1. Partial obstruction of the murine ileum led to changes in the gross morphology and ultrastructure of the tunica muscularis. Populations of interstitial cells of Cajal (ICC) decreased oral, but not aboral, to the site of obstruction. Since ICC generate and propagate electrical slow waves in gastrointestinal muscles, we investigated whether the loss of ICC leads to loss of function in partial bowel obstruction.2. Changes in ICC networks and electrical activity were monitored in the obstructed murine intestine using immunohistochemistry, electron microscopy and intracellular electrophysiological techniques.3. Two weeks following the onset of a partial obstruction, the bowel increased in diameter and hypertrophy of the tunica muscularis was observed oral to the obstruction site. ICC networks were disrupted oral to the obstruction, and this disruption was accompanied by the loss of electrical slow waves and responses to enteric nerve stimulation. These defects were not observed aboral to the obstruction. 6. These data describe the plasticity of ICC networks in response to partial obstruction. After obstruction the ICC phenotype was lost, but these cells regenerated when the obstruction was removed. This model may be an important tool for evaluating the cellular/molecular factors responsible for the regulation and maintenance of the ICC phenotype.

No abstract

The p53-inducible gene 3 (PIG3) is originally isolated as a p53 downstream target gene, but its function remains unknown. Here, we report a role of PIG3 in the activation of DNA damage checkpoints, after UV irradiation or radiomimetic drug neocarzinostatin (NCS). We show that depletion of endogenous PIG3 sensitizes cells to DNA damage agents, and impaired DNA repair. PIG3 depletion also allows for UV-and NCS-resistant DNA synthesis and permits cells to progress into mitosis, indicating that PIG3 knockdown can suppress intra-S phase and G2/M checkpoints. PIG3-depleted cells show reduced Chk1 and Chk2 phosphorylation after DNA damage, which may directly contribute to checkpoint bypass. PIG3 exhibited diffuse nuclear staining in the majority of untreated cells and forms discrete nuclear foci in response to DNA damage. PIG3 colocalizes with c-H2AX and 53BP1 to sites of DNA damage after DNA damage, and binds to a c-H2AX. Notably, PIG3 depletion decreases the efficient induction and maintenance of H2AX phosphorylation after DNA damage. Moreover, PIG3 contributes to the recruitment of 53BP1, Mre11, Rad50 and Nbs1 to the sites of DNA break lesions in response to DNA damage. Our combined results suggest that PIG3 is a critical component of the DNA damage response pathway and has a direct role in the transmission of the DNA damage signal from damaged DNA to the intra-S and G2/M checkpoint machinery in human cells.

Tumors frequently contain mutations in the ras genes, resulting in the constitutive activation of the Ras-activated signaling pathway. The activation of Ras is involved not only in tumor progression but also in the development of resistance of the tumor cells to platinum-based chemotherapeutic agents. To investigate the potential mechanisms underlying this resistance, we analyzed the effect of activated H-Ras on the expression of the nucleotide excision repair genes. Here we identified ERCC1, which is one of the key enzymes involved in nucleotide excision repair, as being markedly up-regulated by the activated H-Ras. From promoter analysis of ERCC1, an increase in the Ap1 transcriptional activity as a result of the expression of the oncogenic H-Ras was found to be crucial for this induction. In addition, ERCC1 small interfering RNA expression was shown to reduce the oncogenic H-Ras-mediated increase in the DNA repair activity as well as to suppress the oncogenic H-Ras-mediated resistance of the cells to platinum-containing chemotherapeutic agents. These results suggest that the oncogenic H-Ras-induced ERCC1, which activates the DNA repair capacity, may be involved in the protection of the cells against platinum-based anticancer agents.

Bcl-2 stimulates mutagenesis after the exposure of cells to DNA-damaging agents. However, the biological mechanisms of Bcl-2-mediated mutagenesis have remained largely obscure. Here we demonstrate that the Bcl-2-mediated suppression of hMSH2 expression results in a reduced cellular capacity to repair mismatches. The pathway linking Bcl-2 expression to the suppression of mismatch repair (MMR) activity involves the hypophosphorylation of pRb, and then the enhancement of the E2F-pRb complex. This is followed by a decrease in hMSH2 expression. MMR has a key role in protection against deleterious mutation accumulation and in maintaining genomic stability. Therefore, the decreased MMR activity by Bcl-2 may be an underlying mechanism for Bcl-2-promoted oncogenesis.