Privileged structure based ligands for melanocortin receptors--4,4-disubstituted piperidine derivatives

Homologation and cyclization back to the chiral methine of compound 3 yields achiral 4,4-disubstituted piperidine privileged structures (e.g., 8a) useful in the construction of melanocortin 4 receptor (MC4R) ligands. The piperidine nitrogen was replaced with carbon, oxygen, sulfur, and sulfone with minor erosion of binding. The methyl cyclohexane substituent was the most potent while significant affinity was still seen for smaller lipophilic groups such as ethyl.     

Caribbean corals in crisis: record thermal stress, bleaching, and mortality in 2005

Background

The rising temperature of the world''s oceans has become a major threat to coral reefs globally as the severity and frequency of mass coral bleaching and mortality events increase. In 2005, high ocean temperatures in the tropical Atlantic and Caribbean resulted in the most severe bleaching event ever recorded in the basin.

Methodology/Principal Findings

Satellite-based tools provided warnings for coral reef managers and scientists, guiding both the timing and location of researchers'' field observations as anomalously warm conditions developed and spread across the greater Caribbean region from June to October 2005. Field surveys of bleaching and mortality exceeded prior efforts in detail and extent, and provided a new standard for documenting the effects of bleaching and for testing nowcast and forecast products. Collaborators from 22 countries undertook the most comprehensive documentation of basin-scale bleaching to date and found that over 80% of corals bleached and over 40% died at many sites. The most severe bleaching coincided with waters nearest a western Atlantic warm pool that was centered off the northern end of the Lesser Antilles.

Conclusions/Significance

Thermal stress during the 2005 event exceeded any observed from the Caribbean in the prior 20 years, and regionally-averaged temperatures were the warmest in over 150 years. Comparison of satellite data against field surveys demonstrated a significant predictive relationship between accumulated heat stress (measured using NOAA Coral Reef Watch''s Degree Heating Weeks) and bleaching intensity. This severe, widespread bleaching and mortality will undoubtedly have long-term consequences for reef ecosystems and suggests a troubled future for tropical marine ecosystems under a warming climate.     

TRIAD3/RNF216 mutations associated with Gordon Holmes syndrome lead to synaptic and cognitive impairments via Arc misregulation

Multiple loss‐of‐function mutations in TRIAD3 (a.k.a. RNF216) have recently been identified in patients suffering from Gordon Holmes syndrome (GHS), characterized by cognitive decline, dementia, and movement disorders. TRIAD3A is an E3 ubiquitin ligase that recognizes and facilitates the ubiquitination of its target for degradation by the ubiquitin‐proteasome system (UPS). Here, we demonstrate that two of these missense substitutions in TRIAD3 (R660C and R694C) could not regulate the degradation of their neuronal target, activity‐regulated cytoskeletal‐associated protein (Arc/Arg 3.1), whose expression is critical for synaptic plasticity and memory. The synaptic deficits due to the loss of endogenous TRIAD3A could not be rescued by TRIAD3A harboring GHS‐associated missense mutations. Moreover, we demonstrate that the loss of endogenous TRIAD3A in the mouse hippocampal CA1 region led to deficits in spatial learning and memory. Finally, we show that these missense mutations abolished the interaction of TRIAD3A with Arc, disrupting Arc ubiquitination, and consequently Arc degradation. Our current findings of Arc misregulation by TRIAD3A variants suggest that loss‐of‐function mutations in TRIAD3A may contribute to dementia observed in patients with GHS driven by dysfunctional UPS components, leading to cognitive impairments through the synaptic protein Arc.     

Excess of de novo deleterious mutations in genes associated with glutamatergic systems in nonsyndromic intellectual disability

