Molecular Chaperone Down-regulator Research Articles

De novo transcriptomic sequencing unraveled the molecular mechanisms of VvMybA1 underlying the alteration of Ficus lyrata leaf color

VvMybA1 belongs to the MYB gene family. It has been shown that overexpression of VvMybA1 in Ficus lyrata results in accumulation of anthocyanin in leaves along with some side effects, but the underlying mechanisms remain unclear. In the present study, transcriptome of purple-leaved F. lyrata overexpressing VvMybA1 was sequenced and compared with non-modified green-leaved F. lyrata. The results revealed that the accumulation of anthocyanin was caused by the upregulation of the UDP glucose:flavonoid glucosyltrasferase (3GT), UDP rhamnose:anthocyanidin-3-glucoside rhamnosyltransferase (3RT), chalcone synthase (CHS) and chalcone isomerase (CHI), which are key genes for anthocyanin biosynthesis. The genes associated with photosynthesis and energy supply were downregulated, probably resulting in thin and wrinkled leaves. Genes involved in defoliation, including beta-glucosidase, shikimate O-hydroxy cinnamoyl transferase (HCT) and anthocyanidin reductase (ANR) were all downregulated, which might be the reason for the easy defoliation of purple-leaved F. lyrata. Endoglucanase, flavonoids and HCT are important to pathogen resistance. Downregulation of these genes in the present study might explain the high susceptibility of purple-leaved F. lyrata to pathogens. In addition, the downregulation of molecular chaperones might also deteriorate its resistance.

Synergistic Lethality of a Binary Inhibitor of Mycobacterium tuberculosis KasA.

We report GSK3011724A (DG167) as a binary inhibitor of β-ketoacyl-ACP synthase (KasA) in Mycobacterium tuberculosis Genetic and biochemical studies established KasA as the primary target. The X-ray crystal structure of the KasA-DG167 complex refined to 2.0-Å resolution revealed two interacting DG167 molecules occupying nonidentical sites in the substrate-binding channel of KasA. The binding affinities of KasA to DG167 and its analog, 5g, which binds only once in the substrate-binding channel, were determined, along with the KasA-5g X-ray crystal structure. DG167 strongly augmented the in vitro activity of isoniazid (INH), leading to synergistic lethality, and also synergized in an acute mouse model of M. tuberculosis infection. Synergistic lethality correlated with a unique transcriptional signature, including upregulation of oxidoreductases and downregulation of molecular chaperones. The lead structure-activity relationships (SAR), pharmacokinetic profile, and detailed interactions with the KasA protein that we describe may be applied to evolve a next-generation therapeutic strategy for tuberculosis (TB).IMPORTANCE Cell wall biosynthesis inhibitors have proven highly effective for treating tuberculosis (TB). We discovered and validated members of the indazole sulfonamide class of small molecules as inhibitors of Mycobacterium tuberculosis KasA-a key component for biosynthesis of the mycolic acid layer of the bacterium's cell wall and the same pathway as that inhibited by the first-line antitubercular drug isoniazid (INH). One lead compound, DG167, demonstrated synergistic lethality in combination with INH and a transcriptional pattern consistent with bactericidality and loss of persisters. Our results also detail a novel dual-binding mechanism for this compound as well as substantial structure-activity relationships (SAR) that may help in lead optimization activities. Together, these results suggest that KasA inhibition, specifically, that shown by the DG167 series, may be developed into a potent therapy that can synergize with existing antituberculars.

Functional genomic analysis of corals from natural CO2 -seeps reveals core molecular responses involved in acclimatization to ocean acidification.

Little is known about the potential for acclimatization or adaptation of corals to ocean acidification and even less about the molecular mechanisms underpinning these processes. Here, we examine global gene expression patterns in corals and their intracellular algal symbionts from two replicate population pairs in Papua New Guinea that have undergone long-term acclimatization to natural variation in pCO2 . In the coral host, only 61 genes were differentially expressed in response to pCO2 environment, but the pattern of change was highly consistent between replicate populations, likely reflecting the core expression homeostasis response to ocean acidification. Functional annotations highlight lipid metabolism and a change in the stress response capacity of corals as key parts of this process. Specifically, constitutive downregulation of molecular chaperones was observed, which may impact response to combined climate change-related stressors. Elevated CO2 has been hypothesized to benefit photosynthetic organisms but expression changes of in hospite Symbiodinium in response to acidification were greater and less consistent among reef populations. This population-specific response suggests hosts may need to adapt not only to an acidified environment, but also to changes in their Symbiodinium populations that may not be consistent among environments, adding another challenging dimension to the physiological process of coping with climate change.

