Neuronal KGB-1 JNK MAPK signaling regulates the dauer developmental decision in response to environmental stress in Caenorhabditis elegans

In response to stressful growth conditions of high population density, food scarcity, and elevated temperature, young larvae of nematode Caenorhabditis elegans can enter a developmentally arrested stage called dauer that is characterized by dramatic anatomic and metabolic remodeling. Genetic analysis of dauer formation of C. elegans has served as an experimental paradigm for the identification and characterization of conserved neuroendocrine signaling pathways. Here, we report the identification and characterization of a conserved c-Jun N-terminal Kinase-like mitogen-activated protein kinase (MAPK) pathway that is required for dauer formation in response to environmental stressors. We observed that loss-of-function mutations in the MLK-1-MEK-1-KGB-1 MAPK pathway suppress dauer entry. A loss-of-function mutation in the VHP-1 MAPK phosphatase, a negative regulator of KGB-1 signaling, results in constitutive dauer formation, which is dependent on the presence of dauer pheromone but independent of diminished food levels or elevated temperatures. Our data suggest that the KGB-1 pathway acts in the sensory neurons, in parallel to established insulin and TGF-β signaling pathways, to transduce the dauer-inducing environmental cues of diminished food levels and elevated temperature.     

The dynamin-related protein DRP-1 and the insulin signaling pathway cooperate to modulate Caenorhabditis elegans longevity

Here, we report that inactivation of the Caenorhabditis elegans dynamin-related protein DRP-1, a key component responsible for mitochondrial fission and conserved from yeast to humans, dramatically enhanced the effect of reduced insulin signaling (IIS) to extend lifespan. This represents the first report of a beneficial impact of manipulating mitochondrial dynamics on animal lifespan and suggests that mitochondrial morphology and IIS cooperate to modulate aging.     

A role for the Erk MAPK pathway in modulating SAX-7/L1CAM-dependent locomotion in Caenorhabditis elegans

L1CAMs are immunoglobulin cell adhesion molecules that function in nervous system development and function. Besides being associated with autism and schizophrenia spectrum disorders, impaired L1CAM function also underlies the X-linked L1 syndrome, which encompasses a group of neurological conditions, including spastic paraplegia and congenital hydrocephalus. Studies on vertebrate and invertebrate L1CAMs established conserved roles that include axon guidance, dendrite morphogenesis, synapse development, and maintenance of neural architecture. We previously identified a genetic interaction between the Caenorhabditis elegans L1CAM encoded by the sax-7 gene and RAB-3, a GTPase that functions in synaptic neurotransmission; rab-3; sax-7 mutant animals exhibit synthetic locomotion abnormalities and neuronal dysfunction. Here, we show that this synergism also occurs when loss of SAX-7 is combined with mutants of other genes encoding key players of the synaptic vesicle (SV) cycle. In contrast, sax-7 does not interact with genes that function in synaptogenesis. These findings suggest a postdevelopmental role for sax-7 in the regulation of synaptic activity. To assess this possibility, we conducted electrophysiological recordings and ultrastructural analyses at neuromuscular junctions; these analyses did not reveal obvious synaptic abnormalities. Lastly, based on a forward genetic screen for suppressors of the rab-3; sax-7 synthetic phenotypes, we determined that mutants in the ERK Mitogen-activated Protein Kinase (MAPK) pathway can suppress the rab-3; sax-7 locomotion defects. Moreover, we established that Erk signaling acts in a subset of cholinergic neurons in the head to promote coordinated locomotion. In combination, these results suggest a modulatory role for Erk MAPK in L1CAM-dependent locomotion in C. elegans.     

Caenorhabditis elegans mounts a p38 MAPK pathway‐mediated defence to Cutibacterium acnes infection

Cutibacterium acnes is capable of inducing inflammation in acne and can lead to a chronic prostatic infection. The diverse pathogenicity among different strains of C. acnes has been presented, but simple appropriate animal models for the evaluation of this bacterium are lacking. In this study, the nematode Caenorhabditis elegans was used as an invertebrate infection model. We revealed that C. acnes type strain ATCC 6919 caused lethal infections to C. elegans in solid and liquid culture media (p < .0001). Compared with the strain ATCC 6919, the antibiotic‐resistant strain HM‐513 was more virulent, resulting in reduced survival (p < .0001). Four different C. acnes strains killed worms with a p value of less than .0001 when provided to C. elegans at 4.8 × 10⁸ CFU/ml. The infection model was also employed to explore host defence responses. An increase in numerous immune effectors in response to C. acnes was detected. We focused on nine C‐type lectins, including: clec‐13, clec‐17, clec‐47, clec‐52, clec‐60, clec‐61, clec‐70, clec‐71 and clec‐227. The induced expression of these C‐type lectin genes was down‐regulated in mutant worms deficient in the p38 mitogen‐activated protein kinase (MAPK) pathway. Meanwhile, PMK‐1 (MAPK) was phosphorylated and activated at the onset of C. acnes infection. By monitoring the survival of mutant worms, we found that PMK‐1, SEK‐1 (MAPKK) and TIR‐1 (MAPKKK) were critical in responding to C. acnes infection. C. elegans pmk‐1 and tir‐1 mutants exhibited higher mortality to C. acnes infection (p < .0001). In conclusion, C. elegans serves as a simple and valuable model to study C. acnes virulence and facilitates improvements in understanding of host innate immune responses.     

