Studies on anaerobic proteolytic bacteria of Leh,India

Five anaerobic proteolytic bacteria were isolated from water bodies of Leh, India, where the ambient temperature varies from –25 to 25 °C. Isolates showed growth at all temperatures ranging from 5 to 37 °C except SPL-4 and SPL-5 which showed no growth at 5 °C. The cultures could grow and produce proteases on various protein substrates and the yield varied with the substrates. Two of the cultures showed the presence of spores. Acetate was the dominant VFA during hydrolysis of protein substrates.     

Bioemulsifier production by Streptomyces sp. S22 isolated from garden soil

Out of 45 actinomycetes isolated from garden soil, pond water and air; fifteen showed good emulsification activity. Streptomyces sp. S22 isolated from garden soil produced maximum bioemulsifier with 0.5% (v/v) sunflower oil during stationary phase at 37 degrees C, pH 6 and 250 rev/min. Emulsification activity was maximum (320 EU/ml) with sunflower oil as substrate. Partially purified bioemulsifier from Streptomyces sp. S22 was a peptidoglycolipid containing lipid (51.25%), protein (30%), non-reducing sugar (17.75%) and reducing sugar (1%). The yield of partially purified bioemulsifier was 1.6 g/l and reduced the surface tension of water by 23.09 mN/m. The bioemulsifier produced by Streptomyces sp. S22 was stable at room temperature for seven days.     

Virus induced chromosomal abnormalities in Chinese hamster lung cell line and human peripheral blood leukocyte culture

Chinese hamster lung (CHL) cells were susceptible to Herpes Simplex type-1 and Chandipura viruses; which induced chromosomal abnormalities in these cells. Chromosomal changes induced in these cells were specific. The cells were refractory to measles virus and chromosomal abnormalities were not detected after inoculation of the virus. On the other hand human peripheral blood (HPB) leukocytes were susceptible to all the 3 viruses studied and exhibited chromosomal abnormalities upon infection. The aberrations induced in HPBL cultures were random. The results suggest that a virus could induce chromosomal changes only in susceptible cells. This is the first report of comparative in vitro study on chromosomes.     

The molecular origin and consequences of escape from miRNA regulation by HLA-C alleles

Differential expression of human leukocyte antigen C (HLA-C) allotypes is mediated by the binding of a microRNA, miR-148a, to the 3′ untranslated region of some, but not all, HLA-C alleles. The binding results in lower levels of HLA-C expression, which is associated with higher levels of HIV-1 viral load among infected individuals. The alternative set of HLA-C alleles has several substitutions in the miR-148a binding site that prevent binding and HLA-C downregulation; these high-expression alleles associate with control of HIV-1 viral load. We show that the common ancestor of all extant HLA-C alleles was suppressed by miR-148a. Substitutions that prevent miR-148a binding arose by a sequence exchange event between an HLA-C allele and an HLA-B (MIM 142830) allele of a B^∗07-like lineage. The event occurred 3–5 million years ago, resulting in an HLA-C variant that escape from miR-148a downregulation. We present evidence suggesting that selection played a role in the successful spread of the HLA-C escape alleles, giving rise to 7 of the 14 extant HLA-C lineages. Notably, critical peptide and KIR binding residues of the escape variants have selectively converged to resemble the sequence of their inhibited counterparts, such that the inhibited and escape groupings differ primarily by their levels of expression.     

Cloning and expression of a tyrosinase from <Emphasis Type="Italic">Aspergillus oryzae</Emphasis> in <Emphasis Type="Italic">Yarrowia lipolytica</Emphasis>: application in <Emphasis Type="SmallCaps">l</Emphasis>-DOPA biotransformation

Amplification of the tyrosinase gene (melO) from the genomic DNA of Aspergillus oryzae NCIM 1212 yielded a 1.6-kb product. This gene was cloned into pYLEX1, and the resulting pTyro-YLEX1 vector was transformed in Yarrowia lipolytica strain Po1g. A clone displaying the highest specific activity for tyrosinase (10.94 U/mg) was used for obtaining the complementary DNA (cDNA) and for protein expression studies. cDNA sequence analysis indicated the splicing of an intron present in the melO gene by Po1g. Native polyacrylamide gel electrophoresis, acidification at pH 3.0 followed by activity staining with l-DOPA indicated the expression of an active tyrosinase. The clone over-expressing the tyrosinase transformed l-tyrosine to l-DOPA. On optimization of conditions for the biotransformation (pH 4.0, temperature 60°C and with 3.5 mg of biomass), 0.4 mg/ml of l-DOPA was obtained.     

