Novel C-seco-taxoids possessing high potency against paclitaxel-resistant cancer cell lines overexpressing class III β-tubulin

Novel C-seco-taxoids were synthesized from 10-deacetylbaccatin III and their potencies evaluated against drug-sensitive and drug-resistant cancer cell lines. The drug-resistant cell lines include ovarian cancer cell lines resistant to cisplatin, topotecan, adriamycin and paclitaxel overexpressing class III β-tubulin, A2780TC1 and A2780TC3. The last two cell lines were selected through chronic exposure of A2780wt to paclitaxel and Pgp blocker cyclosporine. All novel C-seco-taxoids exhibited remarkable potency against A2780TC1 and A2780TC3 cell lines, and no cross resistance to cisplatin- and topotecan-resistant cell lines, A2780CIS and A2780TOP. Four of those C-seco-taxoids exhibit much higher activities than IDN5390 against paclitaxel-resistant cell lines, A2780ADR, A2780TC1 and A2780TC3. SB-CST-10202 possesses the best all-round high potencies across different drug-resistant cell lines. Molecular modeling studies, including molecular dynamics simulations, on the drug-protein complexes of class I and III β-tubulins were performed to identify possible cause of the remarkable potency of these C-seco-taxoids against paclitaxel-resistant cell lines overexpressing class III β-tubulin.     

Pichia anomala DBVPG 3003 Secretes a Ubiquitin-Like Protein That Has Antimicrobial Activity

The yeast strain Pichia anomala DBVPG 3003 secretes a killer toxin (Pikt) that has antifungal activity against Brettanomyces/Dekkera sp. yeasts. Pikt interacts with β-1,6-glucan, consistent with binding to the cell wall of sensitive targets. In contrast to that of toxin K1, secreted by Saccharomyces cerevisiae, Pikt killer activity is not mediated by an increase in membrane permeability. Purification of the toxin yielded a homogeneous protein of about 8 kDa, which showed a marked similarity to ubiquitin in terms of molecular mass and N-terminal sequences. Pikt is also specifically recognized by anti-bovine ubiquitin antibodies and, similar to ubiquitin-like peptides, is not absorbed by DEAE-cellulose. However, Pikt differs from ubiquitin in its sensitivity to proteolytic enzymes. Therefore, Pikt appears to be a novel ubiquitin-like peptide that has killer activity.     

Chronic intracerebroventricular injection of TLQP-21 prevents high fat diet induced weight gain in fast weight-gaining mice

The vgf gene regulates energy homeostasis and the VGF-derived peptide TLQP-21 centrally exerts catabolic effects in mice and hamsters. Here, we investigate the effect of chronic intracerebroventricular (icv) injection of TLQP-21 in mice fed high fat diet (HFD). Fast weight-gaining mice injected with the peptide or cerebrospinal fluid were selected for physiological, endocrine, and molecular analysis. TLQP-21 selectively inhibited the increase in body weight and epididymal white adipose tissue (eWAT) weight induced by HFD in control animals despite both groups having a similar degree of hyperphagia. TLQP-21 normalized the increase in leptin and decrease in ghrelin while increasing epinephrine and epinephrine/norepinephrine ratio when compared to values in controls. Finally, HFD-TLQP-21 mice showed a selective increase of eWAT β3-adrenergic receptor mRNA. Peroxisome-proliferator-activated-receptor-δ and hormone-sensing-lipase mRNA were also upregulated. In conclusion, chronic icv infusion of TLQP-21 prevented the early phase of diet-induced obesity despite overfeeding. These effects were paralleled by activation of catabolic pathways within the eWAT. Our results further support a role for TLQP-21 as a catabolic neuropeptide.     

IgG Autoantibody to Brain Beta Tubulin III Associated with Cytokine Cluster-II Discriminate Cerebral Malaria in Central India

Background

The main processes in the pathogenesis of cerebral malaria caused by Plasmodium falciparum involved sequestration of parasitized red blood cells and immunopathological responses. Among immune factors, IgG autoantibodies to brain antigens are increased in P. falciparum infected patients and correlate with disease severity in African children. Nevertheless, their role in the pathophysiology of cerebral malaria (CM) is not fully defined. We extended our analysis to an Indian population with genetic backgrounds and endemic and environmental status different from Africa to determine if these autoantibodies could be either a biomarker or a risk factor of developing CM.

