The Cbl family proteins: ring leaders in regulation of cell signaling

The proto-oncogenic protein c-Cbl was discovered as the cellular form of v-Cbl, a retroviral transforming protein. This was followed over the years by important discoveries, which identified c-Cbl and other Cbl-family proteins as key players in several signaling pathways. c-Cbl has donned the role of a multivalent adaptor protein, capable of interacting with a plethora of proteins, and has been shown to positively influence certain biological processes. The identity of c-Cbl as an E3 ubiquitin ligase unveiled the existence of an important negative regulatory pathway involved in maintaining homeostasis in protein tyrosine kinase (PTK) signaling. Recent years have also seen the emergence of novel regulators of Cbl, which have provided further insights into the complexity of Cbl-influenced pathways. This review will endeavor to provide a summary of current studies focused on the effects of Cbl proteins on various biological processes and the mechanism of these effects. The major sections of the review are as follows: Structure and genomic organization of Cbl proteins; Phosphorylation of Cbl; Interactions of Cbl; Localization of Cbl; Mechanism of effects of Cbl: (a) Ubiquitylation-dependent events: This section elucidates the mechanism of Cbl-mediated downregulation of EGFR and details the PTK and non-PTKs targeted by Cbl. In addition, it addresses the functional requirements for E3 Ubiquitin ligase activity of Cbl and negative regulation of Cbl-mediated downregulation of PTKs, (b) Adaptor functions: This section discusses the mechanisms of adaptor functions of Cbl in mitogen-activated protein kinase (MAPK) activation, insulin signaling, regulation of Ras-related protein 1 (Rap1), PI-3' kinase signaling, and regulation of Rho-family GTPases and cytoskeleton; Biological functions: This section gives an account of the diverse biological functions of Cbl and includes the role of Cbl in transformation, T-cell signaling and thymus development, B-cell signaling, mast-cell degranulation, macrophage functions, bone development, neurite growth, platelet activation, muscle degeneration, and bacterial invasion; Conclusions and perspectives.     

Photodynamic action of C-phycocyanins obtained from marine and fresh water cyanobacterial cultures: a comparative study using EPR spin trapping technique

C-phycocyanins, major biliproteins of blue green algae (cyanobacteria), widely used as colourants in food and cosmetics are known for their antioxidant as well as therapeutic potential. Recent claims indicating phycobiliproteins exert stronger photodynamic action on tumor cells than clinically approved hematoporphyrin derivatives motivate us to investigate the photodynamic action of two newly isolated C-phycocyanins from Phormidium [PHR] and Lyngbya [LY] spp, respectively in comparison with known C-phycocyanin from Spirulina sp. [SPI]. Photolysis of air saturated solutions of PHR, LY and SPI in the presence of 2,2,6,6-Tetramethyl piperidinol (TEMPL) generated three line EPR spectrum characteristic of 4-hydroxy-2,2,6,6-tetramethylpiperidinyloxyl (TEMPOL). The increase in intensity of the EPR spectrum with time of irradiation and decrease in intensity, in the presence of 1O2 quencher DABCO confirm the formation of 1O2. Photoirradiation in the presence of spin trap 5,5-dimethyl-1-pyrroline-N-oxide (DMPO) generated EPR signal characteristic of O2(-) adduct. Efficiency of 1O2 generation is of the order LY > PHR> SPI. The yield of reactive oxygen species (ROS) generation is found to be 1O2>O2(-) indicating type II mechanism to be the prominent pathway for photosensitation by phycocyanins.     

Coupling ligand structure to specific conformational switches in the beta2-adrenoceptor

G protein-coupled receptors (GPCRs) regulate a wide variety of physiological functions in response to structurally diverse ligands ranging from cations and small organic molecules to peptides and glycoproteins. For many GPCRs, structurally related ligands can have diverse efficacy profiles. To investigate the process of ligand binding and activation, we used fluorescence spectroscopy to study the ability of ligands having different efficacies to induce a specific conformational change in the human beta2-adrenoceptor (beta2-AR). The 'ionic lock' is a molecular switch found in rhodopsin-family GPCRs that has been proposed to link the cytoplasmic ends of transmembrane domains 3 and 6 in the inactive state. We found that most partial agonists were as effective as full agonists in disrupting the ionic lock. Our results show that disruption of this important molecular switch is necessary, but not sufficient, for full activation of the beta2-AR.     

Prevention of selenite induced oxidative stress and cataractogenesis by luteolin isolated from Vitex negundo

Free radical mediated oxidative stress plays a crucial role in the pathogenesis of cataract and the present study was to determine the efficacy of luteolin in preventing selenite induced oxidative stress and cataractogenesis in vitro. Luteolin is a bioactive flavonoid, isolated and characterized from the leaves of Vitex negundo. Lenses were extracted from Sprague-Dawley strain rats and were organ cultured in DMEM medium. They were divided into three groups with eight lenses in each group as follows: lenses cultured in normal medium (G I), supplemented with 0.1mM sodium selenite (G II) and sodium selenite and 2 μg/ml luteolin (G III). Treatment was from the second to fifth day, while selenite administration was done on the third day. After the experimental period, lenses were taken out and various parameters were studied. The antioxidant potential of luteolin was assessed by 2,2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity. In the selenite induced group, morphological examination of the lenses showed dense cortical opacification and vacuolization. Biochemical examinations revealed a significant decrease in activities of antioxidant enzymes and enzymes of the glutathione system. Additionally decreased glutathione level and increased reactive oxygen species (ROS) and thiobarbituric acid reactive substances (TBARS) were observed. Luteolin treatment abated selenite induced oxidative stress and cataractogenesis by maintaining antioxidant status, reducing ROS generation and lipid peroxidation in the lens. These finding demonstrated the anticataractogenic effect of luteolin by virtue of its antioxidant property, which has been reported in this paper for the first time.     

