Computational Mechanistic Study of Thionation of Carbonyl Compounds with Lawesson's Reagent.

The thionation reaction of carbonyl compounds with Lawesson's reagent (LR) has been studied using density functional theory methods and topological analyses. After dissociation of LR, the reaction takes place through a two-step mechanism involving (i) a concerted cycloaddition between one monomer and the carbonyl compound to form a four-membered intermediate and (ii) a cycloreversion leading to the thiocarbonyl derivative and phenyl(thioxo)phosphine oxide. Topological analyses confirmed the concertedness and asynchronicity of the process. The second step is the rate-limiting one, and the whole process resembles the currently accepted mechanism for the lithium salt-free Wittig reaction. No zwitterionic intermediates are formed during the reaction, although stabilizing electrostatic interactions are present in initial stages. Phenyl(thioxo)phosphine oxide formed in the thionation reaction is capable of performing a second thionation, although with energy barriers higher than the first one. The driving force of the thionation reactions is the formation of trimers from the resulting monomers. In agreement with experimental observations, the amides are the most reactive when compared with esters, aldehydes, and ketones and the reaction is slightly influenced by the polarity of the solvent. Whereas for amides and esters substituents have little effect, aldehydes and ketones are influenced by both steric and electronic effects.

RhoA activity is required for fibronectin assembly and counteracts beta1B integrin inhibitory effect in FRT epithelial cells.

FRT thyroid epithelial cells synthesize fibronectin and organize a network of fibronectin fibrils at the basal surface of the cells. Fibronectin fibril formation is enhanced by the overexpression of the ubiquitous beta1A integrin and is inhibited by the expression of the dominant-negative beta1B subunit. We tested the hypotheses that RhoA activity might mediate the integrin-dependent fibronectin fibrillogenesis and might counteract beta1B integrin inhibitory effect. FRT-beta1A cells were transfected with a vector carrying a dominant negative form of RhoA (RhoAN19) or treated with the C3 transferase exoenzyme. Both treatments inhibited fibronectin assembly and caused loss of actin microfilaments and adhesion plaques. On the other hand, FRT-beta1B cells were transfected with the constitutively activated form of RhoA (RhoAV14) or treated with the E. coli cytotoxic necrotizing factor 1, which directly activates RhoA. Either treatment restored microfilament and adhesion plaque assembly and promoted fibronectin fibril organization. A great increase in fibronectin fibril assembly was also obtained by treatment of FRT-beta1B cells with TGF-beta. Our data indicate that RhoA is required to promote fibronectin matrix assembly in FRT cells and that the activation of the signal transduction pathway downstream of RhoA can overcome the inhibitory effect of beta1B integrin.