<i>In vitro</i> genotoxicity analyses of colibactin-producing <i>E. coli</i> isolated from a Japanese colorectal cancer patient

Abstract

Colibactin is a polyketide-peptide genotoxin produced by enteric bacteria such as E. coli, and is considered to contribute to the development of colorectal cancer. We previously isolated E. coli strains from Japanese colorectal cancer patients, and in the present study we investigated the genotoxic potency of the colibactin-producing (clb +) E. coli strains that carry the polyketide synthases "pks" gene cluster (pks +) and an isogenic clbmutant in which the colibactin-producing ability is impaired. Measurement of phosphorylated histone H2AX indicated that DNA double strand breaks were induced in mammalian CHO AA8 cells infected with the clb + E. coli strains. Induction of DNA damage response (SOS response) by crude extract of the clb + strains was 1.7 times higher than that of the clb-E. coli in an umu assay with a Salmonella typhimurium TA1535/pSK1002 tester strain. Micronucleus test with CHO AA8 cells revealed that infection with the clb + strains induced genotoxicity, i.e., the frequencies of micronucleated cells infected with clb + strain were 4-6 times higher than with the clbstrain. Since the intestinal flora are affected by dietary habits that are strongly associated with ethnicity, these data may contribute to both risk evaluation and prevention of colorectal cancer in the Japanese population.

Association of Escherichia coli containing polyketide synthase in the gut microbiota with colorectal neoplasia in Japan

“…Escherichia coli from the B2 phylogenetic group possesses a genomic island named polyketide synthetase ( pks ), which is thought to produce a polyketide‐peptide genotoxin, colibactin. E coli containing pks ( pks + E coli ) has been shown to induce DNA double‐strand breaks, cell cycle arrest, mutations, and chromosomal instability in eukaryotic cells 6‐9 . Colibactin also alkylates DNA in vivo and DNA adducts have been identified in mammalian cells and mice exposed to pks + E coli 10 .…”

Colon Carcinogenesis: The Interplay Between Diet and Gut Microbiota

“…The pathogenicity island encoding colibactin, known as polyketide synthase (pks), are detected in up to 66.7% of CRC patients (Arthur et al, 2012). Moreover, experimental studies show that colibactin could sustain tumor growth by generating DNA cross links in cellulo, which were later transformed into DNA doublestrand breaks (Bossuet-Greif et al, 2018;Kawanishi et al, 2019;Wilson et al, 2019), or by favoring the emergence of senescent cells (Cougnoux et al, 2014;Dalmasso et al, 2014). Senescent cells are cells with an irreversible cell cycle arrest.…”

Genotyping of a gene cluster for production of colibactin and in vitro genotoxicity analysis of Escherichia coli strains obtained from the Japan Collection of Microorganisms

“…U.S.A). The DNA damaging potency of bacterial cell extracts was estimated using the umu test, as previously described [5]. Briefly, E. coli cells were harvested from 10 mL of overnight culture in LB media (O.D.…”

“…The clb + E. coli stimulates growth of colon tumors under conditions of host inflammation, and is found with increased frequency in inflammatory bowel disease, familial adenomatous polyposis, and colorectal cancer patients [2,3]. We previously reported that E. coli strains isolated from a Japanese colorectal cancer patient produced colibactin and showed genotoxicity in in vitro assays [4,5]. However, the chemical structure of the genotoxin, the molecular mechanism of its mutagenesis/carcinogenesis, and distribution of the clb gene cluster among microorganisms have not been fully clarified yet.…”