Prolactin inhibits activity of pyruvate kinase M2 to stimulate cell proliferation

Mitogenic and prosurvival effects underlie the tumorigenic roles of prolactin (PRL) in the pathogenesis of breast cancer. PRL signaling is mediated through its receptor (PRLr). A proteomics screen identified the pyruvate kinase M2 (PKM2), a glycolytic enzyme known to play an important role in tumorigenesis, as a protein that constitutively interacts with PRLr. Treatment of cells with PRL inhibited pyruvate kinase activity and increased the lactate content in human cells in a manner that was dependent on the abundance of PRLr, activation of Janus kinase 2, and tyrosine phosphorylation of the intracellular domain of PRLr. Knockdown of PKM2 attenuated PRL-stimulated cell proliferation. The extent of this proliferation was rescued by the knock-in of the wild-type PKM2 but not of its mutant insensitive to PRL-mediated inhibition. We discuss a hypothesis that the inhibition of PKM2 by PRL contributes to the PRL-stimulated cell proliferation.     

Hymenobacter properus sp. nov., Hymenobacter ruricola sp. nov., and Hymenobacter jeongseonensis sp. nov., three new species isolated from mountain and beach soil in South Korea

Antonie van Leeuwenhoek - Three Gram-stain-negative, aerobic, circular, convex, red-colored and rod-shaped bacterial strains, designated BT439T, BT662T and BT683T were obtained from soil collected...     

Antioxidant effects of Emblica officinalis and Zingiber officinalis on arsenic and lead induced toxicity on Albino rats

It is of interest to document the effect of Emblica officinalis (E. officinalis) and Zingiber officinalae (Z. officinalae) leaf extract on reactive oxygen species, antioxidant potential changes in arsenic and lead-induced toxicity in male rats. We used 8 groups of adult male Wistar rats with 1 control group for this study. The animals were divided into Group I: Control and Group II: Lead and sodium arsenite induced rats (animals were induced for metal toxicity by the combined administration of arsenic (13.8 mg/ kg body weight) and lead (116.4 mg/kg body weight). These doses were administered by gastric intubation during 14 consecutive days using known standard procedures. Arsenic and lead induced rats treated with ethanolic extract of Emblica officinalis (60 mg/kg body weight/day, orally for 45 days) are group III rats. Group IV animals are arsenic and lead induced rats treated orally with ethanolic extracts of E. officinalis (120 mg/kg body weight/day for 45 days). Group V animals are arsenic and lead induced rats treated orally with ethanolic extracts of Z. officinalae (60 mg/kg body weight/day for 45 days). Group VI animals are arsenic and lead induced rats orally treated with ethanolic extracts of Zingiber officinalis (120 mg/kg body weight/day for 45 days). Group VII animals are arsenic and lead induced rats treated orally with ethanolic extracts of E. officinalis and Z. officinalae (60 + 60 mg/kg body weight/day for 45 days). Group VIII animals are arsenic and lead induced rats treated orally with ethanolic extracts of E. officinalis and Z. officinalae (120 + 120 mg/kg body weight/day, orally for 45 days). Normal Control animals were treated orally with ethanolic extracts of E. officinalis (120mg/kg body weight) + Z. officinalae (120mg/kg body weight) for 45 days. The control and experimental animals were then subjected to analysis for oxidative stress markers such as H2O2, *OH, and lipid peroxidation (LPO), antioxidant enzymes in addition to liver and kidney function markers. Results: Arsenic and lead induced rats showed a significant increase in the levels of reactive oxygen species (H2O2, OH* and LPO) with concomitant alterations in the renal and liver tissues. However, enzymic and non-enzymic antioxidant levels were decreased. Nevertheless, an oral effective dose of E. officinalis and Z. officinalae (120 + 120 mg/kg body weight/day increased the antioxidant enzymes and retrieved the altered levels of ROS and LPO that were induced by arsenic and lead. Thus, we show that E. officinalis and Z. officinalae leaf extract exhibits nephroprotective and hepatoprotective role through the restoration of reactive oxygen species and antioxidant enzymes in the kidney and liver tissue of Arsenic and Lead-induced nephrotoxicity and hepatotoxicity in rats. Hence, E. officinalis and Z. officinalae leaf extract are potential therapeutic options for the treatment of metal toxicity-induced kidney and liver diseases.     

Mutational analysis of G-protein coupled receptor – FFA2

FFA2 (GPR43) is a receptor for short-chain fatty acids (SCFAs), acetate, and propionate. FFA2 is predominantly expressed in islets, a subset of immune cells, adipocytes, and the gastrointestinal tract which suggest a possible role in inflammatory and metabolic conditions. We have previously described the identification and characterization of novel phenylacetamides as allosteric agonists of FFA2. In the current study, we have investigated the molecular determinants contributing to receptor activation with the endogenous and synthetic ligands as well as allosteric interactions between these two sites. The mutational analysis revealed previously unidentified sites that may allosterically regulate orthosteric ligand’s function as well as residues potentially important for the interactions between orthosteric and allosteric binding sites.