Methods/Principal Findings

We investigated the significance of these self-reactive antibodies in clinically well-defined groups of P. falciparum infected patients manifesting mild malaria (MM), severe non-cerebral malaria (SM), or cerebral malaria (CM) and in control subjects from Gondia, a malaria epidemic site in central India using quantitative immunoprinting and multivariate statistical analyses. A two-fold complete-linkage hierarchical clustering allows classifying the different patient groups and to distinguish the CM from the others on the basis of their profile of IgG reactivity to brain proteins defined by PANAMA Blot. We identified beta tubulin III (TBB3) as a novel discriminant brain antigen in the prevalence of CM. In addition, circulating IgG from CM patients highly react with recombinant TBB3. Overall, correspondence analyses based on singular value decomposition show a strong correlation between IgG anti-TBB3 and elevated concentration of cluster-II cytokine (IFNγ, IL1β, TNFα, TGFβ) previously demonstrated to be a predictor of CM in the same population.

Conclusions/Significance

Collectively, these findings validate the relationship between antibody response to brain induced by P. falciparum infection and plasma cytokine patterns with clinical outcome of malaria. They also provide significant insight into the immune mechanisms associated to CM by the identification of TBB3 as a new disease-specific marker and potential therapeutic target.     

NMR assignments of the DNA binding domain of Ml4 protein from Mesorhizobium loti

Ml4 protein from Mesorhizobium loti has a 58% sequence identity with the Ros protein from Agrobacterium tumefaciens that contains a prokaryotic Cys₂His₂ zinc finger domain. Interestingly, Ml4 is a zinc-lacking protein that does not contain the Cys₂His₂ motif and is able to bind the Ros DNA target sequence with high affinity. Here we report the ¹H, ¹⁵N and ¹³C NMR assignments of the Ml4 protein DNA binding domain (residue 52–151), as an important step toward elucidating at a molecular level how this prokaryotic domain can overcome the metal requirement for proper folding and DNA-binding activity.     

Avoidance of protein oxidation correlates with the desiccation and radiation resistance of hot and cold desert strains of the cyanobacterium Chroococcidiopsis

To investigate the relationship between desiccation and the extent of protein oxidation in desert strains of Chroococcidiopsis a selection of 10 isolates from hot and cold deserts and the terrestrial cyanobacterium Chroococcidiopsis thermalis sp. PCC 7203 were exposed to desiccation (air-drying) and analyzed for survival. Strain CCMEE 029 from the Negev desert and the aquatic cyanobacterium Synechocystis sp. PCC 6803 were further investigated for protein oxidation after desiccation (drying over silica gel), treatment with H₂O₂ up to 1 M and exposure to γ-rays up to 25 kGy. Then a selection of desert strains of Chroococcidiopsis with different survival rates after prolonged desiccation, as well as Synechocystis sp. PCC 6803 and Chroococcidiopsis thermalis sp. PCC 7203, were analyzed for protein oxidation after treatment with 10 and 100 mM of H₂O₂. Results suggest that in the investigated strains a tight correlation occurs between desiccation and radiation tolerance and avoidance of protein oxidation.

     

Design and expression of peptides with antimicrobial activity against Salmonella typhimurium

We showed previously that insertion of Synechocystis Δ¹²‐desaturase in salmonella's membrane alters membrane physical state (MPS), followed by the expression of stress genes causing inability to survive within murine macrophages (MΦ). Recently, we showed that expression of one membrane lipid domain (MLD) of Δ¹²‐desaturase (ORF200) interferes with salmonella MPS, causing loss of virulence in mice and immunoprotection. Here, we postulate that an α‐antimicrobial peptide (α‐AMP) intercalates within membrane lipids, and depending on its amino acid sequence, it does so within specific key sensors of MLD. In this study, we choose as target for a putative synthetic AMP, PhoP/PhoQ, a sensor that responds to low Mg²⁺ concentration. We synthesised a modified DNA fragment coding for an amino acid sequence (NUF) similar to that fragment and expressed it in salmonella typhimurium. We showed that the pattern of gene expression controlled by PhoP/PhoQ highlights dysregulation of pathways involving phospholipids biosynthesis, stress proteins and genes coding for antigens. RNA‐Seq of strain expressing ORF200 showed that the pattern of those genes is also altered here. Accumulation of NUF conferred temporary immunoprotection. This represents a powerful procedure to address synthetic α‐AMPs to a specific MLD generating live non‐virulent bacterial strains.