Light Effects in Yeast: Evidence for Participation of Cytochromes in Photoinhibition of Growth and Transport in Saccharomyces cerevisiae Cultured at Low Temperatures

Visible light of moderate intensity inhibits growth, respiration, protein synthesis, and membrane transport in bakers' yeast and has a deleterious effect on membrane integrity. The results of this study indicate that these effects require the presence of cytochromes b and a/a(3). The light sensitivities of growth rate and [(14)C]histidine uptake in wild-type rho(+) Y185 and D225-5A strains of Saccharomyces cerevisiae were compared with those in a variety of mutants lacking cytochrome b or a/a(3) or both; a close correlation was found between the presence of these respiratory pigments and photosensitivity. Thus, strain TL5-3C, a nuclear petite lacking cytochromes b, a, and a(3), was resistant to light; strain GL5-6A, another nuclear petite having reduced amounts of cytochromes a and a(3), was partially resistant; strains MB127-20C and MB1-6C, nuclear petites lacking only cytochrome b, were also only partially resistant to light; whereas mutants containing all three cytochromes but having their respiratory chain either nonfunctional (strain ZK3-6B) or uncoupled (strain 18-27/t12) were fully sensitive to light. Finally, an equal-energy, broad-band action spectrum for the light inhibition of growth and transport indicated that blue light (408 nm) was most effective; these wavelengths correspond to the Soret region of the cytochrome absorption spectrum. The results suggest, therefore, that the yeast cytochromes b, a, and a(3) are the primary photoreceptors for the inhibitory effects of light and, perhaps, for other processes, such as the entrainment of biological rhythms in this species.     

Development of an amperometric biosensing method for the determination of L-fucose in pretreated urine

The first amperometric biosensing method for the determination of L-fucose is described. L-Fucose is the objective of much current research, as it is considered as a potential marker for various pathologic disorders. Recombinant L-fucose dehydrogenase, having as cofactor beta-nicotinamide adenine dinucleotide phosphate (NAD+P), was cross-linked in a water-soluble photosensitive polymer matrix, that is, polyvinyl alcohol (PVA) modified with styrylpyridinium (SbQ), in the presence of BSA and glutaraldehyde. The resulting membrane was sandwiched between two polycarbonate membranes and was mounted in an amperometric cell. The oxidation of the enzymatically produced NADPH was monitored at a platinum anode at +0.25 V versus a silver pseudoreference electrode in the presence of ferricyanide. The system was fully optimized with respect to various analytical parameters. Regarding to the mechanical properties of the membrane and the storage stability of the immobilized enzyme, various parameters were also optimized. Several methods for the pretreatment of urine samples were investigated. Treatment of the samples with PbO2 found to eliminate the interference effect of various electroactive species exist in urine; optimum incubation time was determined since at prolonged incubation times L-fucose is also affected. Calibration curves for the direct and the mediated monitoring of NADPH were liner over the concentration ranges 0.04-1.0 mM (r2=0.9995) and 0.03-3.0 mM (r2=0.9997) fucose, respectively. The detection limits (S/N 3) were 2 and 1.5 microM fucose, respectively. The R.S.D. of the mediated biosensor is better than 1.5% (n=10, 0.5 mM fucose). The proposed biosensor correlates well with a reference enzymatic method and exhibits very good working and storage stability.     

Monoclonal antibodies in the treatment of lymphoid malignancies

Rituximab (Rit) was the first monoclonal antibody approved for therapeutic use in cancer patients. Rit is a chimeric mouse/human monoclonal antibody, consisting of the human IgG1 and k constant Fc region, and a mouse variable Fab region specific against the B-cell antigen CD20. Rit exerts its antilymphoma activity through many different mechanisms. Binding of antibody to CD20 antigen, provokes apoptosis through downstream signals that lead to caspase-3 activation. Complement activation by the Fc portion of the antibody results in complement-dependent cytotoxicity. However, the most effective mechanism of action seems to be antigen-dependent cellular cytotoxicity. Effector cytotoxic cells such as natural killer cells (NK) are activated after binding to the Fc portion of the anti-CD20 molecule. Activated NK cells kill the coated lymphoma cells with the use of granzyme-perforin system. More recently, pre-clinical data support the concept that Rituximab can provoke a vaccination-like effect. Finally in-vitro experiments and clinical trials have shown that co-administration of the antibody with cytotoxics confers a strong synergistic effect. The relative contribution of these mechanisms in vivo and in different lymphoma subtypes is not well known and remains to be further evaluated.

Among the different histological groups, follicular lymphoma (FL) has been proven to be the most sensitive to Rit when used as a single agent, with overall response rates of 80% and 50% in untreated and previously treated patients, respectively. Moreover, Rit in combination with chemotherapy is superior to chemotherapy alone in terms of response rate and event-free survival, while early data indicate a significant prolongation in overall survival as well. Similarly, the addition of Rit to standard chemotherapy improves the disease-free and overall survival of patients with diffuse large B-cell lymphoma. There is no doubt that Rit represents one of the greatest achievements of biotechnology engineering. However, we need to understand better the mechanisms of its action as well as the mechanisms of resistance to Rit, in order to design more effective treatment modalities.     

Subjective visual perception: from local processing to emergent phenomena of brain activity

The combination of electrophysiological recordings with ambiguous visual stimulation made possible the detection of neurons that represent the content of subjective visual perception and perceptual suppression in multiple cortical and subcortical brain regions. These neuronal populations, commonly referred to as the neural correlates of consciousness, are more likely to be found in the temporal and prefrontal cortices as well as the pulvinar, indicating that the content of perceptual awareness is represented with higher fidelity in higher-order association areas of the cortical and thalamic hierarchy, reflecting the outcome of competitive interactions between conflicting sensory information resolved in earlier stages. However, despite the significant insights into conscious perception gained through monitoring the activities of single neurons and small, local populations, the immense functional complexity of the brain arising from correlations in the activity of its constituent parts suggests that local, microscopic activity could only partially reveal the mechanisms involved in perceptual awareness. Rather, the dynamics of functional connectivity patterns on a mesoscopic and macroscopic level could be critical for conscious perception. Understanding these emergent spatio-temporal patterns could be informative not only for the stability of subjective perception but also for spontaneous perceptual transitions suggested to depend either on the dynamics of antagonistic ensembles or on global intrinsic activity fluctuations that may act upon explicit neural representations of sensory stimuli and induce perceptual reorganization. Here, we review the most recent results from local activity recordings and discuss the potential role of effective, correlated interactions during perceptual awareness.     

Adducts of guanine with chromium(III), iron(III) and oxovanadium(IV) chlorides

The preparation of well-defined adducts of the M(guH)(₂Cl₃ (M = Cr, Fe) and VO(guH)Cl₂ types (guH = neutral guanine), by refluxing ligand and metal chloride mixtures in ethanol-triethyl orthoformate, is reported. Characterization studies suggest that the new complexes are probably linear chain-like polymeric species, involving single bridges of bidentate guH ligands between adjacent metal ions. Bidentate bridging guH is most probably coordinated through the N(7) and N(9) imidazole nitrogens. The chloro ligands present in the adducts are exclusively terminal. Infrared evidence rules out the possibility of coordination of guanine through either of its exocyclic potential binding sites (i.e., CO oxygen and NH₂ nitrogen) [1].