Metabolic adaptability shifts of cell membrane fatty acids of Komagataeibacter hansenii HDM1-3 improve acid stress resistance and survival in acidic environments.

Komagataeibacter hansenii HDM1-3 (K. hansenii HDM1-3) has been widely applied for producing bacterial cellulose (BC). The yield of BC has been frequently limited by the acidification during sugar metabolism, due to the generation of organic acids such as acetic acid. In this study, the acid resistance mechanism of K. hansenii HDM1-3 has been investigated from the aspect of metabolic adaptability of cell membrane fatty acids. Firstly, we observed that the survival rate of K. hansenii HDM1-3 was decreased with lowered pH values (adjusted with acetic acids), accompanied by increased leakage rate. Secondly, the cell membrane adaptability in response to acid stress was evaluated, including the variations of cell membrane fluidity and fatty acid composition. The proportion of unsaturated fatty acids was increased (especially, C18-1w9c and C19-Cyc), unsaturation degree and chain length of fatty acids were also increased. Thirdly, the potential molecular regulation mechanism was further elucidated. Under acid stress, the fatty acid synthesis pathway was involved in the structure and composition variations of fatty acids, which was proved by the activation of both fatty acid dehydrogenase (des) and cyclopropane fatty acid synthase (cfa) genes, as well as the addition of exogenous fatty acids. The fatty acid synthesis of K. hansenii HDM1-3 may be mediated by the activation of two-component sensor signaling pathways in response to the acid stress. The acid resistance mechanism of K. hansenii HDM1-3 adds to our knowledge of the acid stress adaptation, which may facilitate the development of new strategies for improving the industrial performance of this species under acid stress.

Salt and oxidative stress tolerance in Debaryomyces hansenii and Debaryomyces fabryi.

We report the characterization of five strains belonging to the halotolerant highly related Debaryomyces hansenii/fabryi species. The analysis performed consisted in studying tolerance properties, membrane characteristics, and cation incell amounts. We have specifically investigated (1) tolerance to different chemicals, (2) tolerance to osmotic and salt stress, (3) tolerance and response to oxidative stress, (4) reactive oxygen species (ROS) content, (5) relative membrane potential, (6) cell volume, (7) K(+) and Na(+) ion content, and (8) membrane fluidity. Unexpectedly, no direct relationship was found between one particular strain, Na(+) content and its tolerance to NaCl or between its ROS content and its tolerance to H(2)O(2). Results show that, although in general, human origin D. fabryi strains were more resistant to oxidative stress and presented shorter doubling times and smaller cell volume than food isolated D. hansenii ones, strains belonging to the same species can be significantly different. Debaryomyces fabryi CBS1793 strain highlighted for its extremely tolerant behavior when exposed to the diverse stress factors studied.

Disruption of HLA-DR raft, deregulations of Lck-ZAP-70-Cbl-b cross-talk and miR181a towards T cell hyporesponsiveness in leprosy.

Leprosy, a chronic human disease, results from infection of Mycobacterium leprae. Defective CMI and T cell hyporesponsiveness are the major hallmark of M. leprae pathogenesis. The present study demonstrates immunological-deregulations that eventually lead to T cell anergy/hyporesponsiveness in M. lepare infection. We firstly, evaluated the membrane fluidity and antigen-presenting-lipid-raft (HLA-DR) on macrophages of leprosy patients using fluorescence anisotropy and confocal microscopy, respectively. Increased membrane fluidity and raft-out localizations of over-expressed HLA-DR towards BL/LL pole are pinpointed as major defects, may be leading to defective antigen presentation in leprosy. Furthermore, altered expression and localization of Lck, ZAP-70, etc. and their deregulated cross talks with negative regulators (CD45, Cbl-b and SHP2) turned out to be the major putative reason(s) leading to T cell hyporesponsiveness in leprosy. Deregulations of Lck-ZAP-70 cross-talk in T cells were found to be associated with cholesterol-dependent-dismantling of HLA-DR rafts in macrophages in leprosy progression. Increased molecular interactions between Cbl-b and Lck/ZAP-70 and their subsequent degradation via ubiquitinization pathway, as result of high expression of Cbl-b, were turned out to be one of the principal underlying reason leading to T cell anergy in leprosy patients. Interestingly, overexpression of SHP2 due to gradual losses of miR181a and subsequent dephosphorylation of imperative T cell signaling molecules were emerged out as another important reason associated with prevailing T cell hyporesponsiveness during leprosy progression. Thus, this study for the first time pinpointed overexpression of Cbl-b and expressional losses of miR-181 as important hallmarks of progression of leprosy.

Plasma membrane composition of Debaryomyces hansenii adapts to changes in pH and external salinity.

Debaryomyces hansenii is a marine yeast that has to cope with different stress situations. Since changes in membrane properties can play an important function in adaptation, we have examined the fluidity and lipid composition of purified plasma membranes of D. hansenii grown at different external pH values and salt concentrations. Growth at low pH caused an increase in the sterol-to-phospholipid ratio and a decrease in fatty acid unsaturation which was reflected in decreased fluidity of the plasma membrane. High levels of NaCl increased the sterol-to-phospholipid ratio and fatty acid unsaturation, but did not significantly affect fluidity. The sterol-to-phospholipid ratios obtained in D. hansenii grown under any of these conditions were similar to the ratios that have been reported for halophilic/halotolerant black yeasts, but much smaller than those observed in the model yeast Saccharomyces cerevisiae.

The biosynthesis of cyclopropanated mycolic acids in Mycobacterium tuberculosis. Identification and functional analysis of CMAS-2.

The major mycolic acid produced by Mycobacterium tuberculosis contains two cis-cyclopropanes in the meromycolate chain. The gene whose product cyclopropanates the proximal double bond was cloned by homology to a putative cyclopropane synthase identified from the Mycobacterium leprae genome sequencing project. This gene, named cma2, was sequenced and found to be 52% identical to cma1 (which cyclopropanates the distal double bond) and 73% identical to the gene from M. leprae. Both cma genes were found to be restricted in distribution to pathogenic species of mycobacteria. Expression of cma2 in Mycobacterium smegmatis resulted in the cyclopropanation of the proximal double bond in the alpha 1 series of mycolic acids. Coexpression of both cyclopropane synthases resulted in cyclopropanation of both centers, producing a molecule structurally similar to the M. tuberculosis alpha-dicyclopropyl mycolates. Differential scanning calorimetry of purified cell walls and mycolic acids demonstrated that cyclopropanation of the proximal position raised the observed transition temperature by 3 degrees C. These results suggest that cyclopropanation contributes to the structural integrity of the cell wall complex.