Molecular adaptations underlying susceptibility and resistance to social defeat in brain reward regions

While stressful life events are an important cause of psychopathology, most individuals exposed to adversity maintain normal psychological functioning. The molecular mechanisms underlying such resilience are poorly understood. Here, we demonstrate that an inbred population of mice subjected to social defeat can be separated into susceptible and unsusceptible subpopulations that differ along several behavioral and physiological domains. By a combination of molecular and electrophysiological techniques, we identify signature adaptations within the mesolimbic dopamine circuit that are uniquely associated with vulnerability or insusceptibility. We show that molecular recapitulations of three prototypical adaptations associated with the unsusceptible phenotype are each sufficient to promote resistant behavior. Our results validate a multidisciplinary approach to examine the neurobiological mechanisms of variations in stress resistance, and illustrate the importance of plasticity within the brain's reward circuits in actively maintaining an emotional homeostasis.     

Sphingosine kinase 1 is a critical component of the copper-dependent FGF1 export pathway

Sphingosine kinase 1 catalyzes the formation of sphingosine-1-phosphate, a lipid mediator involved in the regulation of angiogenesis. Sphingosine kinase 1 is constitutively released from cells, even though it lacks a classical signal peptide sequence. Because copper-dependent non-classical stress-induced release of FGF1 also regulates angiogenesis, we questioned whether sphingosine kinase 1 is involved in the FGF1 release pathway. We report that (i) the coexpression of sphingosine kinase 1 with FGF1 inhibited the release of sphingosine kinase 1 at 37 degrees C; (ii) sphingosine kinase 1 was released at 42 degrees C in complex with FGF1; (iii) sphingosine kinase 1 null cells failed to release FGF1 at stress; (iv) sphingosine kinase 1 is a high affinity copper-binding protein which formed a complex with FGF1 in a cell-free system, and (v) sphingosine kinase 1 over expression rescued the release of FGF1 from inhibition by the copper chelator, tetrathiomolybdate. We propose that sphingosine kinase 1 is a component of the copper-dependent FGF1 release pathway.     

Regulation of axon guidance by slit and netrin signaling in the Drosophila ventral nerve cord

Netrin and Slit signaling systems play opposing roles during the positioning of longitudinal tracts along the midline in the ventral nerve cord of Drosophila embryo. It has been hypothesized that a gradient of Slit from the midline interacts with three different Robo receptors to specify the axon tract positioning. However, no such gradient has been detected. Moreover, overexpression of Slit at the midline has no effect on the positioning of these lateral tracts. In this article, we show that Slit is present outside of the midline along the longitudinal and commissural tracts. Sli from the midline, in a Robo-independent manner, is initially taken up by the commissural axon tracts when they cross the midline and is transported along the commissural tracts into the longitudinal connectives. These results are not consistent with a Sli gradient model. We also find that sli mRNA is maternally deposited and embryos that are genetically null for sli can have weaker guidance defects. Moreover, in robo or robo3 mutants, embryos with normal axon tracts are found and such robo embryos reach pupal stages and die, while robo3 mutant embryos develop into normal individuals and produce eggs. Interestingly, embryos from robo3 homozygous individuals fail to develop but have axon tracts ranging from normal to various defects: robo3 phenotype, robo phenotype, and slit-like phenotype, suggesting a more complex functional role for these genes than what has been proposed. Finally, our previous results indicated that netrin phenotype is epistatic to sli or robo phenotypes. However, it seems likely that this previously reported epistatic relationship might be due to the partial penetrance of the sli, robo, robo3 (or robo2) phenotypes. Our results argue that double mutant epistasis is most definitive only if the penetrance of the phenotypes of the mutants involved is complete.     

A role for natural killer cells in intestinal inflammation caused by infection with Salmonella enterica serovar Typhimurium

Acute gastroenteritis caused by Salmonella infection is a significant public health problem. Using a mouse model of this condition, the authors demonstrated previously that the cytokine gamma interferon (IFN-gamma) is required for a normal intestinal inflammatory response to the pathogen. In the present study, these experiments are extended to show that natural killer (NK) cells constitute an early source of intestinal IFN-gamma during Salmonella infection, and that these cells have a significant impact on intestinal inflammation. It was found that infection of mice with Salmonella increased both intestinal IFN-gamma production and the numbers of NK cells in the intestine and mesenteric lymph nodes. NK cells, along with other types of lymphocytes, produced IFN-gamma in response to the bacteria in vitro, while antibody-mediated depletion of NK cells in vivo resulted in a significant reduction in Salmonella-induced intestinal IFN-gamma expression. In a mouse strain lacking NK cells and T and B lymphocytes, intestinal production of IFN-gamma and Salmonella-induced intestinal inflammation were both significantly decreased compared with a strain deficient only in T and B cells. The authors' observations point to an important function for NK cells and NK-derived IFN-gamma in regulating the intestinal inflammatory response to Salmonella.     