Little is known about the genetics of nonsyndromic intellectual disability (NSID). We hypothesized that de novo mutations (DNMs) in synaptic genes explain an important fraction of sporadic NSID cases. In order to investigate this possibility, we sequenced 197 genes encoding glutamate receptors and a large subset of their known interacting proteins in 95 sporadic cases of NSID. We found 11 DNMs, including ten potentially deleterious mutations (three nonsense, two splicing, one frameshift, four missense) and one neutral mutation (silent) in eight different genes. Calculation of point-substitution DNM rates per functional and neutral site showed significant excess of functional DNMs compared to neutral ones. De novo truncating and/or splicing mutations in SYNGAP1, STXBP1, and SHANK3 were found in six patients and are likely to be pathogenic. De novo missense mutations were found in KIF1A, GRIN1, CACNG2, and EPB41L1. Functional studies showed that all these missense mutations affect protein function in cell culture systems, suggesting that they may be pathogenic. Sequencing these four genes in 50 additional sporadic cases of NSID identified a second DNM in GRIN1 (c.1679_1681dup/p.Ser560dup). This mutation also affects protein function, consistent with structural predictions. None of these mutations or any other DNMs were identified in these genes in 285 healthy controls. This study highlights the importance of the glutamate receptor complexes in NSID and further supports the role of DNMs in this disorder.     

Immobilization of Aspergillus oryzae β galactosidase on zinc oxide nanoparticles via simple adsorption mechanism

The present study demonstrates the immobilization of Aspergillus oryzae β galactosidase on native zinc oxide (ZnO) and zinc oxide nanoparticles (ZnO-NP) by simple adsorption mechanism. The binding of enzyme on ZnO-NP was confirmed by Fourier transform-infrared spectroscopy and atomic force microscopy. Native ZnO and ZnO-NP showed 60% and 85% immobilization yield, respectively. Soluble and immobilized enzyme preparations exhibited similar pH-optima at pH 4.5. ZnO-NP bound β galactosidase retained 73% activity at pH 7.0 while soluble and ZnO adsorbed enzyme lost 68% and 53% activity under similar experimental conditions, respectively. There was a marked broadening in temperature-activity profile for ZnO-NP adsorbed β galactosidase; it showed no difference in temperature-optima between 50 °C and 60 °C. Moreover, ZnO-NP adsorbed β galactosidase retained 53% activity after 1 h incubation with 5% galactose while the native ZnO- and soluble β galactosidase exhibited 35% and 28% activity under similar exposure, respectively. Native ZnO and ZnO-NP adsorbed β galactosidase retained 61% and 75% of the initial activity after seventh repeated use, respectively. It was noticed that 54%, 63% and 71% milk lactose was hydrolyzed by soluble, ZnO adsorbed and ZnO-NP adsorbed β galactosidase in batch process after 9 h while whey lactose was hydrolyzed to 61%, 68% and 81% under similar experimental conditions, respectively. In view of its easy production, improved stability against various denaturants and excellent reusability, ZnO-NP bound β galactosidase may find its applications in constructing enzyme-based analytical devices for clinical, environmental and food technology.     

CC and CXC chemokines induce airway smooth muscle proliferation and survival

The increase in airway smooth muscle (ASM) mass is a major structural change in asthma. This increase has been attributed to ASM cell (ASMC) hyperplasia and hypertrophy. The distance between ASMC and the epithelium is reduced, suggesting migration of smooth muscle cells toward the epithelium. Recent studies have suggested a role of chemokines in ASMC migration toward the epithelium; however, chemokines have other biological effects. The objective of the current study is to test the hypothesis that chemokines (eotaxin, RANTES, IL-8, and MIP-1α) can directly influence ASMC mass by increasing the rate of proliferation or enhancing the survival of these cells. Human ASMCs were exposed to different concentrations of eotaxin, RANTES, IL-8, or MIP-1α. To test for proliferation, matched control and stimulated ASMC were pulsed with [(3)H]thymidine, or ASMCs were stained with BrdU and then analyzed with flow cytometry. Apoptosis was measured using Annexin V staining and flow cytometry. Expression of phosphorylated p42/p44 and MAPKs was assessed by Western blot. In a concentration-dependent manner, chemokines including eotaxin, RANTES, IL-8, and MIP-1α increased ASMC's [(3)H]thymidine incorporation and DNA synthesis. IL-8, eotaxin, and MIP-1α decreased the rate of apoptosis of ASMCs compared with the matched controls. A significant increase in phosphorylated p42/p44 MAPKs was seen after treating ASMCs with RANTES and eotaxin. Moreover, inhibition of p42/p44 MAPK phosphorylation reduced the level of chemokine-induced ASM proliferation. We conclude that chemokines might contribute to airway remodeling seen in asthma by enhancing the number and survival of ASMCs.     

