Rather rule than exception? How to evaluate the relevance of dual protein targeting to mitochondria and chloroplasts

Dual targeting of a nuclearly encoded protein into two different cell organelles is an exceptional event in eukaryotic cells. Yet, the frequency of such dual targeting is remarkably high in case of mitochondria and chloroplasts, the two endosymbiotic organelles of plant cells. In most instances, it is mediated by “ambiguous” transit peptides, which recognize both organelles as the target. A number of different approaches including in silico, in organello as well as both transient and stable in vivo assays are established to determine the targeting specificity of such transit peptides. In this review, we will describe and compare these approaches and discuss the potential role of this unusual targeting process. Furthermore, we will present a hypothetical scenario how dual targeting might have arisen during evolution.     

The two Drosophila cytochrome C proteins can function in both respiration and caspase activation

Cytochrome C has two apparently separable cellular functions: respiration and caspase activation during apoptosis. While a role of the mitochondria and cytochrome C in the assembly of the apoptosome and caspase activation has been established for mammalian cells, the existence of a comparable function for cytochrome C in invertebrates remains controversial. Drosophila possesses two cytochrome c genes, cyt-c-d and cyt-c-p. We show that only cyt-c-d is required for caspase activation in an apoptosis-like process during spermatid differentiation, whereas cyt-c-p is required for respiration in the soma. However, both cytochrome C proteins can function interchangeably in respiration and caspase activation, and the difference in their genetic requirements can be attributed to differential expression in the soma and testes. Furthermore, orthologues of the apoptosome components, Ark (Apaf-1) and Dronc (caspase-9), are also required for the proper removal of bulk cytoplasm during spermatogenesis. Finally, several mutants that block caspase activation during spermatogenesis were isolated in a genetic screen, including mutants with defects in spermatid mitochondrial organization. These observations establish a role for the mitochondria in caspase activation during spermatogenesis.     

Plant Growth-Promoting Rhizobacteria Enhance Abiotic Stress Tolerance in Solanum tuberosum Through Inducing Changes in the Expression of ROS-Scavenging Enzymes and Improved Photosynthetic Performance

In this report we address the changes in the expression of the genes involved in ROS scavenging and ethylene biosynthesis induced by the inoculation of plant growth-promoting rhizobacteria (PGPR) isolated from potato rhizosphere. The two Bacillus isolates used in this investigation had earlier demonstrated a striking influence on potato tuberization. These isolates showed enhanced 1-aminocyclopropane-1-carboxylic acid deaminase activity, phosphate solubilization, and siderophore production. Potato plants inoculated with these PGPR isolates were subjected to salt, drought, and heavy-metal stresses. The enhanced mRNA expression levels of the various ROS-scavenging enzymes and higher proline content in tubers induced by PGPR-treated plants contributed to increased plant tolerance to these abiotic stresses. Furthermore, the photosynthetic performance indices of PGPR-inoculated plants clearly exhibited a positive influence of these bacterial strains on the PSII photochemistry of the plants. Overall, these results suggest that the PGPR isolates used in this study are able to confer abiotic stress tolerance in potato plants.     

Efficient Single Muscle Fiber Isolation from Alcohol-Fixed Adult Muscle following ��-Galactosidase Staining for Satellite Cell Detection

Staining for β-galactosidase activity for whole tissues, sections, and cells is a common method to detect expression of β-galactosidase reporter transgene as well as senescence-dependent β-galactosidase activity. Choice of fixatives is a critical step for detection of β-galactosidase activity, subsequent immunostaining, and enzymatic digestion of tissue to dissociate cells. In this report, the authors examined several aldehyde and alcohol fixatives in mouse skeletal muscle tissues for their efficiency at improving detection of β-galactosidase activity as well as detection by immunostaining. In addition, fixatives were also analyzed for their efficiency for collagenase digestion to isolate single muscle fibers on postfixed β-galactosidase-stained whole skeletal muscle tissues. The results show that fixing cells with isopropanol yields the greatest reliability and intensity in both β-galactosidase staining as well as double staining for β-galactosidase activity and antibodies. In addition, isopropanol and ethanol, but not glutaraldehyde or paraformaldehyde, allow for the isolation of single muscle fibers from the diaphragm and tibialis anterior muscles following postfixed β-galactosidase staining. Using this method, it is possible to identify the amount of cells that occupy the satellite cell compartment in single muscle fibers prepared from any muscle tissues, including tibialis anterior muscle and diaphragm.     

