RESUMEN
Here we compared the proteomes of primary fibroblast cultures derived from morphologically normal colonic mucosa of familial adenomatous polyposis (FAP) patients with those obtained from unaffected controls. The expression signature of about 19% of total fibroblast proteins separates FAP mutation carriers from unaffected controls (P < 0.01). More than 4,000 protein spots were quantified by 2D PAGE analysis, identifying 368 non-redundant proteins and 400 of their isoforms. Specifically, all three classes of cytoskeletal filaments and their regulatory proteins were altered as were oxidative stress response proteins. Given that FAP fibroblasts showed heightened sensitivity to transformation by KiMSV and SV40 including elevated levels of the p53 protein, events controlled in large measure by the Ras suppressor protein-1 (RSU-1) and oncogenic DJ-1, here we show decreased RSU1 and augmented DJ-1 expression in both fibroblasts and crypt-derived epithelial cells from morphologically normal colonic mucosa of FAP gene-carriers. The results indicate that heterozygosity for a mutant APC tumor suppressor gene alters the proteomes of both colon-derived normal fibroblasts in a gene-specific manner, consistent with a "one-hit" effect.
Asunto(s)
Poliposis Adenomatosa del Colon/genética , Poliposis Adenomatosa del Colon/metabolismo , Genes APC , Proteínas de Neoplasias/biosíntesis , Proteoma/biosíntesis , Poliposis Adenomatosa del Colon/patología , Adulto , Anciano , Estudios de Casos y Controles , Fibroblastos/metabolismo , Fibroblastos/patología , Regulación Neoplásica de la Expresión Génica , Heterocigoto , Humanos , Péptidos y Proteínas de Señalización Intracelular/genética , Persona de Mediana Edad , Proteínas de Neoplasias/genética , Proteínas Oncogénicas/biosíntesis , Proteínas Oncogénicas/genética , Estrés Oxidativo/genética , Proteína Desglicasa DJ-1 , Proteoma/genética , Factores de Transcripción/biosíntesis , Factores de Transcripción/genética , Adulto JovenRESUMEN
We seek alterations in protein patterns at the earliest possible step on the path to cancer, namely, in cells of the target tissue from normal persons versus the corresponding normally appearing cells from persons who are heterozygous for mutation in a tumor suppressor gene that predisposes strongly to carcinoma in that tissue. To begin a systematic comparison of the proteomes of cells from normal and from neoplastic colons, we have undertaken the isolation of human colon crypts that are derived from the normal-appearing mucosa of left (descending) colon of patients with sporadic colorectal cancer. Two-dimensional (2D) gel electrophoresis is a proteomic approach that excels in the resolution of protein isoforms. Here, we document the practicality of this approach with human samples using gels of three overlapping pH ranges. For the first time, about 800 nonredundant proteins and 900 isoforms from purified human colonic crypts were identified, permitting an assessment of the contributions of protein isoforms. These interactive, searchable, hyperlink-enabled proteome maps and gene ontology analyses will facilitate future studies to discover the earliest markers and intervention targets during progression to colon cancer.