Mycobacterium tuberculosis WhiB3 Responds to Vacuolar pH-induced Changes in Mycothiol Redox Potential to Modulate Phagosomal Maturation and Virulence.

The ability of Mycobacterium tuberculosis to resist intraphagosomal stresses, such as oxygen radicals and low pH, is critical for its persistence. Here, we show that a cytoplasmic redox sensor, WhiB3, and the major M. tuberculosis thiol, mycothiol (MSH), are required to resist acidic stress during infection. WhiB3 regulates the expression of genes involved in lipid anabolism, secretion, and redox metabolism, in response to acidic pH. Furthermore, inactivation of the MSH pathway subverted the expression of whiB3 along with other pH-specific genes in M. tuberculosis. Using a genetic biosensor of mycothiol redox potential (E-MSH), we demonstrated that a modest decrease in phagosomal pH is sufficient to generate redox heterogeneity in E-MSH of the M. tuberculosis population in a WhiB3-dependent manner. Data indicate that M. tuberculosis needs low pH as a signal to alter cytoplasmic E-MSH, which activates WhiB3-mediated gene expression and acid resistance. Importantly, WhiB3 regulates intraphagosomal pH by down-regulating the expression of innate immune genes and blocking phagosomal maturation. We show that this block in phagosomal maturation is in part due to WhiB3-dependent production of polyketide lipids. Consistent with these observations, Mtb Delta whiB3 displayed intramacrophage survival defect, which can be rescued by pharmacological inhibition of phagosomal acidification. Last, Mtb Delta whiB3 displayed marked attenuation in the lungs of guinea pigs. Altogether, our study revealed an intimate link between vacuolar acidification, redox physiology, and virulence in M. tuberculosis and discovered WhiB3 as crucial mediator of phagosomal maturation arrest and acid resistance in M. tuberculosis.

Experimental solubilities of two lipid derivatives in supercritical carbon dioxide and new correlations based on activity coefficient models

The solubilities of two lipid derivatives, geranyl butyrate and 10-undecen-1-ol, in SCCO2 (supercritical carbon dioxide) were measured at different operating conditions of temperature (308.15 to 333.15 K) and pressure (10 to 18 MPa). The solubilities (in mole fraction) ranged from 2.1 x 10(-3) to 23.2 x 10(-3) for geranyl butyrate and 2.2 x 10(-3) to 25.0 x 10(-3) for 10-undecen-1-ol, respectively. The solubility data showed a retrograde behavior in the pressure and temperature range investigated. Various combinations of association and solution theory along with different activity coefficient models were developed. The experimental data for the solubilities of 21 liquid solutes along with geranyl butyrate and 10-undecen-1-ol were correlated using both the newly derived models and the existing models. The average deviation of the correlation of the new models was below 15%.

Regulation of Acetate Metabolism and Acetyl Co-a Synthetase 1 (ACS1) Expression by Methanol Expression Regulator 1 (Mxr1p) in the Methylotrophic Yeast Pichia pastoris

Methanol expression regulator 1 (Mxr1p) is a zinc finger protein that regulates the expression of genes encoding enzymes of the methanol utilization pathway in the methylotrophic yeast Pichia pastoris by binding to Mxr1p response elements (MXREs) present in their promoters. Here we demonstrate that Mxr1p is a key regulator of acetate metabolism as well. Mxr1p is cytosolic in cells cultured in minimal medium containing a yeast nitrogen base, ammonium sulfate, and acetate (YNBA) but localizes to the nucleus of cells cultured in YNBA supplemented with glutamate or casamino acids as well as nutrient-rich medium containing yeast extract, peptone, and acetate (YPA). Deletion of Mxr1 retards the growth of P. pastoris cultured in YNBA supplemented with casamino acids as well as YPA. Mxr1p is a key regulator of ACS1 encoding acetyl-CoA synthetase in cells cultured in YPA. A truncated Mxr1p comprising 400 N-terminal amino acids activates ACS1 expression and enhances growth, indicating a crucial role for the N-terminal activation domain during acetate metabolism. The serine 215 residue, which is known to regulate the expression of Mxr1p-activated genes in a carbon source-dependent manner, has no role in the Mxr1p-mediated activation of ACS1 expression. The ACS1 promoter contains an Mxr1p response unit (MxRU) comprising two MXREs separated by a 30-bp spacer. Mutations that abrogate MxRU function in vivo abolish Mxr1p binding to MxRU in vitro. Mxr1p-dependent activation of ACS1 expression is most efficient in cells cultured in YPA. The fact that MXREs are conserved in genes outside of the methanol utilization pathway suggests that Mxr1p may be a key regulator of multiple metabolic pathways in P. pastoris.

Dual role of arginine metabolism in establishing pathogenesis

Arginine is an integral part of host defense when invading pathogens are encountered. The arginine metabolite nitric oxide (NO) confers antimicrobial properties, whereas the metabolite ornithine is utilized for polyamine synthesis. Polyamines are crucial to tissue repair and anti-inflammatory responses. iNOS/arginase balance can determine Th1/Th2 response. Furthermore, the host arginine pool and its metabolites are utilized as energy sources by various pathogens. Apart from its role as an immune modulator, recent studies have also highlighted the therapeutic effects of arginine. This article sheds light upon the roles of arginine metabolism during pathological conditions and its therapeutic potential.