G protein–coupled receptor kinase 2 modifies the ability of Caenorhabditis elegans to survive oxidative stress

Survival and adaptation to oxidative stress is important for many organisms, and these occur through the activation of many different signaling pathways. In this report, we showed that Caenorhabditis (C.) elegans G protein–coupled receptor kinases modified the ability of the organism to resist oxidative stress. In acute oxidative stress studies using juglone, loss-of-function grk-2 mutants were more resistant to oxidative stress compared with loss-of-function grk-1 mutants and the wild-type N2 animals. This effect was Ce-AKT-1 dependent, suggesting that Ce-GRK2 adjusted C. elegans oxidative stress resistance through the IGF/insulin-like signaling (IIS) pathway. Treating C. elegans with a GRK2 inhibitor, the selective serotonin reuptake inhibitor paroxetine, resulted in increased acute oxidative stress resistance compared with another selective serotonin reuptake inhibitor, fluoxetine. In chronic oxidative stress studies with paraquat, both grk-1 and grk-2 mutants had longer lifespan compared with the wild-type N2 animals in stress. In summary, this research showed the importance of both GRKs, especially GRK2, in modifying oxidative stress resistance.     

Down-regulation of HIV-1 infection by inhibition of the MAPK signaling pathway

The human immunodeficiency virus type 1 (HIV-1) can interact with and exploit the host cellular machinery to replicate and propagate itself. Numerous studies have shown that the Mitogen-activated protein kinase (MAPK) signal pathway can positively regulate the replication of HIV-1, but exactly how each MAPK pathway affects HIV-1 infection and replication is not understood. In this study, we used the Extracellular signal-regulated kinase (ERK) pathway inhibitor, PD98059, the Jun N-terminal kinase (JNK) pathway inhibitor, SP600125, and the p38 pathway inhibitor, SB203580, to investigate the roles of these pathways in HIV-1 replication. We found that application of PD98059 results in a strong VSV-G pseudotyped HIV-1_NL4-3 luciferase reporter virus and HIV-1_NL4-3 virus inhibition activity. In addition, SB203580 and SP600125 also elicited marked VSV-G pseudotyped HIV-1_NL4-3 luciferase reporter virus inhibition activity but no HIV-1_NL4-3 virus inhibition activity. We also found that SB203580 and SP600125 can enhance the HIV-1 inhibition activity of PD98059 when cells were treated with all three MAPK pathway inhibitors in combination. Finally, we show that HIV-1 virus inhibition activity of the MAPK pathway inhibitors was the result of the negative regulation of HIV-1 LTR promoter activity.     

Caenorhabditis elegans mom-4 is required for the activation of the p38 MAPK signaling pathway in the response to Pseudomonas aeruginosa infection

The p38 mitogen-activated protein kinase (MAPK) plays an evolutionarily conserved role in the cellular response to microbial infection and environmental stress. Activation of p38 is mediated through phosphorylation by upstream MAPKK, which in turn is activated by MAPKKK. In the Caenorhabditis elegans, the p38 MAPK (also called PMK-1) signaling pathway has been shown to be required in its resistance to bacterial infection. However, how different upstream MAP2Ks and MAP3Ks specifically contribute to the activation of PMK-1 in response to bacterial infection still is not clearly understood. By using double-stranded RNA-mediated interference (RNAi) and genetic mutants of C. elegans, we demonstrate that C. elegans MOM-4, a mammalian TAK1 homolog, is required for the resistance of C. elegans to a P. aeruginosa infection. We have also found that the MKK-4 of C. elegans is required for P. aeruginosa resistance, but not through the regulation of DLK-1. In summary, our results indicate that different upstream MAPKKKs or MAPKKs regulate the activation of PMK-1 in response to P. aeruginosa.     

The serotonin receptor SER-1 (5HT2ce) contributes to the regulation of locomotion in Caenorhabditis elegans