Troglitazone activates TRPV1 and causes deacetylation of PPARγ in 3T3-L1 cells

Published research suggests that activation of transient receptor potential vanilloid subfamily 1 (TRPV1) enhances the expression and deacetylation of peroxisome proliferator-activated receptor gamma (PPARγ) to cause browning of white adipose tissue. Here, we show that TRPV1 activation by capsaicin significantly prevents high fat diet-induced obesity in mice. This is associated with an increase in the expression and deacetylation of PPARγ in the epididymal fat of these mice. Consistent with the TRPV1 activation in vivo, overexpression of TRPV1 enhanced the PPARγ and other thermogenic genes in cultured 3T3-L1 preadipocytes. To determine the interaction between TRPV1 and PPARγ signaling, we analyzed the effect of Troglitazone (Trog; a thiazolidinedione derivative and an agonist of PAARγ) treatment on cultured 3T3-L1 cells. Trog enhanced the expression of TRPV1, PPARγ and thermogenic proteins in undifferentiated 3T3-L1 cells but not in differentiated cells. Acute application of Trog stimulated a robust Ca²⁺ influx into 3T3-L1 cells and TRPV1 inhibition by capsazepine prevented this. More interestingly, Trog or capsaicin treatment caused the deacetylation of PPARγ in 3T3-L1 cells and inhibition of TRPV1 or Sirtuin 1 - prevented this. Our data suggest a novel effect of Trog to induce PPARγ deacetylation by activating TRPV1. This research has a significant implication on the role of TRPV1 and PPARγ signaling in the browning of white adipose tissue.     

Classical autophagy proteins LC3B and ATG4B facilitate melanosome movement on cytoskeletal tracks

Macroautophagy/autophagy is a dynamic and inducible catabolic process that responds to a variety of hormonal and environmental cues. Recent studies highlight the interplay of this central pathway in a variety of pathophysiological diseases. Although defective autophagy is implicated in melanocyte proliferation and pigmentary disorders, the mechanistic relationship between the 2 pathways has not been elucidated. In this study, we show that autophagic proteins LC3B and ATG4B mediate melanosome trafficking on cytoskeletal tracks. While studying melanogenesis, we observed spatial segregation of LC3B-labeled melanosomes with preferential absence at the dendritic ends of melanocytes. This LC3B labeling of melanosomes did not impact the steady-state levels of these organelles but instead facilitated their intracellular positioning. Melanosomes primarily traverse on microtubule and actin cytoskeletal tracks and our studies reveal that LC3B enables the assembly of microtubule translocon complex. At the microtubule-actin crossover junction, ATG4B detaches LC3B from melanosomal membranes by enzymatic delipidation. Further, by live-imaging we show that melanosomes transferred to keratinocytes lack melanocyte-specific LC3B. Our study thus elucidates a new role for autophagy proteins in directing melanosome movement and reveal the unconventional use of these proteins in cellular trafficking pathways. Such crosstalk between the central cellular function and housekeeping pathway may be a crucial mechanism to balance melanocyte bioenergetics and homeostasis.     

Genetic analyses using GGE model and a mixed linear model approach,and stability analyses using AMMI bi-plot for late-maturity alpha-amylase activity in bread wheat genotypes

Low falling number and discounting grain when it is downgraded in class are the consequences of excessive late-maturity α-amylase activity (LMAA) in bread wheat (Triticum aestivum L.). Grain expressing high LMAA produces poorer quality bread products. To effectively breed for low LMAA, it is necessary to understand what genes control it and how they are expressed, particularly when genotypes are grown in different environments. In this study, an International Collection (IC) of 18 spring wheat genotypes and another set of 15 spring wheat cultivars adapted to South Dakota (SD), USA were assessed to characterize the genetic component of LMAA over 5 and 13 environments, respectively. The data were analysed using a GGE model with a mixed linear model approach and stability analysis was presented using an AMMI bi-plot on R software. All estimated variance components and their proportions to the total phenotypic variance were highly significant for both sets of genotypes, which were validated by the AMMI model analysis. Broad-sense heritability for LMAA was higher in SD adapted cultivars (53%) compared to that in IC (49%). Significant genetic effects and stability analyses showed some genotypes, e.g. ‘Lancer’, ‘Chester’ and ‘LoSprout’ from IC, and ‘Alsen’, ‘Traverse’ and ‘Forefront’ from SD cultivars could be used as parents to develop new cultivars expressing low levels of LMAA. Stability analysis using an AMMI bi-plot revealed that ‘Chester’, ‘Lancer’ and ‘Advance’ were the most stable across environments, while in contrast, ‘Kinsman’, ‘Lerma52’ and ‘Traverse’ exhibited the lowest stability for LMAA across environments.     

