TY - JOUR
T1 - Identification of target genes of the p16INK4A-pRB-E2F pathway.
AU - Vernell, Richard
AU - Helin, Kristian
AU - Müller, Heiko
N1 - Keywords: Amino Acid Motifs; Binding Sites; Blotting, Northern; Cell Cycle Proteins; Cell Division; Cell Line, Tumor; Cyclin-Dependent Kinase Inhibitor p16; DNA-Binding Proteins; Databases as Topic; Down-Regulation; E2F Transcription Factors; Exons; Gene Expression Regulation; Humans; Microscopy, Fluorescence; Mutation; Oligonucleotide Array Sequence Analysis; Phosphorylation; Promoter Regions (Genetics); Protein Synthesis Inhibitors; RNA; Retinoblastoma Protein; Reverse Transcriptase Polymerase Chain Reaction; Tetracycline; Transcription Factors; Transcription, Genetic; Up-Regulation
PY - 2003
Y1 - 2003
N2 - Deregulation of the retinoblastoma protein (pRB) pathway is a hallmark of human cancer. The core members of this pathway include the tumor suppressor protein, pRB, which through binding to a number of cellular proteins, most notably members of the E2F transcription factor family, regulates progression through the cell division cycle. With the aim of identifying transcriptional changes provoked by deregulation of the pRB pathway, we have used cell lines that conditionally express a constitutively active phosphorylation site mutant of pRB (pRBDeltaCDK) or p16INK4A (p16). The expression of pRBDeltaCDK and p16 resulted in significant repression and activation of a large number of genes as measured by high density oligonucleotide array analysis. Transcriptional changes were found in genes that are essential for DNA replication and cell proliferation. In agreement with previous results, we found a high degree of overlap between genes regulated by p16 and pRB. Data we have obtained previously for E2F family members showed that 74 of the genes repressed by pRB and p16 were induced by the E2Fs and 23 genes that were induced by pRB and p16 were repressed by the E2Fs. Thus, we have identified 97 genes as physiological targets of the pRB pathway, and the further characterization of these genes should provide insights into how this pathway controls proliferation. We show that Gibbs sampling detects enrichment of several sequence motifs, including E2F consensus binding sites, in the upstream regions of these genes and use this enrichment in an in silico filtering process to refine microarray derived gene lists.
AB - Deregulation of the retinoblastoma protein (pRB) pathway is a hallmark of human cancer. The core members of this pathway include the tumor suppressor protein, pRB, which through binding to a number of cellular proteins, most notably members of the E2F transcription factor family, regulates progression through the cell division cycle. With the aim of identifying transcriptional changes provoked by deregulation of the pRB pathway, we have used cell lines that conditionally express a constitutively active phosphorylation site mutant of pRB (pRBDeltaCDK) or p16INK4A (p16). The expression of pRBDeltaCDK and p16 resulted in significant repression and activation of a large number of genes as measured by high density oligonucleotide array analysis. Transcriptional changes were found in genes that are essential for DNA replication and cell proliferation. In agreement with previous results, we found a high degree of overlap between genes regulated by p16 and pRB. Data we have obtained previously for E2F family members showed that 74 of the genes repressed by pRB and p16 were induced by the E2Fs and 23 genes that were induced by pRB and p16 were repressed by the E2Fs. Thus, we have identified 97 genes as physiological targets of the pRB pathway, and the further characterization of these genes should provide insights into how this pathway controls proliferation. We show that Gibbs sampling detects enrichment of several sequence motifs, including E2F consensus binding sites, in the upstream regions of these genes and use this enrichment in an in silico filtering process to refine microarray derived gene lists.
U2 - 10.1074/jbc.M304930200
DO - 10.1074/jbc.M304930200
M3 - Journal article
C2 - 12923195
SN - 0021-9258
VL - 278
SP - 46124
EP - 46137
JO - Journal of Biological Chemistry
JF - Journal of Biological Chemistry
IS - 46
ER -