Physiological Response of Escherichia coli O157:H7 Sakai to Dynamic Changes in Temperature and Water Activity as Experienced during Carcass Chilling

Enterohemeorrhagic Escherichia coli is a leading cause of foodborne illness, with the majority of cases linked to foods of bovine origin. Currently, no completely effective method for controlling this pathogen during carcass processing exists. Understanding how this pathogen behaves under those stress conditions experienced on the carcass during chilling in cold air could offer opportunities for development or improvement of effective decontamination processes. Therefore, we studied the growth kinetics and physiological response of exponential phase E. coli O157:H7 Sakai cultures upon an abrupt downshift in temperature and water activity (from 35 °C aw 0.993 to 14 °C aw 0.967). A parallel Biolog study was conducted to follow the phenotypic responses to 190 carbon sources. Exposure of E. coli to combined cold and water activity stresses resulted in a complex pattern of population changes. This pattern could be divided into two main phases, including adaptation and regrowth phases, based on growth kinetics and clustering analyses. The transcriptomic and proteomic studies revealed that E. coli exhibited a “window” of cell susceptibility (i.e. weaknesses) during adaptation phase. This included apparent DNA damage, the downregulation of molecular chaperones and proteins associated with responses to oxidative damage. However, E. coli also displayed a transient induction in the RpoE-controlled envelope stress response and activation of the master stress regulator RpoS and the Rcs phosphorelay system involved in colanic acid biosynthesis. Increased expression was observed for several genes and/or proteins involved in DNA repair, protein and peptide degradation, amino acid biosynthesis, and carbohydrate catabolism and energy generation. Furthermore, the Biolog study revealed reduced carbon source utilization during adaptation phase, indicating the disruption of energy-generating processes. This study provides insight into the physiological response of E. coli during exposure to combined cold and water activity stress, which could be exploited to enhance the microbiological safety of carcasses and related foods.

Synthetic studies of the spirocyclic cyclohexene part of versipelostatin, a novel GRP78/Bip molecular chaperone downregulator

The spirocyclic part consisting of an α-acylated tetronic acid and a multisubstituted cyclohexene embedded in versipelostatin, a novel GRP78/Bip molecular chaperone downregulator, has been synthesized in enantiomerically pure form. The asymmetric synthesis of the targeted spiro[4.5]-1-oxa-7-decen-2,4-dione derivative was characterized by (1) stereoselective allylation at the α-carbon of methylmalonate diester, in which one carboxylic acid was esterified with a D-glucose-derived chiral template, (2) construction of the tetrasubstituted cyclohexenone substructure by high-yielding ring-closing metathesis and (3) stereoselective construction of the spirocyclic tetronic acid part starting from the cyclohexenone obtained as the ring-closing metathesis product.

New glycosylated derivatives of versipelostatin, the GRP78/Bip molecular chaperone down-regulator, from Streptomyces versipellis 4083-SVS6

Four novel glycosylated derivatives of versipelostatin (1), versipelostatins B-E (2-5), were isolated from the culture broth of Streptomyces versipellis 4083-SVS6. The inhibitory activities of the isolated compounds against the expression of molecular chaperone GRP78 induced by 2-deoxyglucose were evaluated. Of the five versipelostatin family members, 1 and 4 were the more potent with IC(50) values of 3.5 and 4.3 microM. These results suggest that the alpha-L-oleandropyranosyl (1-->4)-beta-D-digitoxopyranosyl residue in the sugar moiety may play an important role in down-regulating GRP78 expression induced by 2-deoxyglucose.

Total Synthesis of the Trisaccharide Unit of the Molecular Chaperone Down-Regulator Versipelostatin

The first synthesis of the trisaccharide unit of versipelostatin, a down regulator of grp78, was achieved, utilizing methyl­ triflate promoted glycosidations of thioethyl glycosyl donors of d-digitoxose and d-cymarose with protected digitoxal. The synthesis was terminated by the preparation of a thioglycoside based on diethyldithionophosphonic acid which is ready for further glycosidations with aglycons. Comparison of NMR spectroscopic data of two trisaccharides with the data reported for versipelostatin supported the configuration of the monosaccharide units as well as glycosidic linkages.

Prunustatin A, a Novel GRP78 Molecular Chaperone Down-regulator Isolated from Streptomyces violaceoniger

In the course of our screening program for regulators of a molecular chaperone GRP78 expression, we isolated a novel inhibitor of GRP78 expression, designated as prunustatin A, from Streptomyces violaceoniger 4521-SVS3. The structure of prunustatin A was determined by a series of NMR analyses to be an oxidized type of the neoantimycin family. Prunustatin A inhibited the expression of GRP78 induced by 2-deoxyglucose in human fibrosarcoma HT1080 cells accompanied by global cell death without showing cytotoxicity under normal nutrient condition.

Versipelostatin, a novel GRP78/Bip molecular chaperone down-regulator of microbial origin

We discovered a novel down-regulator of grp78 gene expression designated as versipelostatin from the culture broth of Streptomyces versipellis 4083-SVS6. Its structure was determined to be a macrocyclic compound consisting of an α-acyltetronic acid and sugar moieties.