Serotonin (5-hydroxytryptamine: 5HT) is an important neuroactive substance in the model roundworm, Caenorhabditis elegans. Aside from having effects in feeding and egg-laying, 5HT inhibits motility and also modulates several locomotory behaviors, notably food-induced slowing and foraging. Recent evidence showed that a serotonergic 5HT2-like receptor named SER-1 (also known as 5HT2ce) was responsible for the effect of 5HT on egg-laying. Here we confirm this observation and show that SER-1 also plays an important role in locomotion. A mutant lacking SER-1 was found to be highly resistant to exogenous 5HT in the absence of food and this resistant phenotype was rescued by reintroducing the SER-1 gene in a mutant background. Pharmacological studies showed that the same antagonists that blocked the activity of recombinant SER-1 in vitro also inhibited the effect of 5HT on motility, suggesting the same receptor was responsible for both effects. When tested for locomotory behaviors, the SER-1 mutant was found to be moderately defective in food-induced slowing. In addition, the mutant changed direction more frequently than the wildtype when searching for food, suggesting that SER-1 may play a role in navigational control during foraging. Both these effects required the presence of MOD-1, a 5HT gated chloride channel, and the results indicate that SER-1 and MOD-1 modulate these behaviors through a common pathway. On the basis of expression analysis of a ser-1::GFP translational fusion, SER-1 is prominently located in central, integrating neurons of the head ganglia (RIA and RIC) but not the body wall musculature. The evidence suggests that SER-1 controls locomotion through indirect modulation of neuromuscular circuits and has effects both on speed and direction of movement.     

线虫PCR:直接针对秀丽线虫进行PCR扩增基因组DNA的新方法

目的:直接针对秀丽线虫进行PCR反应,以便快速扩增基因组DNA,从而提高钓取目的基因和鉴定基因组是否发生突变的效率.方法:根据生物信息学分析,针对不同基因设计单重或多重PCR引物;在不含砌DNA聚合酶的PCR反应体系中加入蛋白酶K消化秀丽线虫染色体中的组蛋白,然后加入Taq酶,直接针对野生型或突变型秀丽线虫个体进行PC...     

Evodiamine inhibits adipogenesis via the EGFR-PKCα-ERK signaling pathway

The molecular mechanism of the anti-adipogenic effect of evodiamine (which has several capsaicin-like pharmacological actions) was investigated. The evodiamine effect was not blocked by the specific TRPV1 antagonist capsazepine in 3T3-L1 preadipocytes, whereas its effect was greatly curtailed by inhibitors of protein kinase C (PKC) and epidermal growth factor receptor (EGFR). Signal analyses showed that evodiamine stimulated the phosphorylation of EGFR, PKCα, and ERK, all of which were reduced by an EGFR inhibitor. Silencing experiments of EGFR mRNA supported the involvement of these signaling molecules in the inhibitory effect of evodiamine. An unidentified mechanism whereby evodiamine inhibits adipogenesis via the EGFR-PKCα-ERK signaling pathway was revealed.     

Predicting aging/longevity-related genes in the nematode Caenorhabditis elegans

We present a novel mathematical/computational strategy for predicting genes/proteins associated with aging/longevity. The novelty of our method arises from the topological analysis of an organismal longevity gene/protein network (LGPN), which extends the existing cellular networks. The LGPN nodes represent both genes and corresponding proteins. Links stand for all known interactions between the nodes. The LGPN of C. elegans incorporated 362 genes/proteins, 160 connecting and 202 age-related ones, from a list of 321 with known impact on aging/longevity. A 'longevity core' of 129 directly interacting genes or proteins was identified. This core may shed light on the large-scale mechanisms of aging. Predictions were made, based upon the finding that LGPN hubs and centrally located nodes have higher likelihoods of being associated with aging/longevity than do randomly selected nodes. Analysis singled-out 15 potential aging/longevity-related genes for further examination: mpk-1, gei-4, csp-1, pal-1, mkk-4, 4O210, sem-5, gei-16, 1O814, 5M722, ife-3, ced-10, cdc-42, 1O776Co, and 1O690.     

pkc-1 regulates daf-2 insulin/IGF signalling-dependent control of dauer formation in Caenorhabditis elegans

In Caenorhabditis elegans, the insulin/IGF pathway participates in the decision to initiate dauer development. Dauer is a diapause stage that is triggered by environmental stresses, such as a lack of nutrients. Insulin/IGF receptor mutants arrest constitutively in dauer, an effect that can be suppressed by mutations in other elements of the insulin/IGF pathway or by a reduction in the activity of the nuclear hormone receptor daf‐12. We have isolated a pkc‐1 mutant that acts as a novel suppressor of the dauer phenotypes caused by insulin/IGF receptor mutations. Interactions between insulin/IGF mutants and the pkc‐1 suppressor mutant are similar to those described for daf‐12 or the DAF‐12 coregulator din‐1. Moreover, we show that the expression of the DAF‐12 target daf‐9, which is normally elevated upon a reduction in insulin/IGF receptor activity, is suppressed in a pkc‐1 mutant background, suggesting that pkc‐1 could link the daf‐12 and insulin/IGF pathways. pkc‐1 has been implicated in the regulation of peptide neurosecretion in C. elegans. Although we demonstrate that pkc‐1 expression in the nervous system regulates dauer formation, our results suggest that the requirement for pkc‐1 in neurosecretion is independent of its role in modulating insulin/IGF signalling. pkc‐1 belongs to the novel protein kinase C (nPKC) family, members of which have been implicated in insulin resistance and diabetes in mammals, suggesting a conserved role for pkc‐1 in the regulation of the insulin/IGF pathway.