouse organs. In addition to cell cycle partners, we observed several proteins involved in transcription. Genome-wide location analyses (chromatin immunoprecipitation coupled to DNA microarray; ChIP-chip) showed that during mouse development cyclin D1 occupies promoters of abundantly expressed genes. In particular, we found that in developing mouse retinas-an organ that critically requires cyclin D1 function-cyclin D1 binds the upstream regulatory region of the Notch1 gene, where it serves to recruit CREB binding protein (CBP) histone acetyltransferase. Genetic ablation of cyclin D1 resulted in decreased CBP recruitment, decreased histone acetylation of the Notch1 promoter region, and led to decreased levels of the Notch1 transcript and protein in cyclin D1-null (Ccnd1(-/-)) retinas. Transduction of an activated allele of Notch1 into Ccnd1(-/-) retinas increased proliferation of retinal progenitor cells, indicating that upregulation of Notch1 signalling alleviates the phenotype of cyclin D1-deficiency. These studies show that in addition to its well-established cell cycle roles, cyclin D1 has an in vivo transcriptional function in mouse development. Our approach, which we term 'genetic-proteomic', can be used to study the in vivo function of essentially any protein.
Bienvenu
Frédéric
F
Department of Cancer Biology, Dana-Farber Cancer Institute, Boston, Massachusetts 02115, USA.
Jirawatnotai
Siwanon
S
Elias
Joshua E
JE
Meyer
Clifford A
CA
Mizeracka
Karolina
K
Marson
Alexander
A
Frampton
Garrett M
GM
Cole
Megan F
MF
Odom
Duncan T
DT
Odajima
Junko
J
Geng
Yan
Y
Zagozdzon
Agnieszka
A
Jecrois
Marie
M
Young
Richard A
RA
Liu
X Shirley
XS
Cepko
Constance L
CL
Gygi
Steven P
SP
Sicinski
Piotr
P
D000483:Alleles; D000818:Animals; D050882:CREB-Binding Protein / Q000378:metabolism; D047369:Chromatin Immunoprecipitation; D019938:Cyclin D1 / Q000172:deficiency / Q000235:genetics / Q000378:metabolism*; D018507:Gene Expression Regulation, Developmental; D016678:Genome / Q000235:genetics; D057166:High-Throughput Screening Assays; D051548:Histone Acetyltransferases / Q000378:metabolism; D013058:Mass Spectrometry; D051379:Mice; D020411:Oligonucleotide Array Sequence Analysis; D011401:Promoter Regions, Genetic / Q000235:genetics; D011485:Protein Binding; D040901:Proteomics / Q000379:methods; D051381:Rats; D051881:Receptor, Notch1 / Q000235:genetics / Q000378:metabolism; D012160:Retina / Q000166:cytology / Q000196:embryology / Q000378:metabolism; D013234:Stem Cells / Q000166:cytology / Q000378:metabolism; D014158:Transcription, Genetic
C530105:Ccnd1 protein, mouse; C497762:Notch1 protein, mouse; D051881:Receptor, Notch1; D019938:Cyclin D1; D050882:CREB-Binding Protein; D051548:Histone Acetyltransferases