Somatic mutations in the p53 tumor suppressor gene in rheumatoid arthritis synovium

Abstract

The factors that regulate the perpetuation and invasiveness of rheumatoid synovitis have been the subject of considerable inquiry, and the possibility that nonimmunologic defects can contribute to the disease has not been rigorously addressed. Using a mismatch detection system, we report that synovial tissue from the joints of severe chronic rheumatoid arthritis patients contain mutant p53 transcripts, which were not found in skin samples from the same patients or in joints of patients with osteoarthritis. Mutant p53 transcripts also were identified in synoviocytes cultured from rheumatoid joints. The predicted amino acid substitutions in p53 were identical or similar to those commonly observed in a variety of tumors and might influence growth and survival of rheumatoid synoviocytes. Thus, mutations in p53 and subsequent selection of the mutant cells may occur in the joints of patients as a consequence of inflammation and contribute to the pathogenesis of the disease.

Figure 1

RMDA for p53 from RA and OA tissue samples. RNA was prepared from skin or synovium (SYN) obtained from RA or OA patients, and RMDA was performed. A p53 mutant cDNA (Ambion) served as a control for positive mismatch, which appears as smaller fragments after RNase digestion. p53 from normal blood was used as wild-type template, and wild type hybridized to wild type served as a no-mismatch control. (Upper) RMDAs resulting from samples transcribed in the sense direction hybridized to wild-type antisense transcripts. RA467 SYN and RA460 SYN lanes demonstrated intense complex banding patterns, suggesting the presence of multiple mutations. RA467 SKIN, OA440 SYN, and OA458 SYN showed only very faint nonspecific bands. (Lower) The converse hybridizations with wild-type sense and sample antisense transcripts, with similar results.

Figure 2

p53 protein detection by immunoprecipitation. p53 protein expression was analyzed in synovial tissue (RA and OA), skin (OA), fibroblast-like synoviocytes [RA, OA, and normal (NML)], and dermal fibroblasts (DF). Different samples are from different patients and are numbered arbitrarily. Immunoprecipitation was performed as described in Methods using either PAb240 (detects mutant but not wild-type p53) or DO1 (detects mutant and wild-type p53). Note the discordant expression of DO1 and PAb240 precipitable protein in OA joint samples, whereas abundant p53 was detected in RA joint tissue using PAb240.