Acetylation of Rb by PCAF is required for nuclear localization and keratinocyte differentiation

Although the retinoblastoma protein (Rb) functions as a checkpoint in the cell cycle, it also regulates differentiation. It has recently been shown that Rb is acetylated during differentiation; however, the role of this modification has not been identified. Depletion of Rb levels with short hairpin RNA resulted in inhibition of human keratinocyte differentiation, delayed cell cycle exit and allowed cell cycle re-entry. Restoration of Rb levels rescued defects in differentiation and cell cycle exit and re-entry; however, re-expression of Rb with the major acetylation sites mutated did not. During keratinocyte differentiation, acetylation of Rb is mediated by PCAF and it is further shown that PCAF acetyltransferase activity is also required for normal differentiation. The major acetylation sites in Rb are located within the nuclear localization sequence and, although mutation did not alter Rb localization in cycling cells, the mutant is mislocalized to the cytoplasm during differentiation. Studies indicate that acetylation is a mechanism for controlling Rb localization in human keratinocytes, with either reduction of the PCAF or exogenous expression of the deacetylase SIRT1, resulting in mislocalization of Rb. These findings identify PCAF-mediated acetylation of Rb as an event required to retain Rb within the nucleus during keratinocyte differentiation.

Molecular Logic Gates and Luminescent Sensors Based on Photoinduced Electron Transfer

The competition between Photoinduced electron transfer (PET) and other de-excitation pathways such as fluorescence and phosphorescence can be controlled within designed molecular structures. Depending on the particular design, the resulting optical output is thus a function of various inputs such as ion concentration and excitation light dose. Once digitized into binary code, these input-output patterns can be interpreted according to Boolean logic. The single-input logic types of YES and NOT cover simple sensors and the double- (or higher-) input logic types represent other gates such as AND and OR. The logic-based arithmetic processors such as half-adders and half-subtractors are also featured. Naturally, a principal application of the more complex gates is in multi-sensing contexts.

Molecular cloning, expression, and regulation in Escherichia coli K-12 of a chromosome-mediated aerobactin iron transport system from a human invasive isolate of E. coli K1

We have cloned chromosomal genes determining the aerobactin iron transport system from the Escherichia coli K1 strain VW187. Mapping and hybridization experiments showed that the VW187 aerobactin region was identical to that of the plasmid ColV-K30. However, in the E. coli K-12 background, the biosynthesis of both siderophore and ferric aerobactin receptor encoded by the VW187-derived recombinant plasmids was not repressed by iron to the same extent found when a recombinant plasmid derived from pColV-K30 was used. RNA-DNA dot-blot hybridization experiments demonstrated that the aerobactin-specific mRNA synthesized by the VW187-derived clones was not iron regulated in E. coli K-12. In contrast, the synthesis of aerobactin and its receptor in strain VW187 was completely repressed by iron regardless of whether the recombinant plasmids originated from VW187 or pColV-K30. Similar results were obtained with gene fusions in which a promoterless lac operon was placed under the control of aerobactin promoter regions of either chromosome- or plasmid-mediated aerobactin systems. DNA sequencing of the chromosomal aerobactin promoter region showed changes in bases located immediately upstream to the -35 region compared with the corresponding region in pColV-K30, which is known to be part of the binding site for the Fur repressor protein.

Green tea supplementation increases glutathione and plasma antioxidant capacity in adults with the metabolic syndrome

