Anin vitro system to study drug sensitivity ofMycobacterium leprae using infected human tissue

A reliable screening technique for assessing the sensitivity ofMycobacterium leprae to drugs has been developed. The method is based on the susceptibility or otherwise ofM. leprae- infected tissues from lepromatous leprosy patients to the action of diaminodiphenyl sulphone (dapsone) or rifampicin on the incorporation of [¹⁴C]-acetate into lipids. The extent of inhibition or lack of inhibition correlated very well with the drug sensitivity or resistance of the bacteria isolated from the patients to the above drugs. A similar trend was observed when the incorporation into individual fractions of neutral lipids was measured. There was no incorporation by heat-killed tissues. This method correlates well with the 3,4-dihydroxyphenylalanine uptake studies.     

RAPD profile based genetic characterization of chemotypic variants of Artemisia annua L.

The annual herbaceous plant, Artemisia annua L., belonging to family Asteraceae, is the natural source of the highly potent antimalarial compound, artemisinin, besides producing valuable essential oil. The plant is at present the sole commercial source for artemisinin production since all the chemical syntheses are non-viable. Therefore, economic and practical considerations dictate that plants with maximum content of artemisinin be found and/or ways to increase their artemisinin content be sought. The key to this selection and breeding is a comprehension of chemical and genetic variability and suitable selection(s) of elites from within the available population. In the present study, RAPD analyses of selected chemotypes from a decade old introduced population in India were carried out using arbitrary primers. The RAPD data clearly indicate the distinction amongst these plants. Further, the detection of highly polymorphic profiles (97 polymorphic markers out of a total of 101 markers) suggests the existence of very high levels of genetic variation in the Indian population despite geographical isolation and opens out a strong possibility of further genetic improvement for superior artemisinin content. UPGMA analyses of RAPD and phytochemical trait data indicate that the wide phytochemical diversity is included within the genetic diversity. These results further support the prospects for selection and breeding of superior artemisinin containing lines.     

Isolation of methanogens from Arabian sea sediments and their salt tolerance

Abstract Sea sediments in tropical regions have been less studied for methanogenesis and methanogens present therein. Three species of methanogens viz. Methanobacterium bryantii, Methanococcus voltae and Methanosarcina barkeri were isolated from Arabian sea sediments collected near the west coast of India. Maximum methane was formed by M. voltae at 3.0% (w/v) NaCl and other two methanogens at 0.06% (w/v) NaCl. M. bryantii and M. barkeri tolerated 2.5 and 3.0% (w/v) NaCl respectively due to which these methanogens must have survived in salt conditions of the sea sediments.     

Effects of Ageing on Amylase Activity and Scutellar Cell Structure during Imbibition in Wheat Seed

In wheat seed the scutellum plays an important role in the hydrolysisof stored substrate during germination. This layer is activatedfirst, whilst the aleurone becomes activated later. A good correlationexists between the initiation of visible germination and theappearance of enzyme activity in the scutellum. Enzyme activityin the aleurone becomes apparent only when the germinating seedlingreaches the rapid growth phase. Electron microscopic observationsshow that during the later stages of germination the scutellarcells develop finger like projections. These may serve to absorbendospermic reserves hydrolysed by aleurone amylase. The scutellumof aged non-germinating seeds showed no amylase activity andno finger like projections were produced even after prolongedimbibition.Copyright 1993, 1999 Academic Press Wheat (Triticum aestivum L.), deteriorated, germination, scutellum, scanning electron microscopy, aleurone     

Alteration of macromolecular events and elemental levels in the skin of UVC exposed hairless mice

Swiss mice (S/RV/Cri-ba) were exposed to a spectral range of UV light emitting predominantly lambda 253.7 nm. Following a cumulative dose of 22.5 X 10(3) KJ/m2 tumours were induced. The tumour development occurred after 52 weeks, relatively a longer time interval following exposure, compared with shorter time intervals required for production of tumours in mice by spectral range UVA and UVB as observed by other workers. A study of biochemical events viz. levels of protein, DNA, zinc, iron, sodium and potassium has been made in the skin samples of control and irradiated animals following ultraviolet irradiation with a dose of 22.5 X 10(3) KJ/m2. Study of macromolecular events in the skin of control and irradiated mice, showed fluctuations in the levels of DNA. A particularly notable event is the occurrence of increased levels of DNA and zinc and their persistence during the 9-39 weeks post UV interval prior to tumour production. No such variation was observed in the control group in any of the intervals. Increased levels were also seen in case of iron, sodium and potassium at different intervals in the post UV periods. These fluctuations in various biochemical events are deemed to be indicative of UV initiated biochemical changes.     

MITOL-dependent ubiquitylation negatively regulates the entry of PolγA into mitochondria

Mutations in mitochondrial replicative polymerase PolγA lead to progressive external ophthalmoplegia (PEO). While PolγA is the known central player in mitochondrial DNA (mtDNA) replication, it is unknown whether a regulatory process exists on the mitochondrial outer membrane which controlled its entry into the mitochondria. We now demonstrate that PolγA is ubiquitylated by mitochondrial E3 ligase, MITOL (or MARCH5, RNF153). Ubiquitylation in wild-type (WT) PolγA occurs at Lysine 1060 residue via K6 linkage. Ubiquitylation of PolγA negatively regulates its binding to Tom20 and thereby its mitochondrial entry. While screening different PEO patients for mitochondrial entry, we found that a subset of the PolγA mutants is hyperubiquitylated by MITOL and interact less with Tom20. These PolγA variants cannot enter into mitochondria, instead becomes enriched in the insoluble fraction and undergo enhanced degradation. Hence, mtDNA replication, as observed via BrdU incorporation into the mtDNA, was compromised in these PEO mutants. However, by manipulating their ubiquitylation status by 2 independent techniques, these PEO mutants were reactivated, which allowed the incorporation of BrdU into mtDNA. Thus, regulated entry of non-ubiquitylated PolγA may have beneficial consequences for certain PEO patients.

This study shows that mitochondrial entry of the replicative polymerase PolgA is regulated by ubiquitylation by the E3 ligase MITOL; however, by manipulating their ubiquitylation status, some progressive external ophthalmoplegia mutants whose PolgA is polyubiquitylated and cannot enter the mitochondrion can be reactivated and hence become functionally active.