SCCRO3 (DCUN1D3) Antagonizes the Neddylation and Oncogenic Activity of SCCRO (DCUN1D1)

The activity of cullin-RING type ubiquitination E3 ligases is regulated by neddylation, a process analogous to ubiquitination that culminates in covalent attachment of the ubiquitin-like protein Nedd8 to cullins. As a component of the E3 for neddylation, SCCRO/DCUN1D1 plays a key regulatory role in neddylation and, consequently, cullin-RING ligase activity. The essential contribution of SCCRO to neddylation is to promote nuclear translocation of the cullin-ROC1 complex. The presence of a myristoyl sequence in SCCRO3, one of four SCCRO paralogues present in humans that localizes to the membrane, raises questions about its function in neddylation. We found that although SCCRO3 binds to CAND1, cullins, and ROC1, it does not efficiently bind to Ubc12, promote cullin neddylation, or conform to the reaction processivity paradigms, suggesting that SCCRO3 does not have E3 activity. Expression of SCCRO3 inhibits SCCRO-promoted neddylation by sequestering cullins to the membrane, thereby blocking its nuclear translocation. Moreover, SCCRO3 inhibits SCCRO transforming activity. The inhibitory effects of SCCRO3 on SCCRO-promoted neddylation and transformation require both an intact myristoyl sequence and PONY domain, confirming that membrane localization and binding to cullins are required for in vivo functions. Taken together, our findings suggest that SCCRO3 functions as a tumor suppressor by antagonizing the neddylation activity of SCCRO.     

A species difference between hamster and rat in the effect of oestrogens on growth of large preantral follicles

Intact or hypophysectomized 23-day-old hamsters and rats were injected s.c. with 2 mg diethylstilboestrol (DES) or 1 mg oestradiol cyclopentylpropionate (OECP) on Days 23-25 and killed on Day 26. Although serum oestradiol was elevated to the same high levels by OECP, ovarian and uterine weights were increased in the rat by OECP or DES whereas only the uterus responded in the hamster. This correlated with the ability of the oestrogens to increase significantly the number of large preantral and antral follicles in the intact rat but only the number of follicles with 2-3 layers of granulosa cells in the immature hamster. Qualitative study revealed that DES and OECP increased the number of large preantral follicles in the adult hypophysectomized rat but were ineffective in the adult hamster. It is concluded that for the immature and adult hamster oestrogens do not play a major role in the recruitment of large preantral follicles.     

The endosomal trafficking regulator LITAF controls the cardiac Nav1.5 channel via the ubiquitin ligase NEDD4-2

The QT interval is a recording of cardiac electrical activity. Previous genome-wide association studies identified genetic variants that modify the QT interval upstream of LITAF (lipopolysaccharide-induced tumor necrosis factor-α factor), a protein encoding a regulator of endosomal trafficking. However, it was not clear how LITAF might impact cardiac excitation. We investigated the effect of LITAF on the voltage-gated sodium channel Nav1.5, which is critical for cardiac depolarization. We show that overexpressed LITAF resulted in a significant increase in the density of Nav1.5-generated voltage-gated sodium current I_Na and Nav1.5 surface protein levels in rabbit cardiomyocytes and in HEK cells stably expressing Nav1.5. Proximity ligation assays showed co-localization of endogenous LITAF and Nav1.5 in cardiomyocytes, whereas co-immunoprecipitations confirmed they are in the same complex when overexpressed in HEK cells. In vitro data suggest that LITAF interacts with the ubiquitin ligase NEDD4-2, a regulator of Nav1.5. LITAF overexpression down-regulated NEDD4-2 in cardiomyocytes and HEK cells. In HEK cells, LITAF increased ubiquitination and proteasomal degradation of co-expressed NEDD4-2 and significantly blunted the negative effect of NEDD4-2 on I_Na. We conclude that LITAF controls cardiac excitability by promoting degradation of NEDD4-2, which is essential for removal of surface Nav1.5. LITAF-knockout zebrafish showed increased variation in and a nonsignificant 15% prolongation of action potential duration. Computer simulations using a rabbit-cardiomyocyte model demonstrated that changes in Ca²⁺ and Na⁺ homeostasis are responsible for the surprisingly modest action potential duration shortening. These computational data thus corroborate findings from several genome-wide association studies that associated LITAF with QT interval variation.     

