Mitochondrial kinases in Parkinson’s disease: Converging insights from neurotoxin and genetic models

Alterations in mitochondrial biology have long been implicated in neurotoxin, and more recently, genetic models of parkinsonian neurodegeneration. In particular, kinase regulation of mitochondrial dynamics and turnover are emerging as central mechanisms at the convergence of neurotoxin, environmental and genetic approaches to studying Parkinson’s disease (PD). Kinases that localize to mitochondria during neuronal injury include mitogen activated protein kinases (MAPK) such as extracellular signal regulated protein kinases (ERK) and c-Jun N-terminal kinases (JNK), protein kinase B/Akt, and PTEN-induced kinase 1 (PINK1). Although site(s) of action within mitochondria and specific kinase targets are still unclear, these signaling pathways regulate mitochondrial respiration, transport, fission–fusion, calcium buffering, reactive oxygen species (ROS) production, mitochondrial autophagy and apoptotic cell death. In this review, we summarize accelerating experimental evidence gathered over the last decade that implicate a central role for kinase signaling at the mitochondrion in Parkinson’s and related neurodegenerative disorders. Interactions involving α-synuclein, leucine rich repeat kinase 2 (LRRK2), DJ-1 and Parkin are discussed. Converging mechanisms from different model systems support the concept of common pathways in parkinsonian neurodegeneration that may be amenable to future therapeutic interventions.     

Assessing facilitative responses to a nurse shrub at the community level: the example of Potentilla fruticosa in a sub‐alpine grassland of northwest China

Biotic interaction studies have revealed a large discrepancy among experiments in target responses to the effects of neighbours, which may in part be due to both high species‐specificity of plant responses and low number of target species used in experiments. Our aim was to assess facilitative responses at the community level and the role of both functional groups and ecological attributes of target species. In a sub‐alpine grassland on the eastern Tibet plateau, we assessed growth responses of all species in the community to removal of a dominant shrub. We also measured changes in the main environmental variables. Species responses were analysed by functional group and in relation to their mean regional altitudinal distribution. All significant interactions were positive and affected one‐third of the total species richness of the community. All functional groups were facilitated but forbs were less strongly facilitated than in the two other groups. High‐alpine species were less strongly facilitated than low‐sub‐alpine species, but the strength of this relationship was weaker than that reported in previous work. There was evidence of a decrease in extreme temperatures below the canopy of the shrub but no variations in soil moisture. We conclude that the highly stressful conditions induced by the dry continental climate of the eastern Tibet plateau are a main driver of the exclusive dominance of positive interactions. Assessing interactive responses at the community level is likely to provide a useful tool to better understand the role of biotic interactions in community responses to environmental changes.     

Peroxisome proliferator-activated receptors gamma reverses hepatic nutritional fibrosis in mice and suppresses activation of hepatic stellate cells in vitro

Nonalcoholic steatohepatitis with fibrosis is a more severe form of nonalcoholic fatty liver disease, one of the most common liver diseases. We have previously shown that peroxisome proliferator-activated receptors gamma (PPARγ) ligand, rosiglitazone, prevented the development of the methionine choline deficient (MCD) diet-induced fibrosing steatohepatitis. We have now tested whether overexpression of PPARγ ameliorates established steatohepatitis and fibrosis. Male C57BL6 mice fed with MCD diet for 8 weeks developed hepatic fibrosis with increased hepatic expression of collagen1α(I), inhibitors of fibrosis reversal-1, regulator involved in matrix degradation-9 and connective tissue growth factor. After 2 weeks of transduction of PPARγ through an adenovirus-expressing PPARγ (Ad-PPARγ), expression of these genes was reduced in a manner that paralleled the reduction in activated hepatic stellate cells (HSCs) and resolution of liver fibrosis. On the in vitro study, PPARγ is expressed in primary quiescent HSC, but depleted in culture activated HSC. Conversely, ectopic expression of PPARγ in activated HSC achieved the phenotypic reversal to the quiescent cell. Such induction markedly suppressed cell viability and cell proliferation, downregulated proliferating cell nuclear antigen, and caused cell cycle arrest at G0/G1 phase. Further, introduction of PPARγ in HSC increased cell apoptosis, this was confirmed by enhanced expression of FasL, cleaved caspase-3, cleaved caspase-7 and poly ADP-ribose polymerase, indicating an extrinsic apoptosis pathway. In conclusion, the present study shows that MCD diet-induced fibrosing steatohepatitis can be reversed by overexpression of PPARγ. It is likely that PPARγ reverses fibrosis by reducing HSCs proliferation, inducing cell cycle arrest and apoptosis.     

