RESUMO
DPC4/SMAD4 is a candidate tumor suppressor gene with a strikingly high frequency of gene alterations in pancreatic cancer that suggests a discrete role for DPC4 in these tumors. DPC4 tumor-suppressive function has been implicated to mediate the transforming growth factor-beta (TGFbeta)-suppressive pathway; however, in a DPC4-null pancreatic cancer cell line, TGFbeta growth-inhibitory and transcriptional responses were found to be DPC4-independent. This was observed within native cells having a natural homozygous deletion and in clones engineered for stable expression of wild-type DPC4 integrated into the genome. This observation contrasted with the absolute DPC4 dependence of TGFbeta responses in a breast cancer cell line studied in parallel. This growth-inhibitory response to TGFbeta in DPC4-null cells relied on an intact ras effector pathway. These data further suggest a major categorization of TGFbeta responses into DPC4-dependent and -independent signaling pathways and specifically suggest that disruption of the TGFbeta-independent signal might be a basis of selection for the emergence of DPC4 alterations during tumorigenesis in the pancreas and other sites.
Assuntos
Neoplasias da Mama/genética , Proteínas de Ligação a DNA/genética , Genes Supressores de Tumor , Neoplasias Pancreáticas/genética , Transativadores/genética , Fator de Crescimento Transformador beta/fisiologia , Alelos , Proteínas Quinases Dependentes de Cálcio-Calmodulina/antagonistas & inibidores , Proteínas Quinases Dependentes de Cálcio-Calmodulina/fisiologia , Proteínas de Ligação a DNA/biossíntese , Proteínas de Ligação a DNA/metabolismo , Deleção de Genes , Humanos , Transdução de Sinais/fisiologia , Proteína Smad4 , Transativadores/biossíntese , Transativadores/deficiência , Transativadores/metabolismo , Transfecção , Fator de Crescimento Transformador beta/genética , Células Tumorais Cultivadas , Proteínas ras/antagonistas & inibidores , Proteínas ras/fisiologiaRESUMO
The identification of homozygous deletions in sporadic neoplasms has been pivotal in the positional cloning of several tumor suppressor genes. Chromosomal arm 18q harbors the DPC4, SMAD2, and DCC genes and is suspected on the basis of high frequencies of allelic loss to harbor additional tumor suppressor genes. We applied high-resolution sequence-tagged site (STS) marker scanning to a panel of 106 pancreatic adenocarcinomas to identify novel regions of homozygous deletions on 18q. Three homozygous deletions were identified. Physical mapping of these deletions showed them to be nonoverlapping, but clustered in an approximately 7- to 10-Mb region of chromosome band 18q22. Each deletion spanned physical distances of nearly 1.3 to 3 Mb. A number of transcribed genes map within these deletions. The identification of these homozygous deletions might aid in the identification of novel tumor suppressor genes on chromosomal arm 18q. Genes Chromosomes Cancer 25:370-375, 1999.
Assuntos
Adenocarcinoma/genética , Deleção Cromossômica , Cromossomos Humanos Par 18/genética , Perda de Heterozigosidade/genética , Neoplasias Pancreáticas/genética , Sitios de Sequências Rotuladas , Bandeamento Cromossômico , Marcadores Genéticos , Humanos , Mapeamento Físico do Cromossomo/métodos , Células Tumorais CultivadasRESUMO
Chromosomal arm 1p has long been suspected, on the basis of loss of heterozygosity (LOH) and other data, to harbor a tumor suppressor gene important in pancreatic carcinomas and other tumors. We constructed a high-resolution map of LOH at I p in a panel of pancreatic adenocarcinomas. Using 44 markers, we identified LOH on I p in 49% of 43 cancers. Breakpoints in 1p were identified in 15 of the carcinomas and could be used to ascertain consensus patterns. We found a major consensus region of LOH at 1p35 between loci D1S233 and D1S247. This region participates in the majority of LOH events on 1 p in pancreatic cancer. These data provide a roadmap for further regional mapping, homozygous deletion searches, comparison to LOH patterns seen in other tumor types, and prioritization of studies using candidate genes.
Assuntos
Adenocarcinoma/genética , Mapeamento Cromossômico/métodos , Cromossomos Humanos Par 1/genética , Neoplasias Pancreáticas/genética , Animais , Sequência Consenso , Humanos , Perda de Heterozigosidade/genética , Camundongos , Camundongos Nus , Transplante HeterólogoRESUMO
Dpc4 (Smad4) is implicated in mediation of signals from transforming growth factor (TGF) beta and related ligands, and wild-type Dpc4 mediates TGF-beta-stimulated gene transcription at specific DNA sequences bound by Dpc4 [Smad binding element (SBE)]. We characterized panels of DPC4 tumor mutations and cancer cell lines. Amino acid substitutions within the NH2-terminal third of Dpc4 weakened or ablated SBE-mediated gene regulation by a disruption of DNA binding. An interaction of the COOH-terminal end with the DNA-binding domain of Dpc4 was evident but was not required to explain the functional impairment produced by NH2-terminal DPC4 mutations. Both substitution and truncation mutations of the COOH-terminal half of DPC4 lacked the ability to regulate transcription while retaining the sequence-specific DNA-binding function, but through differing mechanisms. A modular domain to redistribute Dpc4 to the nuclear compartment allowed SBE-mediated transcriptional activation in a cell line having a TGF-1 receptor defect and was sufficient to restore SBE-mediated transactivation ability to COOH-terminal DPC4 missense mutants. Cells harboring DPC4 alterations had a universal impairment of the TGF-beta-stimulated SBE transcriptional response. These studies identify the loss of SBE-mediated gene regulation as a uniform outcome of the selection for DPC4 alterations during tumorigenesis. They raise the possibility of restoration of some Dpc4-associated transcriptional events in cancer cells through the targeted redistribution of wild-type and some missense mutant forms of Dpc4 to the nucleus.
