Path-preference cellular-automaton model for traffic flow through transit points and its application to the transcription process in human cells.

We all use path routing everyday as we take shortcuts to avoid traffic jams, or by using faster traffic means. Previous models of traffic flow of RNA polymerase II (RNAPII) during transcription, however, were restricted to one dimension along the DNA template. Here we report the modeling and application of traffic flow in transcription that allows preferential paths of different dimensions only restricted to visit some transit points, as previously introduced between the 5' and 3' end of the gene. According to its position, an RNAPII protein molecule prefers paths obeying two types of time-evolution rules. One is an asymmetric simple exclusion process (ASEP) along DNA, and the other is a three-dimensional jump between transit points in DNA where RNAPIIs are staying. Simulations based on our model, and comparison experimental results, reveal how RNAPII molecules are distributed at the DNA-loop-formation-related protein binding sites as well as CTCF insulator proteins (or exons). As time passes after the stimulation, the RNAPII density at these sites becomes higher. Apparent far-distance jumps in one dimension are realized by short-range three-dimensional jumps between DNA loops. We confirm the above conjecture by applying our model calculation to the SAMD4A gene by comparing the experimental results. Our probabilistic model provides possible scenarios for assembling RNAPII molecules into transcription factories, where RNAPII and related proteins cooperatively transcribe DNA.

Global analysis of RNA expression profile in human vascular cells treated with statins.

In addition to a lipid-lowering effect, 3-hydroxy-3-methylglutaryl coenzyme A reductase inhibitors (statins) have an effect on the expression levels of many genes. In order to elucidate the range of this effect as comprehensively as possible, we investigated the changes in gene expression profiles brought about by atorvastatin or pitavastatin in cultured human umbilical vein endothelial cells (HUVEC), cultured human coronary artery smooth muscle cells (HCASMC) and cultured human hepatocarcinoma Hep G2 cells by means of DNA microarrays. Among the 6146 genes in the array, statins affected the expression levels of genes involved in coagulation, vascular constriction and cell growth in a cell-type specific manner. In HUVEC, they induced integrin beta4 and thrombomodulin profoundly, and profoundly suppressed pentraxin 3 both at 8 and 24 hours. In HCASMC, the statins induced thrombomodulin and urokinase inhibitor, and potently suppressed the cysteine-rich angiogenic inducer 61 and cyclin B. Many genes related to the cell cycle and/or growth were also regulated in HUVEC and HCASMC by the statins. These results indicate that many aspects of the pleiotropic effect can be mediated by transcriptional control by statins. Genes newly identified by this study may be useful in statin therapy.

A comparison of differences in the gene expression profiles of phorbol 12-myristate 13-acetate differentiated THP-1 cells and human monocyte-derived macrophage.

In order to characterize the monocytic cell line THP-1 and its mature, macrophage-like form treated with phorbol 12-myristate 13-acetate (PMA), we have conducted an oligonucleotide microarray assay and compared the results with those from an assay of human monocytes and macrophages. We found that early THP-1 cells have a pattern of gene expression distinct from monocytes, and when treated with PMA, certain genes which are induced in macrophages, such as apolipoprotein-E, matrix metalloproteinase 9 and alpha2 macroglobulin are also induced in the PMA-treated THP-1 cells (THP1PMA cells). However, these were some genes which are conversely regulated among macrophages and THP1PMA cells such as interleukin-1-beta and the overall correlation coefficient was not very high. It is shown that, although certain morphological and other characteristics of PMA-differentiated THP-1 cells are similar to macrophages, from a transcriptomic view, the two are different. This suggests a need for careful recognition of and allowance for this difference when interpreting the results of experiments done with THP-1 cells in which it is otherwise assumed they are representative of the macrophage.

Cl(-) concentration dependence of photovoltage generation by halorhodopsin from Halobacterium salinarum.

The photovoltage generation by halorhodopsin from Halobacterium salinarum (shR) was examined by adsorbing shR-containing membranes onto a thin polymer film. The photovoltage consisted of two major components: one with a sub-millisecond range time constant and the other with a millisecond range time constant with different amplitudes, as previously reported. These components exhibited different Cl(-) concentration dependencies (0.1-9 M). We found that the time constant for the fast component was relatively independent of the Cl(-) concentration, whereas the time constant for the slow component increased sigmoidally at higher Cl(-) concentrations. The fast and the slow processes were attributed to charge (Cl(-)) movements within the protein and related to Cl(-) ejection, respectively. The laser photolysis studies of shR-membrane suspensions revealed that they corresponded to the formation and the decay of the N intermediate. The photovoltage amplitude of the slow component exhibited a distorted bell-shaped Cl(-) concentration dependence, and the Cl(-) concentration dependence of its time constant suggested a weak and highly cooperative Cl(-)-binding site(s) on the cytoplasmic side (apparent K(D) of approximately 5 M and Hill coefficient > or =5). The Cl(-) concentration dependence of the photovoltage amplitude and the time constant for the slow process suggested a competition between spontaneous relaxation and ion translocation. The time constant for the relaxation was estimated to be >100 ms.

Detection of an up-regulation of a group of chemokine genes in murine cardiac allograft in the absence of interferon-gamma by means of DNA microarray.

BACKGROUND: [corrected] Interferon (IFN)-gamma and the IFN-gamma-dependent pathway are prominent in vascularized allograft during acute rejection. However, IFN-gamma deficient (IFN-gamma-/-) mice can rapidly reject cardiac allografts. To bring the alternative pathway during allograft rejection into more precise focus, we investigated the gene expression profile in murine cardiac allografts in IFN-gamma-/- mice by means of DNA microarray. MATERIAL AND METHOD: We screened for gene expression changes in murine cardiac allografts of BALB/c H-2d into both wild-type C57BL/6 H-2b (n=3) and IFN-gamma-/- C57BL/6 H-2b(IFN-gamma-/-, n=4) using Affymetrix oligonucleotide arrays to monitor more than 11,000 genes and expressed sequence tag (ESTs). The heart was heterotopically transplanted. Transplanted hearts were harvested on day 5. As a control, isografts (C57BL/6 to C57BL/6) were also harvested on day 5. RESULTS: On day 5, 64 of the 84 genes induced in the allografts in wild-type mice were not up-regulated in IFN-gamma-/- mice. We identified a group of 54 genes that were up-regulated in allografts in IFN-gamma-/- mice. Several chemokine genes, including monocyte chemoattractant protein=1 and macrophage inflammatory protein, were induced in the allografts in both wild-type and IFN-gamma-/- mice. Interestingly, a group of genes, including C10-like chemokine and platelet factor 4, were specifically induced in the IFN-gamma-/- mice. CONCLUSION: DNA microarray analysis reveals a unique pattern of mRNA expression in allografts in IFN-gamma-/- mice as well as a group of genes induced in cardiac allografts in both wild-type and IFN-gamma-/- mice, including monocyte chemoattractant protein-1 and monocyte chemoattractant protein-1.

Two forms of NAD-dependent D-mandelate dehydrogenase in Enterococcus faecalis IAM 10071.

Two forms of NAD-dependent D-mandelate dehydrogenase (D-ManDHs) were purified from Enterococcus faecalis IAM 10071. While these two enzymes consistently exhibited high activity toward large 2-ketoacid substrates that were branched at the C3 or C4 position, they gave distinctly different K(m) and V(max) values for these substrates and had distinct molecular weights by gel electrophoresis and gel filtration.