ZBTB2, a Novel Master Regulator of the p53 Pathway

Abstract

We found that ZBTB2, a POK family transcription factor, is a potent repressor of the ARF-HDM2-p53-p21 pathway important in cell cycle regulation. ZBTB2 repressed transcription of the ARF, p53, and p21 genes, but activated the HDM2 gene. In particular, ZBTB2 repressed transcription of the p21 gene by acting on the two distal p53 binding elements and the proximal Sp1 binding GC-box 5/6 elements. ZBTB2 directly interacted with Sp1 via its POZ domain and zinc fingers, which was important in the repression of transcription activation by Sp1. ZBTB2 and Sp1 competed with each other in binding to the GC-box 5/6 elements and the two p53 binding elements. ZBTB2 directly interacted with p53 via its zinc fingers, inhibiting p53 binding and repressing transcription activation by p53. The POZ domain, required for transcription repression, interacted with corepressors such as BCoR, NCoR, and SMRT. The interactions deacetylated histones Ac-H3 and -H4 at the proximal promoter. Although ectopic ZBTB2 stimulated cell proliferation, knockdown of ZBTB2 expression decreased cell proliferation and DNA synthesis. Overall, our data suggest that ZBTB2 is a potential proto-oncogenic master control gene of the p53 pathway and, in particular, is a potent transcription repressor of the cell cycle arrest gene p21 by inhibiting p53 and Sp1.The POZ domain is an evolutionarily conserved proteinprotein interaction motif found in many cellular regulatory proteins (1, 2). POZ domain genes, first identified in Drosophila and poxvirus, have since been found in organisms ranging from yeast to humans (3, 4). As many as 184 known human proteins, 96 Drosophila proteins, and 137 Caenorhabditis elegans proteins are estimated to contain the POZ domain.POZ domain proteins are involved in many critical cellular processes such as apoptosis (5), development (6, 7), ion channel activity (4), oncogenesis (8 -10), and transcription (10 -16). In particular, some of the POZ domain Krüppel-like zinc finger (POK) 3 proteins are the major determinants of development, differentiation, and oncogenesis. PLZF-null mice display severe defects in limb development and germ stem cell maintenance (7, 17). T helper-inducing POZ/Krüppel-like factor (Th-POK/cKrox) has been recently reported as a master regulator of T-cell lineage commitment (18). BCL-6, PLZF, and HIC1 have been implicated in non-Hodgkin lymphoma, acute promyelocytic leukemia, and spontaneous malignant tumors, respectively (8,9,19). Recently, FBI-1 (also called Pokemon) has been shown to act as a proto-oncogene by repressing transcription of the ARF gene, causing down-regulation of p53 and promoting oncogenic cellular transformation (10).The most striking property of some POZ domain transcription factors is their ability to repress transcription via their POZ domains (10 -16, 20), although a few POZ domain transcription factors activate transcription (21,22). This characteristic probably underlies many biological processes controlled by these factors. The ability of the domain to interact with key regulatory protei...

POZ/BTB and AT-hook-containing zinc finger protein 1 (PATZ1) inhibits endothelial cell senescence through a p53 dependent pathway

“…27 ZBTB2 was found to increase cell proliferation by repressing transcription of the p21 CIP1 gene, a well known p53 target gene. 28 HIC1 suppresses age-dependent development of cancer by mediating SIRT1-and p53-dependent apoptotic DNA damage responses. 29 One critical question has been determining what signal mediates the cellular senescence induced by PATZ1 knockdown in young HUVECs.…”

Regulation of the Cyclin-dependent Kinase Inhibitor 1A Gene (CDKN1A) by the Repressor BOZF1 through Inhibition of p53 Acetylation and Transcription Factor Sp1 Binding

“…Previously, we have shown that FBI-1 (ZBTB7A), ZBTB2, ZBTB5, Kr-pok, and MIZ-1 also regulate the transcription of CDKN1A (16,(43)(44)(45)(46). FBI-1, ZBTB2, and ZBTB5 interact with Sp1 and compete with Sp1 for binding to the GC boxes of the proximal promoter.…”

“…Cells were fixed with 100% methanol and washed with PBS, and cellular DNA was stained with propidium iodide (50 g/ml) in RNase A (100 g/ml) solution (43). A cell cycle profile and forward scatter were determined using a BD FACSCalibur, and the data were analyzed using the CellQuest program (BD Biosciences).…”

“…The pGL2-CDKN1A-Luc plasmid series was reported elsewhere (43). pGL2-CDKN1A-Luc-mC/ EBP␣, -␤, and -mRAR were prepared by site-directed mutagenesis.…”

“…For the analysis of transcriptional regulation of various promoters by BOZF1, promoter-reporter fusion plasmids, pcDNA3.1-BOZF1, or the pcDNA3.1 vector in various combinations were transiently transfected into HEK293 and HCT116 p53 ϩ/ϩ cells using Lipofectamine Plus reagent (Invitrogen) and analyzed for luciferase activity as reported elsewhere (43).…”