No abstract

Gadd45a, a p53-and BRCA1-regulated stress protein, has been implicated in the maintenance of genomic fidelity, probably through its roles in the control of cell cycle checkpoint and apoptosis. However, the mechanism(s) by which Gadd45a is involved in the induction of apoptosis remains unclear. We show here that inducible expression of Gadd45a protein causes dissociation of Bim, a Bcl2 family member, from microtubuleassociated components and translocation to mitochondria. The Bim accumulation in mitochondria enhances interaction of Bim with Bcl-2, relieves Bax from Bcl-2-bound complexes, and subsequently results in release of cytochrome c into the cytoplasm. Suppression of endogenous Bim greatly inhibits Gadd45a induction of apoptosis. Interestingly, Gadd45a interacts with elongation factor 1␣ (EF-1␣), a microtubule-severing protein that plays an important role in maintaining cytoskeletal stability, and inhibits EF-1␣-mediated microtubule bundling, indicating that the interaction of Gadd45a with EF-1␣ disrupts cytoskeletal stability. A mutant form of Gadd45a harboring a deletion of EF-1␣-binding domain fails to inhibit microtubule stability and to induce Bim translocation to mitochondria. Furthermore, coexpression of EF-1␣ antagonizes Gadd45a's property of suppressing cell growth and inducing apoptosis. These findings identify a novel link that connects stress protein Gadd45a to the apoptotic machinery and address the importance of cytoskeletal stability in apoptotic response to DNA damage.

The S100 gene family is closely associated with ESCC.

Gadd45a, a growth arrest and DNA-damage gene, plays important roles in the control of cell cycle checkpoints, DNA repair and apoptosis. We show here that Gadd45a is involved in the control of cell contact inhibition and cellcell adhesion. Gadd45a can serve as an adapter to enhance the interaction between b-catenin and Caveolin-1, and in turn induces b-catenin translocation to cell membrane for maintaining cell-cell adhesion/contact inhibition. This is coupled with reduction of b-catenin in cytoplasm and nucleus following Gadd45a induction, which is reflected by the downregulation of cyclin D1, one of the b-catenin targeted genes. Additionally, Gadd45a facilitates ultraviolet radiation-induced degradation of cytoplasmic and nuclear b-catenin in a p53-dependent manner via activation of p38 kinase. These findings define a novel link that connects Gadd45a to cell-cell adhesion and cell contact inhibition, which might contribute to the role of Gadd45a in inhibiting tumorigenesis.

The prevalence of obesity among child-bearing women has increased significantly. The adverse consequences of maternal obesity on the descendants have been well accepted, although few studies have examined the underlying mechanisms. We investigated whether neonatal overfeeding in female mice alters metabolic phenotypes in the offspring and whether hypothalamic leptin signalling is involved. Neonatal overfeeding was induced by reducing the litter size to three pups per litter, in contrast to normal litter size of 10 pups per litter. Normal and neonatally overfed female mice were bred with normal male mice, and offspring of overfeeding mothers (OOM) and control mothers (OCM) were generated. We examined body weight, daily food intake, leptin responsiveness and the number of positive neurones for phosphorylated-signal transducer and activator of transcription 3 (pSTAT3) along with neuropeptide Y (NPY) in the arcuate nucleus of the hypothalamus (ARH) and NPY in the nucleus tractus solitarius (NTS) of the brain stem. The body weight and daily food intake of OOM were significantly higher than those of OCM. Leptin significantly reduced food intake and increased the number of pSTAT3 positive neurones in the ARH of OCM mice, whereas no significant changes in food intake and pSTAT3 neurones were found in leptin-treated OOM mice. The number of NPY neurones in the ARH and NTS of the OOM mice was significantly higher than that of OCM mice. The results of the present study indicate that the obese phenotype from mothers can be passed onto the subsequent generation, which is possibly associated with hypothalamic leptin resistance.

BRCA1 is a well established tumor suppressor gene, which is involved in many cellular processes, including DNA damage repair, cell cycle control, apoptosis, as well as transcriptional control. In this work, we have found that BRCA1 is involved in regulating TGF‐β1/Smad pathway. The loss of endogenous BRCA1 greatly attenuated TGF‐β1‐induced growth inhibition and cell cycle G1 arrest. BRCA1 greatly maintains stability of Smad4 protein, and the loss of BRCA1 results in Smad4 down‐regulation, which is likely related to its downstream gene Gadd45a. Gadd45a is able to interact with β‐Trcp1, a‐F‐box protein of SCF E3 ligase, and consequently suppresses the ubiquitin‐degradation of Smad4 by SCFβ‐trcp1, as reflected by the observations that the induction of Gadd45a substantially stabilizes Smad4 protein. In addition, exogenous expression of Gadd45a can largely rescue the protein level of Smad4 in BRCA1 deficient cells. These results further demonstrate that BRCA1 may act as an important negative regulator in cell cycle progression and tumorigenesis through regulating the stability of Smad4, and define a novel link that connects BRCA1 to TGF‐β1/Smad pathway.

p53 mutations have been found in many esophageal squamous cell carcinoma (ESCC) clinical specimens and cell lines. We reasoned that functional inactivation of wild-type p53 or the functional activation of mutant-type p53 might exist in these specimens and cell lines. In this study, we identified the correlation between p53 functional activation and its genotype in five different ESCC cell lines. To examine the potential p53 activation in a certain ESCC cell line, DNA damage methods including X-ray exposure and cisplatin treatment were employed to treat cells. Further, the expression of p53 protein and four transcripts of well-known p53 target genes were investigated using Western blot and reverse transcriptionpolymerase chain reaction (RT-PCR) after cell exposure to DNA damage. The results showed that in KYSE 30 cell line with mutant p53 and KYSE 150 with wild-type p53, p53 could be activated by DNA damages. However, p53 could not be activated following the DNA damages in YES 2 with wild-type p53, KYSE 70 with mutant p53, and EC9706 with unknown p53 genotype. All our data indicated that p53 function in certain cells is not closely correlated with its genotype. To judge p53 function in a particular cell line, it is important to examine the p53 functional activation, but not to simply rely on the p53 genotype.

ABSTRACT. Liver receptor homologue 1 (Lrh-1) is a member of the nuclear receptor belonging to the second subfamily of the nuclear receptor family 5A (NR5A), also named NR5A2, which is important for lipid homeostasis, embryogenesis, and regulation of aromatics. The present study aimed to understand the sequence of ovine Lrh-1 and the expression traits in reproductive organ tissues. Initially, we cloned Lrh-1 from the liver of Hu sheep through degenerate primer of reverse transcriptionpolymerase chain reaction and rapid amplification of cDNA ends. Characteristic functional domains of DNA binding and ligand binding, conserved among transcription factors of the nuclear receptor superfamily, were identified in Lrh-1 of Hu sheep. The Lrh-1 protein levels in the tissues detected by Western blotting correlated significantly with the transcript levels measured by quantitative real-time polymerase chain reaction (qRT-PCR). To understand the Lrh-1 expression change in the hypothalamus and hypophysis during the estrous cycle, we analyzed the expression pattern of Lrh-1 mRNA and protein by qRT-PCR and Western blotting, respectively. This analysis revealed that Lrh-1 expression in the hypothalamus was highest during the metestrus phase, while the Lrh-1 level was similar during other phases. In the hypophysis, the expression was significantly different during the 4 phases of the estrous cycle but highest during the estrus phase, Characterization of Lrh-1 in ovine significantly correlating with FSH concentration. These results indicate that Lrh-1 expression is correlated with gonadotropic hormone secretion, influencing follicular formation in the ovary.