Signalling mechanisms in the regulation of vacuolar ion release in guard cells

Pharmacological agents were used to investigate the possible involvement of actin in signalling chains associated with abscisic acid (ABA)-induced ion release from the guard cell vacuole, a process which is absolutely essential for stomatal closure. Effects on the ABA-induced transient stimulation of tonoplast efflux were measured, using (86)Rb in isolated guard cells of Commelina communis, together with effects on stomatal apertures. In the response to 10 microm ABA (triggered by Ca(2+) influx rather than internal Ca(2+) release), jasplakinolide (stabilizing actin filaments) and latrunculin B (depolymerizing actin filaments) had opposite effects. Both closure and the vacuolar efflux transient were inhibited by jasplakinolide but enhanced by latrunculin B. At 10 microm ABA prevention of mitogen-activated protein (MAP) kinase activation by PD98059 partially inhibited closure and reduced the efflux transient. By contrast, latrunculin B inhibited the efflux transient at 0.1 microm ABA (involving internal Ca(2+) release rather than Ca(2+) influx). The results suggest that 10 microm ABA activates Ca(2+)-dependent vacuolar ion efflux via a Ca(2+)-permeable influx channel which is maintained closed by interaction with F-actin. A MAP kinase is also involved, in a chain similar to that postulated for Ca(2+)-dependent gene expression in cold acclimation.     

Molecular genetics of schizophrenia: past, present and future

Schizophrenia is a severe neuropsychiatric disorder with a polygenic mode of inheritance which is also governed by non-genetic factors. Candidate genes identified on the basis of biochemical and pharmacological evidence are being tested for linkage and association studies. Neurotransmitters, especially dopamine and serotonin have been widely implicated in its etiology. Genome scan of all human chromosomes with closely spaced polymorphic markers is being used for linkage studies. The completion and availability of the first draft of Human Genome Sequence has provided a treasure-trove that can be utilized to gain insight into the so far inaccessible regions of the human genome. Significant technological advances for identification of single nucleotide polymorphisms (SNPs) and use of microarrays have further strengthened research methodologies for genetic analysis of complex traits. In this review, we summarize the evolution of schizophrenia genetics from the past to the present, current trends and future direction of research.     

Characterization of porcine stable kidney cell line adapted to hyperthermic temperature

The optimum temperature for the growth of porcine stable (PS) kidney cell line is 37 degrees C. We have adapted the cell line to grow at 40 degrees C. The original cell line grown at 37 degrees C has been denoted as PS-37, and the adapted new strain has been denoted as PS-40. Both the cell lines were screened for mycoplasma by Hoechst staining and tritiated uridine-uracil uptake and were found to be negative. Comparative characterization of PS-40 and its progenitor PS-37 cell line was done by using various parameters. The antigenic studies indicated that the new cell strain was not cross-contaminated with any other cell lines. It was observed that PS-40 cells were more fibroblastic with clean cytoplasm and appeared healthy. The growth of PS-40 cells was faster than the original cell line. The karyological study showed heteroploid chromosome number in PS-40 cells. The modal chromosome number of PS-40 cells was 58, whereas that of PS-37 cell line was 38. The lactic dehydrogenase isoenzyme pattern showed a cathodal shift of bands. The PS-40 cell strain could be cryopreserved and revived. The viability of PS-37 as well as PS-40 cell lines is in the range of 90-95%, and the growth characteristics of thawed cells showed six- to eightfold multiplications within 5 d. The virus susceptibility study revealed that the cytopathic effect was more profound and observed 1 d earlier in PS-40 cell line. Increased yields of Japanese encephalitis, Sindbis, and Semliki forest viruses were obtained by 1.8, 1.75, and 1.5 log plaque-forming units/ml, respectively. The yield of West Nile virus was, however, comparable to that in PS-37 cell line. Both the cell lines were refractory to Dengue viruses.