Green tea, a popular polyphenol-containing beverage, has been shown to alleviate clinical features of the metabolic syndrome. However, its effects in endogenous antioxidant biomarkers are not clearly understood. Thus, we tested the hypothesis that green tea supplementation will upregulate antioxidant parameters (enzymatic and nonenzymatic) in adults with the metabolic syndrome. Thirty-five obese participants with the metabolic syndrome were randomly assigned to receive one of the following for 8 weeks: green tea (4 cups per day), control (4 cups water per day), or green tea extract (2 capsules and 4 cups water per day). Blood samples and dietary information were collected at baseline (0 week) and 8 weeks of the study. Circulating carotenoids (a-carotene, ß-carotene, lycopene) and tocopherols (a-tocopherol, ?-tocopherol) and trace elements were measured using high-performance liquid chromatography and inductively coupled plasma mass spectroscopy, respectively. Serum antioxidant enzymes (glutathione peroxidase, glutathione, catalase) and plasma antioxidant capacity were measured spectrophotometrically. Green tea beverage and green tea extract significantly increased plasma antioxidant capacity (1.5 to 2.3 µmol/L and 1.2 to 2.5 µmol/L, respectively; P <.05) and whole blood glutathione (1783 to 2395 µg/g hemoglobin and 1905 to 2751 µg/g hemoglobin, respectively; P <.05) vs controls at 8 weeks. No effects were noted in serum levels of carotenoids and tocopherols and glutathione peroxidase and catalase activities. Green tea extract significantly reduced plasma iron vs baseline (128 to 92 µg/dL, P <.02), whereas copper, zinc, and selenium were not affected. These results support the hypothesis that green tea may provide antioxidant protection in the metabolic syndrome.

Molecular pedigree reconstruction and estimation of evolutionary parameters in a wild Atlantic salmon river system with incomplete sampling: a power analysis

Background: Pedigree reconstruction using genetic analysis provides a useful means to estimate fundamental population biology parameters relating to population demography, trait heritability and individual fitness when combined with other sources of data. However, there remain limitations to pedigree reconstruction in wild populations, particularly in systems where parent-offspring relationships cannot be directly observed, there is incomplete sampling of individuals, or molecular parentage inference relies on low quality DNA from archived material. While much can still be inferred from incomplete or sparse pedigrees, it is crucial to evaluate the quality and power of available genetic information a priori to testing specific biological hypotheses. Here, we used microsatellite markers to reconstruct a multi-generation pedigree of wild Atlantic salmon (Salmo salar L.) using archived scale samples collected with a total trapping system within a river over a 10 year period. Using a simulation-based approach, we determined the optimal microsatellite marker number for accurate parentage assignment, and evaluated the power of the resulting partial pedigree to investigate important evolutionary and quantitative genetic characteristics of salmon in the system.

Results: We show that at least 20 microsatellites (ave. 12 alleles/locus) are required to maximise parentage assignment and to improve the power to estimate reproductive success and heritability in this study system. We also show that 1.5 fold differences can be detected between groups simulated to have differing reproductive success, and that it is possible to detect moderate heritability values for continuous traits (h(2) similar to 0.40) with more than 80% power when using 28 moderately to highly polymorphic markers.

Conclusion: The methodologies and work flow described provide a robust approach for evaluating archived samples for pedigree-based research, even where only a proportion of the total population is sampled. The results demonstrate the feasibility of pedigree-based studies to address challenging ecological and evolutionary questions in free-living populations, where genealogies can be traced only using molecular tools, and that significant increases in pedigree assignment power can be achieved by using higher numbers of markers.

Nuclear translocation and functions of growth factor receptors

Cellular signal transduction in response to environmental signals involves a relay of precisely regulated signal amplifying and damping events. A prototypical signaling relay involves ligands binding to cell surface receptors and triggering the activation of downstream enzymes to ultimately affect the subcellular distribution and activity of DNA-binding proteins that regulate gene expression. These so-called signal transduction cascades have dominated our view of signaling for decades. More recently evidence has accumulated that components of these cascades can be multifunctional, in effect playing a conventional role for example as a cell surface receptor for a ligand whilst also having alternative functions for example as transcriptional regulators in the nucleus. This raises new challenges for researchers. What are the cues/triggers that determine which role such proteins play? What are the trafficking pathways which regulate the spatial distribution of such proteins so that they can perform nuclear functions and under what circumstances are these alternative functions most relevant?