1: Eur J Neurosci. 2005 Jun;21(11):2903-11. A bipotent neural progenitor cell line cloned from a cerebellum of an adult p53-deficient mouse generates both neurons and oligodendrocytes. Tominaga M, Honda S, Okada A, Ikeda A, Kinoshita S, Tomooka Y. Department of Biological Science and Technology and Tissue Engineering Research Center, Tokyo University of Science, 2641 Yamazaki, Noda, Chiba 278-8510, Japan. Here we report developmental characteristics of a clonal cell line 2Y-3t established from a multifocal neoplasm that arose in a cerebellum of an adult p53-deficient mouse. The tumorigenicity of the line was not observed in soft agar assay or in nude mouse assay. In serum-containing medium, 2Y-3t cells were epithelial-like in morphology and were mitotic. When they were cultured in serum-free medium, the expressions of neural stem and/or progenitor cell markers were decreased. Concomitantly, the expressions of neuronal and oligodendrocyte markers were increased in concert with morphological differentiation, and DNA synthesis ceased. None of astrocyte markers were detected under these culture conditions. Double-labelling studies revealed that two cell populations coexisted, expressing neuronal or oligodendrocyte markers. Triiodothyronine (T3) increased the oligodendrocyte population when 2Y-3t cells were cultured in serum-free medium. Recloning of the line gave rise to three types of subclones. Sixteen subclones were capable of generating both neurons and oligodendrocytes, four subclones were capable of generating only neurons and one subclone was capable of generating only oligodendrocytes. Thus, 2Y-3t cells have characteristics of bipotent neural progenitor cells capable of generating both neurons and oligodendrocytes. In addition, the line expressed mRNA for Pax-2 and had GAD67-positive cells when cultured in serum-free medium. However, none of the mRNAs for Zic-1, Math1, zebrin or Calbindin-D28k were detected, suggesting that the 2Y-3t line might generate the GABAergic interneuron lineage of the mouse cerebellum. PMID: 15978002 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 2: J Biol Chem. 2004 Dec 31;279(53):55562-9. Epub 2004 Sep 27. Human PTIP facilitates ATM-mediated activation of p53 and promotes cellular resistance to ionizing radiation. Jowsey PA, Doherty AJ, Rouse J. Medical Research Council Protein Phosphorylation Unit, Wellcome Trust Biocentre/Medical Sciences Institute Complex, Dow Street, University of Dundee, Dundee DD1 5EH, Scotland, United Kingdom. Mus musculus Pax2 transactivation domain-interacting protein (Ptip) is an essential gene required for the maintenance of genome stability, although its precise molecular role is unclear. Human PTIP (hPTIP) was recently isolated in a screen for proteins, translated from cDNA pools, capable of interacting with peptides phosphorylated by the ATM (ataxia telangiectasia-mutated)/ATR (ataxia telangiectasia-related) protein kinases. hPTIP was described as a 757-amino acid protein bearing four BRCT domains. Here we report that instead full-length endogenous hPTIP contains 1069 amino acids and six BRCT domains. hPTIP shows increased association with 53BP1 in response to ionizing radiation (IR) but not in response to other DNA-damaging agents. Whereas translocation of both 53BP1 and hPTIP to sites of IR-induced DNA damage occurs independently of ATM, IR-induced association of PTIP and 53BP1 requires ATM. Deletion analysis identified the domains of 53BP1 and hPTIP required for protein-protein interaction and focus formation. Data characterizing the cellular roles of hPTIP are also presented. Small interfering RNA was used to show that hPTIP is required for ATM-mediated phosphorylation of p53 at Ser(15) and for IR-induced up-regulation of the cyclin-dependent kinase inhibitor p21. Lowering hPTIP levels also increased cellular sensitivity to IR, suggesting that this protein plays a critical role in maintaining genome stability. PMID: 15456759 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 3: Oncogene. 