Focal adhesion kinase is negatively regulated by phosphorylation at tyrosine 407

Focal adhesion kinase (FAK) mediates signal transduction in response to multiple extracellular inputs via tyrosine phosphorylation at specific residues. Although several tyrosine phosphorylation events have been linked to FAK activation and downstream signal transduction, the function of FAK phosphorylation at Tyr(407) was previously unknown. Here, we show for the first time that phosphorylation of FAK Tyr(407) increases during serum starvation, contact inhibition, and cell cycle arrest, all conditions under which activating FAK Tyr(397) phosphorylation decreases. Transfection of NIH3T3 cells with a phosphorylation-mimicking FAK 407E mutant decreased autophosphorylation at Tyr(397) and inhibited both FAK kinase activity in vitro and FAK-mediated functions such as cell adhesion, spreading, proliferation, and migration. The opposite effects were observed in cells transfected with nonphosphorylatable mutant FAK 407F. Taken together, these data suggest the novel concept that FAK Tyr(407) phosphorylation negatively regulates the enzymatic and biological activities of FAK.     

Impaired long-term memory and long-term potentiation in N-type Ca2+ channel-deficient mice

Voltage-dependent N-type Ca(2+) channels, along with the P/Q-type, have a crucial role in controlling the release of neurotransmitters or neuromodulators at presynaptic terminals. However, their role in hippocampus-dependent learning and memory has never been examined. Here, we investigated hippocampus-dependent learning and memory and synaptic plasticity at hippocampal CA3-CA1 synapses in mice deficient for the alpha(1B) subunit of N-type Ca(2+) channels. The mutant mice exhibited impaired learning and memory in the Morris water maze and the social transmission of food preference tasks. In particular, long-term memory was impaired in the mutant mice. Interestingly, among activity-dependent long-lasting synaptic changes, theta burst- or 200-Hz-stimulation-induced long-term potentiation (LTP) was decreased in the mutant, compared with the wild-type mice. This type of LTP is known to require brain-derived neurotrophic factor (BDNF). It was found that both BDNF-induced potentiation of field excitatory postsynaptic potentials and facilitation of the frequency of miniature excitatory postsynaptic currents (mEPSCs) were reduced in the mutant. Taken together, these results demonstrate that N-type Ca(2+) channels are required for hippocampus-dependent learning and memory, and certain forms of LTP.     

Differential analysis of Bacillus anthracis after pX01 plasmid curing and comprehensive data on Bacillus anthracis infection in macrophages and glial cells

Bacillus anthracis is a gram-positive bacterial organism responsible for anthrax. This organism has two pathogenic plasmids: pX01 and pX02. The genetic function of pX01, which comprises about 198 kb, is not known, except for a region called the pathogenic island, which contains three genes-pag, lef, and cya-that code for three toxic proteins. A 2-D difference gel electrophoresis (2-D DIGE) system was used to verify the existence of proteins controlled by the pX01 plasmid, and protein regulation data were obtained using DeCyder software. A total of 1728 proteins were identified in the wild-type strain of this organism and 1684 in the pX01 plasmid. Twenty-seven of these proteins disappeared and eight appeared when the pX01 plasmid was removed. An additional 52 proteins were downregulated and 15 were upregulated when this plasmid was removed. A total of 102 proteins have been identified using the MALDI-TOF method of analysis, including 49 whose functions are unknown. Among these, 31 participate in metabolic processes, two in cellular processes, 15 in the processing of genetic information, and five in the processing of extracellular information. Another seven proteins participate in bacterial virulence and pathogenesis. We investigated the functions of these proteins in other bacteria, particularly the B. anthracis derivative H9041. Bacterial growth differed between pX01+/pX02+ B. anthracis and its pX01-/pX02+ derivative as did the cytotoxicity of macrophages infected by pX01+/pX02+ B. anthracis and the pX01-pX02+ derivative. We also found that S100B protein levels increased in the host infected with pX01+/pX02+ B. anthracis or its pX01-/pX02+ derivative. These data suggest that the pX01 plasmid plays a key role in the regulation of protein functions in B. anthracis.     

Tomographic phase microscopy

We report a technique for quantitative three-dimensional (3D) mapping of refractive index in live cells and tissues using a phase-shifting laser interferometric microscope with variable illumination angle. We demonstrate tomographic imaging of cells and multicellular organisms, and time-dependent changes in cell structure. Our results will permit quantitative characterization of specimen-induced aberrations in high-resolution microscopy and have multiple applications in tissue light scattering.     

