Synthesis,Antitubercular Activity and Mechanism of Resistance of Highly Effective Thiacetazone Analogues

Defining the pharmacological target(s) of currently used drugs and developing new analogues with greater potency are both important aspects of the search for agents that are effective against drug-sensitive and drug-resistant Mycobacterium tuberculosis. Thiacetazone (TAC) is an anti-tubercular drug that was formerly used in conjunction with isoniazid, but removed from the antitubercular chemotherapeutic arsenal due to toxic side effects. However, several recent studies have linked the mechanisms of action of TAC to mycolic acid metabolism and TAC-derived analogues have shown increased potency against M. tuberculosis. To obtain new insights into the molecular mechanisms of TAC resistance, we isolated and analyzed 10 mutants of M. tuberculosis that were highly resistant to TAC. One strain was found to be mutated in the methyltransferase MmaA4 at Gly101, consistent with its lack of oxygenated mycolic acids. All remaining strains harbored missense mutations in either HadA (at Cys61) or HadC (at Val85, Lys157 or Thr123), which are components of the β-hydroxyacyl-ACP dehydratase complex that participates in the mycolic acid elongation step. Separately, a library of 31 new TAC analogues was synthesized and evaluated against M. tuberculosis. Two of these compounds, 15 and 16, exhibited minimal inhibitory concentrations 10-fold lower than the parental molecule, and inhibited mycolic acid biosynthesis in a dose-dependent manner. Moreover, overexpression of HadAB HadBC or HadABC in M. tuberculosis led to high level resistance to these compounds, demonstrating that their mode of action is similar to that of TAC. In summary, this study uncovered new mutations associated with TAC resistance and also demonstrated that simple structural optimization of the TAC scaffold was possible and may lead to a new generation of TAC-derived drug candidates for the potential treatment of tuberculosis as mycolic acid inhibitors.     

Whole-Exome Sequencing Identifies Mutated C12orf57 in Recessive Corpus Callosum Hypoplasia

The corpus callosum is the principal cerebral commissure connecting the right and left hemispheres. The development of the corpus callosum is under tight genetic control, as demonstrated by abnormalities in its development in more than 1,000 genetic syndromes. We recruited more than 25 families in which members affected with corpus callosum hypoplasia (CCH) lacked syndromic features and had consanguineous parents, suggesting recessive causes. Exome sequence analysis identified C12orf57 mutations at the initiator methionine codon in four different families. C12orf57 is ubiquitously expressed and encodes a poorly annotated 126 amino acid protein of unknown function. This protein is without significant paralogs but has been tightly conserved across evolution. Our data suggest that this conserved gene is required for development of the human corpus callosum.     

Evidence for new C-terminally truncated variants of α- and β-tubulins

Cellular α-tubulin can bear various carboxy-terminal sequences: full-length tubulin arising from gene neosynthesis is tyrosinated, and two truncated variants, corresponding to detyrosinated and Δ2 α‑tubulin, result from the sequential cleavage of one or two C-terminal residues, respectively. Here, by using a novel antibody named 3EG that is highly specific to the –EEEG C-terminal sequence, we demonstrate the occurrence in neuronal tissues of a new αΔ3‑tubulin variant corresponding to α1A/B‑tubulin deleted of its last three residues (EEY). αΔ3‑tubulin has a specific distribution pattern: its quantity in the brain is similar to that of αΔ2-tubulin around birth but is much lower in adult tissue. This truncated α1A/B-tubulin variant can be generated from αΔ2-tubulin by the deglutamylases CCP1, CCP4, CCP5, and CCP6 but not by CCP2 and CCP3. Moreover, using 3EG antibody, we identify a C‑terminally truncated β-tubulin form with the same –EEEG C-terminal sequence. Using mass spectrometry, we demonstrate that β2A/B-tubulin is modified by truncation of the four C-terminal residues (EDEA). We show that this newly identified βΔ4-tubulin is ubiquitously present in cells and tissues and that its level is constant throughout the cell cycle. These new C-terminally truncated α- and β-tubulin variants, both ending with –EEEG sequence, are expected to regulate microtubule physiology. Of interest, the αΔ3-tubulin seems to be related to dynamic microtubules, resembling tyrosinated-tubulin rather than the other truncated variants, and may have critical function(s) in neuronal development.     