Overexpression of hypoxia-inducible factor and metabolic pathways: possible targets of cancer

Cancer, the main cause of human deaths in the modern world is a group of diseases. Anticancer drug discovery is a challenge for scientists because of involvement of multiple survival pathways of cancer cells. An extensive study on the regulation of each step of these pathways may help find a potential cancer target. Up-regulated HIF-1 expression and altered metabolic pathways are two classical characteristics of cancer. Oxygen-dependent (through pVHL, PHDs, calcium-mediated) and independent (through growth factor signaling pathway, mdm2 pathway, HSP90) regulation of HIF-1α leads to angiogenesis, metastasis, and cell survival. The two subunits of HIF-1 regulates in the same fashion through different mechanisms. HIF-1α translation upregulates via mammalian target of rapamycin and mitogen-activated protein kinase signaling pathways, whereas HIF-1β through calmodulin kinase. Further, the stabilized interactions of these two subunits are important for proper functioning. Also, metabolic pathways crucial for the formation of building blocks (pentose phosphate pathway) and energy generation (glycolysis, TCA cycle and catabolism of glutamine) are altered in cancer cells to protect them from oxidative stress and to meet the reduced oxygen and nutrient supply. Up-regulated anaerobic metabolism occurs through enhanced expression of hexokinase, phosphofructokinase, triosephosphate isomerase, glucose 6-phosphate dehydrogenase and down-regulation of aerobic metabolism via pyruvate dehydrogenase kinase and lactate dehydrogenase which compensate energy requirements along with high glucose intake. Controlled expression of these two pathways through their common intermediate may serve as potent cancer target in future.     

Structure–activity relationship of dihydroxy-4-methylcoumarins as powerful antioxidants: Correlation between experimental & theoretical data and synergistic effect

The chain-breaking antioxidant activities of eight coumarins [7-hydroxy-4-methylcoumarin (1), 5,7-dihydroxy-4-methylcoumarin (2), 6,7-dihydroxy-4-methylcoumarin (3), 6,7-dihydroxycoumarin (4), 7,8-dihydroxy-4-methylcoumarin (5), ethyl 2-(7,8-dihydroxy-4-methylcoumar-3-yl)-acetate (6), 7,8-diacetoxy-4-methylcoumarin (7) and ethyl 2-(7,8-diacetoxy-4-methylcoumar-3-yl)-acetate (8)] during bulk lipid autoxidation at 37 °C and 80 °C in concentrations of 0.01–1.0 mM and their radical scavenging activities at 25 °C using TLC–DPPH test have been studied and compared. It has been found that the o-dihydroxycoumarins 3–6 demonstrated excellent activity as antioxidants and radical scavengers, much better than the m-dihydroxy analogue 2 and the monohydroxycoumarin 1. The substitution at the C-3 position did not have any effect either on the chain-breaking antioxidant activity or on the radical scavenging activity of the 7,8-dihydroxy- and 7,8-diacetoxy-4-methylcoumarins 6 and 8. The comparison with DL-α-tocopherol (TOH), caffeic acid (CA) and p-coumaric acid (p-CumA) showed that antioxidant efficiency decreases in the following sequence:     

Evolution of pollination syndromes and corolla symmetry in Balsaminaceae reconstructed using phylogenetic comparative analyses