Increased beta-carotene production in recombinant Escherichia coli harboring an engineered isoprenoid precursor pathway with mevalonate addition

When pT-LYCm4 containing lycopene synthetic genes was co-transformed with pSUcrtY or pSHcrtY containing crtY gene of Pantoea ananatis (P. ananatis) or Pantoea agglomerans (P. agglomerans), beta-carotene productions of 36 and 35 mg/L were obtained, respectively. No lycopene was detected in the beta-carotene production culture. pT-HB, constructed by addition of P. ananatis crtY gene into pT-LYCm4, was used for co-transformation with pSdxs and pSSN12Didi, which increased isopentenyl diphosphate and dimethylallyl diphosphate synthesis. beta-Carotene production significantly increased 1.5-fold (51 mg/L) with the amplification of the dxs gene through pSdxs and 4-fold (135 mg/L) with the mevalonate bottom pathway of pSSN12Didi in the presence of 3.3 mM mevalonate. The pT-DHB, constructed by integrating the dxs gene into pT-HB, was used for cotransformation of Escherichia coli (E. coli) harboring pSSN12Didi, resulting in beta-carotene production of 141 mg/L. Recombinant E. coli harboring pT-DHB and pSSN12Didi was used to maximize beta-carotene production by adjusting the available amounts of glycerol, a carbon source, and mevalonate, the precursor of the mevalonate bottom pathway. When recombinant E. coli was given 16.5 mM mevalonate and 2.5% (w/v) glycerol, beta-carotene production of 503 mg/L in concentration and 49.3 mg/g DCW in content was obtained at 144 h, which was the highest level of carotenoid production in E. coli ever reported in the literature.     

Co-treatment with interferon-γ and 1-methyl tryptophan ameliorates cardiac fibrosis through cardiac myofibroblasts apoptosis

Molecular and Cellular Biochemistry - Electron transfer occurs through heme-Fe across the cytochrome c protein. The current models of long range electron transfer pathways in proteins include...     

Generation of single-chain Fv antibody fragments against Mu-2-related death-inducing gene in Escherichia coli

Molecular Biology Reports - Mu-2-related death-inducing (MuD) gene is involved in apoptosis in tumor cells. Although we have previously produced mouse monoclonal antibodies (MAbs) that specifically...     

Comparative genomic analysis of Lactobacillus mucosae LM1 identifies potential niche-specific genes and pathways for gastrointestinal adaptation

Lactobacillus mucosae is currently of interest as putative probiotics due to their metabolic capabilities and ability to colonize host mucosal niches. L. mucosae LM1 has been studied in its functions in cell adhesion and pathogen inhibition, etc. It demonstrated unique abilities to use energy from carbohydrate and non-carbohydrate sources. Due to these functions, we report the first complete genome sequence of an L. mucosae strain, L. mucosae LM1. Analysis of the pan-genome in comparison with closely-related Lactobacillus species identified a complete glycogen metabolism pathway, as well as folate biosynthesis, complementing previous proteomic data on the LM1 strain. It also revealed common and unique niche-adaptation genes among the various L. mucosae strains. The aim of this study was to derive genomic information that would reveal the probable mechanisms underlying the probiotic effect of L. mucosae LM1, and provide a better understanding of the nature of L. mucosae sp.     

Effects of various LED light spectra on circadian rhythm during starvation in the olive flounder (Paralichthys olivaceus)

In aquaculture, feeding is essential for the maintenance of metabolic processes and homoeostasis of fish. However, fasting acts as a stressor. In this study, we investigated the effect of circadian rhythm under various LED wavelengths [blue (460 nm), green (520 nm) and red (630 nm)] and two light intensities (0.3 and 0.6 W m^?2) over a 9-days period in the olive flounder (Paralichthys olivaceus). We analysed clock genes like period 2 (Per 2) and cryptochrome 1 (Cry 1), and serotonin and arylalkylamine-N-acetyltransferase 2 (AANAT 2), which control circadian rhythms. Per 2, Cry 1, serotonin and AANAT 2 were significantly decreased during the starvation period compared to the normal feeding group. Nevertheless, their levels increased in the groups exposed to green- and blue LED light during the experimental period. These results confirmed that green and blue wavelengths are effective in maintaining the circadian rhythm in olive flounder.