The histone-like protein HupB influences biofilm formation and virulence in Xanthomonas citri ssp. citri through the regulation of flagellar biosynthesis

Citrus canker is an important disease of citrus, whose causal agent is the bacterium Xanthomonas citri ssp. citri (Xcc). In previous studies, we found a group of Xcc mutants, generated by the insertion of the Tn5 transposon, which showed impaired ability to attach to an abiotic substrate. One of these mutants carries the Tn5 insertion in hupB, a gene encoding a bacterial histone-like protein, homologue to the β-subunit of the Heat-Unstable (HU) nucleoid protein of Escherichia coli. These types of protein are necessary to maintain the bacterial nucleoid organization and the global regulation of gene expression. Here, we characterized the influence of the mutation in hupB regarding Xcc biofilm formation and virulence. The mutant strain hupB was incapable of swimming in soft agar, whereas its complemented strain partially recovered this phenotype. Electron microscope imaging revealed that impaired motility of hupB was a consequence of the absence of the flagellum. Comparison of the expression of flagellar genes between the wild-type strain and hupB showed that the mutant exhibited decreased expression of fliC (encoding flagellin). The hupB mutant also displayed reduced virulence compared with the wild-type strain when they were used to infect Citrus lemon plants using different infection methods. Our results therefore show that the histone-like protein HupB plays an essential role in the pathogenesis of Xcc through the regulation of biofilm formation and biosynthesis of the flagellum.     

The appendage role of insect disco genes and possible implications on the evolution of the maggot larval form

Though initially identified as necessary for neural migration, Disconnected and its partially redundant paralog, Disco-related, are required for proper head segment identity during Drosophila embryogenesis. Here, we present evidence that these genes are also required for proper ventral appendage development during development of the adult fly, where they specify medial to distal appendage development. Cells lacking the disco genes cannot contribute to the medial and distal portions of ventral appendages. Further, ectopic disco transforms dorsal appendages toward ventral fates; in wing discs, the medial and distal leg development pathways are activated. Interestingly, this appendage role is conserved in the red flour beetle, Tribolium (where legs develop during embryogenesis), yet in the beetle we found no evidence for a head segmentation role. The lack of an embryonic head specification role in Tribolium could be interpreted as a loss of the head segmentation function in Tribolium or gain of this function during evolution of flies. However, we suggest an alternative explanation. We propose that the disco genes always function as appendage factors, but their appendage nature is masked during Drosophila embryogenesis due to the reduction of limb fields in the maggot style Drosophila larva.     

A Review of the Salivary Proteome and Peptidome and Saliva-derived Peptide Therapeutics

Saliva is a glandular secretion that is vital in the maintenance of healthy oral tissues. In this review we outline the high abundance salivary proteins, summarise the status of the salivary proteome and peptidome, the genetic origin and recognised functions of these proteins, the diseases associated with salivary disorders, and the emerging saliva-derived peptide therapeutics. Different proteomic approaches have reported the identification of over 1,300 proteins in saliva. However there are fewer than 100 high abundance proteins, identified by multiple methods including, two-dimensional polyacrylamide gel electrophoresis and HPLC combined with mass spectrometry. Analysis of the genes coding for the salivary proteins demonstrated a non-uniform chromosomal distribution with chromosome 4 having the largest proportion of genes expressed in salivary glands. Several diseases are associated with salivary disorders including Sjögren’s syndrome, Prader-Willi syndrome, dental caries and stress related disorders. Saliva as a diagnostic medium for various biochemical tests has provided a non-invasive and accessibility advantage over other more regularly tested body fluids such as blood and urine. To-date the emerging saliva-based therapeutics include artificial salivas and antimicrobial agents based on histatins and mucins.     

Livestock disturbances in Mediterranean temporary ponds: A mesocosm experiment with sheep manure and simulated trampling

相似文献   

Helicobacter pylori CagA protein targets the c-Met receptor and enhances the motogenic response

Infection with the human microbial pathogen Helicobacter pylori is assumed to lead to invasive gastric cancer. We find that H. pylori activates the hepatocyte growth factor/scatter factor receptor c-Met, which is involved in invasive growth of tumor cells. The H. pylori effector protein CagA intracellularly targets the c-Met receptor and promotes cellular processes leading to a forceful motogenic response. CagA could represent a bacterial adaptor protein that associates with phospholipase Cgamma but not Grb2-associated binder 1 or growth factor receptor-bound protein 2. The H. pylori-induced motogenic response is suppressed and blocked by the inhibition of PLCgamma and of MAPK, respectively. Thus, upon translocation, CagA modulates cellular functions by deregulating c-Met receptor signaling. The activation of the motogenic response in H. pylori-infected epithelial cells suggests that CagA could be involved in tumor progression.