A novel combination immunotherapy for cancer by IL-13Rα2-targeted DNA vaccine and immunotoxin in murine tumor models

Optimum efficacy of therapeutic cancer vaccines may require combinations that generate effective antitumor immune responses, as well as overcome immune evasion and tolerance mechanisms mediated by progressing tumor. Previous studies showed that IL-13Rα2, a unique tumor-associated Ag, is a promising target for cancer immunotherapy. A targeted cytotoxin composed of IL-13 and mutated Pseudomonas exotoxin induced specific killing of IL-13Rα2(+) tumor cells. When combined with IL-13Rα2 DNA cancer vaccine, surprisingly, it mediated synergistic antitumor effects on tumor growth and metastasis in established murine breast carcinoma and sarcoma tumor models. The mechanism of synergistic activity involved direct killing of tumor cells and cell-mediated immune responses, as well as elimination of myeloid-derived suppressor cells and, consequently, regulatory T cells. These novel results provide a strong rationale for combining immunotoxins with cancer vaccines for the treatment of patients with advanced cancer.     

Association of mutation and hypermethylation of p21 gene with susceptibility to breast cancer: a study from north India

p21 gene located at chromosome 6p21.2 is a possible tumour suppressor gene involved in the pathogenesis of breast cancer. Both genetic and epigenetic alterations in p21 have been implicated in breast carcinoma. In the present study, our main aim was to study the impact of these two kinds of alterations of p21 gene in Indian female breast cancer patients. A total of 150 female breast cancer patients of north India were screened by PCR-SSCP followed by direct sequencing and methylation specific PCR. Mutational screening of p21 gene revealed significant amount of mutations [32.66 % (49/150)] in exon 2, whereas p21 promoter was found hypermethylated in 42 of 150 (28 %) breast cancer patients in our population. The intriguing feature of the study was the G>T transition (GAG>TAG) at codon 107 and the A>C transition (AGC>CGC) at codon 146 possibly rendering p21 completely ineffective in its anti- proliferative activity. Our results suggest a significant association between the mutational and hypermethylation profile of p21 gene. Therefore, we show for the first time that the significant association of p21 mutation and hypermethylation leads to the complete inactivation of p21 gene in Indian female breast cancer patients. Complete silencing of the p21 gene seems to be the result not only of genetic alterations but also of epigenetic modification.     

Integrated metagenomic and metaproteomic analyses of marine biofilm communities

Metagenomic and metaproteomic analyses were utilized to determine the composition and function of complex air–water interface biofilms sampled from the hulls of two US Navy destroyers. Prokaryotic community analyses using PhyloChip-based 16S rDNA profiling revealed two significantly different and taxonomically rich biofilm communities (6,942 taxa) in which the majority of unique taxa were ascribed to members of the Gammaproteobacteria, Alphaproteobacteria and Clostridia. Although metagenomic sequencing indicated that both biofilms were dominated by prokaryotic sequence reads (> 91%) with the majority of the bacterial reads belonging to the Alphaproteobacteria, the Ship-1 metagenome harbored greater organismal and functional diversity and was comparatively enriched for sequences from Cyanobacteria, Bacteroidetes and macroscopic eukaryotes, whereas the Ship-2 metagenome was enriched for sequences from Proteobacteria and microscopic photosynthetic eukaryotes. Qualitative liquid chromatography-tandem mass spectrometry metaproteome analyses identified 678 unique proteins, revealed little overlap in species and protein composition between the ships and contrasted with the metagenomic data in that ~80% of classified and annotated proteins were of eukaryotic origin and dominated by members of the Bacillariophyta, Cnidaria, Chordata and Arthropoda (data deposited to the ProteomeXchange, identifier PXD000961). Within the shared metaproteome, quantitative ¹⁸O and iTRAQ analyses demonstrated a significantly greater abundance of structural proteins from macroscopic eukaryotes on Ship-1 and diatom photosynthesis proteins on Ship-2. Photosynthetic pigment composition and elemental analyses confirmed that both biofilms were dominated by phototrophic processes. These data begin to provide a better understanding of the complex organismal and biomolecular composition of marine biofilms while highlighting caveats in the interpretation of stand-alone environmental ‘-omics’ datasets.