2003 Sep 11;22(39):7989-97. Paired-Box genes are frequently expressed in cancer and often required for cancer cell survival. Muratovska A, Zhou C, He S, Goodyer P, Eccles MR. Department of Pathology, Dunedin School of Medicine, University of Otago, PO Box 913, Dunedin, New Zealand. The paired-box (PAX) genes encode a family of nine well-characterized paired-box transcription factors, with important roles in development and disease. Although PAX genes are primarily expressed in the embryo, constitutive expression promotes tissue hyperplasia. Rare tumor-specific mutations of PAX genes implicate an oncogenic role, and persistent PAX expression characterizes several tumors. Yet, a cancer-wide analysis of PAX gene expression to investigate a general role for PAX genes has not been performed. We analysed the pattern and requirement for PAX gene expression in a panel of common cancer cell lines. Very frequent PAX gene expression was identified in tumor cell lines, including lymphoma, breast, ovarian, lung, and colon cancer. In addition, the PAX2 gene was frequently expressed in a panel of 406 common primary tumor tissues. Apoptosis was rapidly induced in ovarian and bladder cancer cell lines following RNA interference to silence PAX2 expression, despite concomitant TP53 and/or HRAS mutations. These data suggest that PAX genes are frequently expressed in cancer, and that endogenous PAX gene expression is required for the growth and survival of cancer cells. PMID: 12970747 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 4: Oncogene. 1997 Jun 5;14(22):2689-700. Differential regulation of the human Wilms tumour suppressor gene (WT1) promoter by two isoforms of PAX2. McConnell MJ, Cunliffe HE, Chua LJ, Ward TA, Eccles MR. Department of Biochemistry, University of Otago, Dunedin, New Zealand. PAX2 is a member of the paired box family of genes with an important role in kidney, genital tract and eye development. Another gene essential for kidney and genital tract development is the Wilms tumour gene, WT1. PAX2 and WT1 encode transcription factors expressed during fetal kidney development in patterns that overlap both spatially and temporally. The overlap of PAX2 and WT1 expression in fetal kidney prompted us to determine whether PAX2 regulates the WT1 gene. To investigate this possibility, the WT1 promoter and a series of WT1 promoter deletion fragments were cloned into a luciferase reporter vector, and used in co-transfection experiments with PAX2 expression constructs. PAX2 transactivated the WT1 promoter up to 35-fold in CHO-K1 cells, and from four- to sevenfold in 293 cells. Two regions of the WT1 promoter were required in the same promoter construct for efficient transactivation by PAX2 in CHO-K1 cells, and purified recombinant PAX2 protein was found to bind to two sites in the WT1 promoter, at -205/-230 and +377/+402. Removal of WT1 promoter sequences containing the -205/-230, or +377/+402 binding sites abolished transactivation of the WT1 promoter by PAX2 in CHO-K1 cells, and had a differential effect on transactivation of the WT1 promoter in 293 cells, depending on the PAX2 isoform used. A fragment containing the -205/-230 site alone could be transactivated by PAX2. These findings suggest that PAX2 is a tissue-specific modulator of WT1 expression, and is involved in cell growth control via WT1. PMID: 9178767 [PubMed - indexed for MEDLINE] --------------------------------------------------------------- 5: EMBO J. 1995 Nov 15;14(22):5638-45. Loss of p53 function through PAX-mediated transcriptional repression. Stuart ET, Haffner R, Oren M, Gruss P. Department of Molecular Cell Biology, Max-Planck Institute for Biophysical Chemistry, Gottingen, Germany. Direct interactions between the genes that regulate development and those which regulate the cell cycle would provide a mechanism by which numerous biological events could be better understood. We have identified a direct role for PAX5 in the control of p53 transcription. In primary human diffuse astrocytomas, PAX5 expression inversely correlated with p53 expression. The human p53 gene harbours a PAX binding site within its untranslated first exon that is conserved throughout evolution. PAX5 and its paralogues PAX2 and PAX8 are capable of inhibiting both the p53 promoter and transactivation of a p53-responsive reporter in cell culture. Mutation of the identified binding site eliminates PAX protein binding in vitro and renders the promoter inactive in cells. These data suggest that PAX proteins might regulate p53 expression during development and propose a novel alternative mechanism for tumour initiation or progression, by which loss of p53 function occurs at the transcriptional level. PMID: 8521821 [PubMed - indexed for MEDLINE] ---------------------------------------------------------------