Background and AimsFloral diversity as a result of plant–pollinator interactions can evolve by two distinct processes: shifts between pollination systems or divergent use of the same pollinator. Although both are pollinator driven, the mode, relative importance and interdependence of these different processes are rarely studied simultaneously. Here we apply a phylogenetic approach using the Balsaminaceae (including the species-rich genus Impatiens) to simultaneously quantify shifts in pollination syndromes (as inferred from the shape and colour of the perianth), as well as divergent use of the same pollinator (inferred from corolla symmetry).MethodsFor 282 species we coded pollination syndromes based on associations between floral traits and known pollination systems, and assessed corolla symmetry. The evolution of these traits was reconstructed using parsimony- and model-based approaches, using phylogenetic trees derived from phylogenetic analyses of nuclear ribosomal and plastid DNA sequence data.Key ResultsA total of 71 % of studied species have a bee pollination syndrome, 22 % a bimodal syndrome (Lepidoptera and bees), 3 % a bird pollination syndrome and 5 % a syndrome of autogamy, while 19 % of species have an asymmetrical corolla. Although floral symmetry and pollination syndromes are both evolutionarily labile, the latter shifts more frequently. Shifts in floral symmetry occurred mainly in the direction towards asymmetry, but there was considerable uncertainty in the pattern of shift direction for pollination syndrome. Shifts towards asymmetrical flowers were associated with a bee pollination syndrome.ConclusionFloral evolution in Impatiens has occurred through both pollination syndrome shifts and divergent use of the same pollinator. Although the former appears more frequent, the latter is likely to be underestimated. Shifts in floral symmetry and pollination syndromes depend on each other but also partly on the region in which these shifts take place, suggesting that the occurrence of pollinator-driven evolution may be determined by the availability of pollinator species at large geographical scales.     

Industrial potential of organic solvent tolerant bacteria

Most bacteria and their enzymes are destroyed or inactivated in the presence of organic solvents. Organic solvent tolerant bacteria are a relatively novel group of extremophilic microorganisms that combat these destructive effects and thrive in the presence of high concentrations of organic solvents as a result of various adaptations. These bacteria are being explored for their potential in industrial and environmental biotechnology, since their enzymes retain activity in the presence of toxic solvents. This property could be exploited to carry out bioremediation and biocatalysis in the presence of an organic phase. Because a large number of substrates used in industrial chemistry, such as steroids, are water-insoluble, their bioconversion rates are affected by poor dissolution in water. This problem can be overcome by carrying out the process in a biphasic organic-aqueous fermentation system, wherein the substrate is dissolved in the organic phase and provided to cells present in the aqueous phase. In bioprocessing of fine chemicals such as cis-diols and epoxides using such cultures, organic solvents can be used to extract a toxic product from the aqueous phase, thereby improving the efficiency of the process. Bacterial strains reported to grow on and utilize saturated concentrations of organic solvents such as toluene can revolutionize the removal of such pollutants. It is now known that enzymes display striking new properties in the presence of organic solvents. The role of solvent-stable enzymes in nonaqueous biocatalysis needs to be explored and could result in novel applications.     

Lactate and malignant tumors: A therapeutic target at the end stage of glycolysis

Metabolic aberrations in the form of altered flux through key metabolic pathways are primary hallmarks of many malignant tumors. Primarily the result of altered isozyme expression, these adaptations enhance the survival and proliferation of the tumor at the expense of surrounding normal tissue. Consequently, they also expose a unique set of targets for tumor destruction while sparing healthy tissues. Despite this fact, development of drugs to directly target such altered metabolic pathways of malignant tumors has been under-investigated until recently. One such target is the ultimate step of glycolysis, which, as expected, presents itself as a metabolic aberration in most malignant tumors. Termed “aerobic glycolysis” due to abnormal conversion of pyruvic acid to lactic acid even under normoxia, the altered metabolism requires these tumors to rapidly efflux lactic acid to the microenvironment in order to prevent poisoning themselves. Thus, exposed is a prime “choke-point” to target these highly malignant, frequently chemo- and radio- resistant tumors. This review will focus on current outcomes in targeting lactate efflux in such tumors using glioma as a model, an ongoing project in our laboratory for the past half-decade, as well as supporting evidence from recent studies by others on targeting this “tail-end” of glycolysis in other tumor models.     

Weak base dispiro-1,2,4-trioxolanes: potent antimalarial ozonides

Thirty weak base 1,2,4-dispiro trioxolanes (secondary ozonides) were synthesized. Amino amide trioxolanes had the best combination of antimalarial and biopharmaceutical properties. Guanidine, aminoxy, and amino acid trioxolanes had poor antimalarial activity. Lipophilic trioxolanes were less stable metabolically than their more polar counterparts.