Generation and characterization of a Tet-On (rtTA-M2) transgenic rat

Background  

The tetracycline-inducible gene regulation system is a powerful tool that allows temporal and dose-dependent regulation of target transgene expression in vitro and in vivo. Several tetracycline-inducible transgenic mouse models have been described with ubiquitous or tissue-specific expression of tetracycline-transactivator (tTA), reverse tetracycline-transactivator (rtTA) or Tet repressor (TetR). Here we describe a Tet-On transgenic rat that ubiquitously expresses rtTA-M2 driven by the murine ROSA 26 promoter.     

Distribution and abundance of marine turtles in the Socialist Republic of Viet Nam

To establish baseline data on the distribution, abundance and threats to marine turtles in Viet Nam we conducted surveys with local fishers, community members and provincial Ministry of Fisheries staff from 17 of Viet Nam’s 29 coastal provinces. These data indicate that five species of marine turtle reside in Viet Nam’s waters (loggerhead, olive ridley, leatherback, green and hawksbill turtles), and four species nest on Viet Nam’s beaches (all of the above except the loggerhead turtle). It is evident from these data that significant declines have occurred in both foraging and nesting populations of all five marine turtle species found in Viet Nam. The greatest current threats to marine turtle populations in Viet Nam are habitat degradation, the accidental and opportunistic of turtles capture by fishers and the direct take of nesting females and their eggs. Successful conservation efforts have been made in recent years through collaboration between international Non Government Organisations and several Vietnamese Government Ministries. Continued success of these projects and the development and implementation of marine conservation policy will depend upon building awareness among Government employees, fishers and the general public about marine turtle biology, ecology, and the need to protect them.     

Synthesis and preliminary biological evaluation of the 99mTc labeled nitrobenzoimidazole and nitrotriazole as tumor hypoxia markers

1-(3-1,2,4-Nitrotriazole-1-yl)-propanhydroxyiminoamide and 1-(6-nitrobenzoimidazole-1-yl)-propanhydroxyiminoamide were synthesized and radiolabeled with (99m)Tc. The (99m)Tc labeled complexes continuously accumulated in hypoxic murine sarcoma S180 cells in vitro but not in aerobic cells. Biodistribution results in mice bearing S180 tumor indicated that the tracers could localize in tumor and eliminate from it slowly. These results suggested that the (99m)Tc labeled nitrobenzoimidazole and nitrotriazole might be the novel tumor hypoxia markers.     

Synthesis, radiolabeling, and in vivo evaluation of an 18F-labeled isatin analog for imaging caspase-3 activation in apoptosis

A non-peptide-based isatin sulfonamide analog, WC-II-89, was synthesized and its inhibition toward recombinant human caspase-3 and other caspases was determined. This compound showed high potency for inhibiting caspase-3 and -7, and high selectivity against caspases-1, -6, and -8. [(18)F]WC-II-89 was synthesized via a nucleophilic substitution of the corresponding mesylate precursor in high yield and radiochemical purity. Biodistribution studies using [(18)F]WC-II-89 revealed higher uptake in liver and spleen of cycloheximide-treated rats, an animal model of apoptosis, relative to control animals. Western blot analysis confirmed the presence of activated caspase-3 in the liver and spleen of cycloheximide-treated animals. MicroPET imaging studies revealed a high uptake of the radiotracer in the liver of a cycloheximide-treated rat relative to the untreated control. These data suggest that [(18)F]WC-II-89 is a potential radiotracer for imaging caspase-3 activation in tissues undergoing apoptosis.     

Novel phenylamino acetamide derivatives as potent and selective kappa opioid receptor agonists

A novel series of phenylamino acetamide derivatives was synthesized. These amides were shown to be potent and selective kappa opioid receptor agonists.     

Identification of novel quaternary domain interactions in the Hsp90 chaperone, GRP94

The structural basis for the coupling of ATP binding and hydrolysis to chaperone activity remains a central question in Hsp90 biology. By analogy to MutL, ATP binding to Hsp90 is thought to promote intramolecular N-terminal dimerization, yielding a molecular clamp functioning in substrate protein activation. Though observed in studies with recombinant domains, whether such quaternary states are present in native Hsp90s is unknown. In this study, native subunit interactions in GRP94, the endoplasmic reticulum Hsp90, were analyzed using chemical cross-linking in conjunction with tandem mass spectrometry. We report the identification of two distinct intermolecular interaction sites. Consistent with previous studies, one site comprises the C-terminal dimerization domain. The remaining site represents a novel intermolecular contact between the N-terminal and middle (M) domains of opposing subunits. This N+M domain interaction was present in the nucleotide-empty, ADP-, ATP-, or geldanamycin-bound states and could be selectively disrupted upon addition of synthetic geldanamycin dimers. These results identify a compact, intertwined quaternary conformation of native GRP94 and suggest that intersubunit N+M interactions are integral to the structural biology of Hsp90.