Solution structure and backbone dynamics of the human DNA ligase IIIalpha BRCT domain.

BRCT (BRCA1 carboxyl terminus) domains are found in a number of DNA repair enzymes and cell cycle regulators and are believed to mediate important protein-protein interactions. The DNA ligase IIIalpha BRCT domain partners with the distal BRCT domain of the DNA repair protein XRCC1 (X1BRCTb) in the DNA base excision repair (BER) pathway. To elucidate the mechanisms by which these two domains can interact, we have determined the solution structure of human ligase IIIalpha BRCT (L3[86], residues 837-922). The structure of L3[86] consists of a beta2beta1beta3beta4 parallel sheet with a two-alpha-helix bundle packed against one face of the sheet. This fold is conserved in several proteins having a wide range of activities, including X1BRCTb [Zhang, X. D., et al. (1998) EMBO J. 17, 6404-6411]. L3[86] exists as a dimer in solution, but an insufficient number of NOE restraints precluded the determination of the homodimer structure. However, 13C isotope-filtered and hydrogen-deuterium exchange experiments indicate that the N-terminus, alpha1, the alpha1-beta2 loop, and the three residues following alpha2 are involved in forming the dimer interface, as similarly observed in the structure of X1BRCTb. NOE and dynamic data indicate that several residues (837-844) in the N-terminal region appear to interconvert between helix and random coil conformations. Further studies of other BRCT domains and of their complexes are needed to address how these proteins interact with one another, and to shed light on how mutations can lead to disruption of function and ultimately disease.     

Detection of virulent Shigella and enteroinvasive Escherichia coli by induction of the 43 kDa invasion plasmid antigen, ipaC

The invasion plasmid antigen, ipaC (43 kDa) of Shigella spp. and enteroinvasive Escherichia coli (EIEC) could be induced in vitro by growing them in the presence of Congo red. An enzyme-linked immunosorbent assay (ELISA) using antibodies to the 43 kDa protein of Shigella has been developed for specific detection of virulent Shigella spp and EIEC. The test is independent of initial isolation of individual colonies. As few as 10² CFU/ml of virulent Shigella present in mixed cultures could be detected and concurrently their susceptibility to antibiotics could be analysed after an initial growth of 8–16 h in Congo red-containing medium. The test may prove useful in the diagnosis and treatment of bacillary dysentery caused either by Shigella or EIEC through their rapid identification and proper antimicrobial therapy.     

Evolution of inspiratory and expiratory muscle pressures during endurance exercise.

We investigated the relationship between minute ventilation (VE) and net respiratory muscle pressure (Pmus) throughout the breathing cycle [Total Pmus = mean Pmus, I (inspiratory) + mean Pmus, E (expiratory)] in six normal subjects performing constant-work heavy exercise (CWHE, at approximately 80% maximum) to exhaustion on a cycle ergometer. Pmus was calculated as the sum of chest wall pressure (elastic + resistive) and pleural pressure, and all mean Pmus variables were averaged over the total breath duration. Pmus, I was also expressed as a fraction of volume-matched, flow-corrected dynamic capacity of the inspiratory muscles (P(cap, I)). VE increased significantly from 3 min to the end of CWHE and was the result of a significantly linear increase in Total Pmus (Delta = 43 +/- 9% from 3 min to end exercise, P < 0.005) in all subjects (r = 0. 81-0.99). Although mean Pmus, I during inspiratory flow increased significantly (Delta = 35 +/- 10%), postinspiratory Pmus, I fell (Delta = -54 +/- 10%) and postexpiratory expiratory activity was negligible or absent throughout CWHE. There was a greater increase in mean Pmus, E (Delta = 168 +/- 48%), which served to increase VE throughout CWHE. In five of six subjects, there were significant linear relationships between VE and mean Pmus, I (r = 0.50-0.97) and mean Pmus, E (r = 0.82-0.93) during CWHE. The subjects generated a wide range of Pmus, I/P(cap, I) values (25-80%), and mean Pmus, I/P(cap, I) increased significantly (Delta = 42 +/- 16%) and in a linear fashion (r = 0.69-0.99) with VE throughout CWHE. The progressive increase in VE during CWHE is due to 1) a linear increase in Total Pmus, 2) a linear increase in inspiratory muscle load, and 3) a progressive fall in postinspiratory inspiratory activity. We conclude that the relationship between respiratory muscle pressure and VE during exercise is linear and not curvilinear.