Vertebrate embryo somite formation is temporally controlled by the cyclic expression of somitogenesis clock genes in the presomitic mesoderm (PSM). The somitogenesis clock is believed to be an intrinsic property of this tissue, operating independently of embryonic midline structures and the signaling molecules produced therein, namely Sonic hedgehog (Shh). This work revisits the notochord signaling contribution to temporal control of PSM segmentation by assessing the rate and number of somites formed and somitogenesis molecular clock gene expression oscillations upon notochord ablation. The absence of the notochord causes a delay in somite formation, accompanied by an increase in the period of molecular clock oscillations. Shh is the notochord-derived signal responsible for this effect, as these alterations are recapitulated by Shh signaling inhibitors and rescued by an external Shh supply. We have characterized chick smoothened expression pattern and have found that the PSM expresses both patched1 and smoothened Shh signal transducers. Upon notochord ablation, patched1, gli1, and fgf8 are down-regulated, whereas gli2 and gli3 are overexpressed. Strikingly, notochord-deprived PSM segmentation rate recovers over time, concomitant with raldh2 overexpression. Accordingly, exogenous RA supplement rescues notochord ablation effects on somite formation. A model is presented in which Shh and RA pathways converge to inhibit PSM Gli activity, ensuring timely somite formation. Altogether, our data provide evidence that a balance between different pathways ensures the robustness of timely somite formation and that notochord-derived Shh is a component of the molecular network regulating the pace of the somitogenesis clock.somitogenesis | molecular clock | notochord

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The reactions of two series of benzoyl derivatives of nucleophiles were investigated and the results were compared with those for N-benzoylimidazole. The general structure is ArCO-Nu, where Ar ¼ X-C 6 H 4 -and/or X 2 -C 6 H 3 -; X ¼ 4-Me, H, 4-Cl, 4-CN, 4-NO 2 and X 2 ¼ 3,5-dinitro; Nu ¼ iodosobenzoate, ArCO-Iba, and phosphate dianion, ArCO-Phos. Catalytic rate constants, kinetic solvent isotope effects, kinetic substituent effects (Hammett equation) and the dependence of ÁH 6 ¼ and ÁS 6 ¼ on X were determined. For ArCO-Iba, the hydrolysis occurs via two pathways, uncatalyzed (k H 2 O ) and specific base-catalyzed (k OH ) water attack on the iodine atom of the iodosobenzoate ring. This conclusion is based on theoretical calculations of the partial charges on ArCO-Iba, and the small values calculated at 25 C, À0.22 and 0.92 for k H 2 O and k OH , respectively. The data for the reaction of ArCO-Phos are consistent with a dissociative transition state, leading to elimination of the metaphosphate monoanion (PO 3 À ). The dependence of the mechanistic pathway on the nucleophile is discussed. Two results are relevant to the reactions of ArCO-Iba: (i) moderate to large substituent effects on the activation entropies suggest that solvation of the leaving benzoate anion and desolvation of the entering nucleophile contribute to the OH À -mediated reaction; (ii) the negative and positive signs of ÁS 6 ¼ indicate large differences in solvation of the transition states of the k H 2 O and k OH pathways. For the spontaneous decomposition of ArCO-Phos, sizeable substituent effects on both ÁH 6 ¼ and ÁS 6 ¼ were observed. This shows the contribution of solvation of the leaving benzoate and substituent-induced shift of the structure of the transition state.

IntroductionPolymorphic variants in the 5p15, 6p12, 6p21, and 15q25 loci were demonstrated to potentially contribute to lung cancer carcinogenesis. Therefore, this study was performed to assess the role of those variants in non-small cell lung cancer (NSCLC) risk and prognosis in a Portuguese population.Materials and MethodsBlood from patients with NSCLC was prospectively collected. To perform an association study, DNA from these patients and healthy controls were genotyped for a panel of 19 SNPs using a Sequenom® MassARRAY platform. Kaplan-Meier curves were used to assess the overall survival (OS) and progression-free survival (PFS).ResultsOne hundred and forty-four patients with NSCLC were successfully consecutively genotyped for the 19 SNPs. One SNP was associated with NSCLC risk: rs9295740 G/A. Two SNPs were associated with non-squamous histology: rs3024994 (VEGF intron 2) T/C and rs401681 C/T. Three SNPs were associated with response rate: rs3025035 (VEGF intron 7) C/T, rs833061 (VEGF –460) C/T and rs9295740 G/A. One SNP demonstrated an influence on PFS: rs401681 C/T at 5p15, p = 0.021. Four SNPs demonstrated an influence on OS: rs2010963 (VEGF +405 G/C), p = 0.042; rs3025010 (VEGF intron 5 C/T), p = 0.047; rs401681 C/T at 5p15, p = 0.046; and rs31489 C/A at 5p15, p = 0.029.ConclusionsOur study suggests that SNPs in the 6p12, 6p21, and 5p15 loci may serve as risk, predictive and prognostic NSCLC biomarkers. In the future, SNPs identified in the genomes of patients may improve NSCLC screening strategies and therapeutic management as well.

Along with nodal status and hormone receptor expression, DNA ploidy is an independent predictor of long-term survival in IDC.