Conformational variability of different sulfonamide inhibitors with thienyl-acetamido moieties attributes to differential binding in the active site of cytosolic human carbonic anhydrase isoforms

The X-ray crystal structures of the adducts of human carbonic anhydrase (hCA, EC 4.2.1.1) II complexed with two aromatic sulfonamides incorporating 2-thienylacetamido moieties are reported here. Although, the two inhibitors only differ by the presence of an additional 3-fluoro substituent on the 4-amino-benzenesulfonamide scaffold, their inhibition profiles against the cytosolic isoforms hCA I, II, III, VII and XIII are quite different. These differences were rationalized based on the obtained X-ray crystal structures, and their comparison with other sulfonamide CA inhibitors with clinical applications, such as acetazolamide, methazolamide and dichlorophenamide. The conformations of the 2-thienylacetamido tails in the hCA II adducts of the two sulfonamides were highly different, although the benzenesulfonamide parts were superimposable. Specific interactions between structurally different inhibitors and amino acid residues present only in some considered isoforms have thus been evidenced. These findings can explain the high affinity of the 2-thienylacetamido benzenesulfonamides for some pharmacologically relevant CAs (i.e., isoforms II and VII) being also useful to design high affinity, more selective sulfonamide inhibitors of various CAs.     

A role for phasic dopamine neuron firing in habit learning

In this issue of Neuron, Wang et?al. (2011) show that mice with dopamine neuron-specific NMDAR1 deletion have attenuated phasic dopamine neuron firing and a deficit in habit learning. These findings indicate that brain regions sensitive to phasic dopamine signals may underlie habit learning.     

Noisy galvanic vestibular stimulation promotes GABA release in the substantia nigra and improves locomotion in hemiparkinsonian rats

Background

The vestibular system is connected to spinal, cerebellar and cerebral motor control structures and can be selectively activated with external electrodes. The resulting sensation of disturbed balance can be avoided by using stochastic stimulation patterns. Adding noise to the nervous system sometimes improves function. Small clinical trials suggest that stochastic vestibular stimulation (SVS) may improve symptoms in Parkinson''s disease. We have investigated this claim and possible mechanisms using the 6-hydroxydopamine (6-OHDA) hemilesion model of Parkinson''s disease.

Methodology/Principal Findings

Animals were tested in the accelerating rod test and the Montoya staircase test of skilled forelimb use. In 6-OHDA hemilesioned animals, SVS improved rod performance by 56±11 s. At group level L-DOPA treatment had no effect, but positive responders improved time on rod by 60±19 s. Skilled forelimb use was not altered by SVS. To investigate how SVS may influence basal ganglia network activity, intracerebral microdialysis was employed in four regions of interest during and after SVS. In presence of the γ-amino buturic acid (GABA) transporter inhibitor NNC 711, SVS induced an increase in GABA to 150±15% of baseline in the substantia nigra (SN) of unlesioned animals, but had no effect in the pedunculopontine nucleus (PPN), the striatum or the ventromedial thalamus (VM). Dopamine release remained stable in all areas, as did GABA and amine concentrations in the SN of unstimulated controls. Following SVS, a sustained increase in GABA concentrations was observed in the ipsilesional, but not in the contralesional SN of 6-OHDA hemilesioned rats. In contrast, L-DOPA treatment produced a similar increase of GABA in the ipsi- and contra-lesional SN.

Conclusions/Significance

SVS improves rod performance in a rat model of Parkinson''s disease, possibly by increasing nigral GABA release in a dopamine independent way. We propose that SVS could be useful for treating symptoms of Parkinson''s disease.     

Role of Vital Trace Elements in Nanocurcumin-Centered Formulation: A Novel Approach to Resuscitate the Immune System

Electromagnetic fields (EMFs) can affect living cells due to biochemical changes, followed by changes in levels of trace elements in serum and different organs. This study focuses on the effect of whole body exposure to EMF, presented everywhere in our environment, and on the levels of trace elements in serum, femur, brain, kidney, and liver tissues. The analyses performed on 29 guinea pigs were divided into five groups. Guinea pigs were exposed to a magnetic field of 50 Hz of 1.5 mT. Groups A and B were exposed to the magnetic field for a period of 4 h/day continuously (4 h/day) for 4 and 7 days, respectively. Groups C and D were exposed to the magnetic field for a period of 4 h/day intermittently for 4 and 7 days, respectively. Group E animals were enrolled as control. Copper (Cu), zinc (Zn), calcium (Ca), and magnesium (Mg) levels were determined by atomic absorption spectroscopy in serum, femur, brain, kidney, and liver tissues in all guinea pigs. When compared to the control groups, the changes in the levels of Cu in serum samples, femur, and kidney tissues of the treated groups were statistically significant. The same was also true for the levels of Mg in the brain, kidney, and lung tissues. Our results suggest that in vivo continuous and intermittent exposure to EMF may cause disturbances in homeostasis of bioelements. These effects could be important risk factors for toxic effects of EMF, especially in relation to deterioration of bioelements.