The Study On The Avoidance Behavior Of Pseudomonas Aeruginosa PA14 By Classical Wnt Pathway Of C.elegans

Whether the higher mammals or the lowly single-celled animals respond to environmental stimuli,such as attraction,escape,etc.Avoidance behavior is the most important and effective response to adverse stimuli,and in the case of higher animals,it involves complex molecular mechanisms and neural circuits.So we chose the simpler model organism,Caenorhabditis elegans,to study complex neurobiological behavior as our subject.C.elegans nervous system structure is simple and relatively perfect,hermaphrodite nematodes.There are 302 neurons and 5000 synapses,and hundreds of gap junction,has a variety of higher animal with classical neurotransmitters,and used c.elegans as a model of the neural biology technology,such as orientation,Ca²⁺imaging,fluorescent protein neurons neural electrophysiological detection,microfluidic technology,and laser damage or specific neuron necrosis technology is mature and easy.Wnt signaling pathway is a regulated and conservative approach to Wingless-type?Wnt?protein.Currently,the known Wnt signaling pathway is mainly divided into classical Wnt pathway and non-classical Wnt pathway.Wnt pathway for many aspects of metazoan development is very important,including regulating cell fate,cell proliferation,cell morphology,cell migration,cell apoptosis and differentiation,and stem cell update and so on.In addition,Wnt signaling pathway plays an important role in the regulation mechanism of tumor.However,we found in the data analysis of the transcriptome Wnt signaling pathways in NPR-1 downstream of neuropeptide receptors involved in c.elegans for pathogen Pseudomonas aeruginosaPA14?later PA14?escape behavior.Therefore,our existing research first confirmed by RNAi that the pathway did participate in the avoidance behavior of nematodes.On this basis,the signaling pathway involved in the process is improved,and the upstream and downstream relationships in the pathway are further verified,and the participating neurons are also preliminarily explored.The main results of this study include:1.Determine the classical Wnt pathway the ligands cwn-2,the receptor cfz-2,Dishevelled protein?Dvl?dsh-1,glycogen synthesis kinase gsk-3,and the nucleus transcription factor?-catenin bar-1 to participate in the avoidance of PA14.Through to the Wnt pathway of five ligands,four protein receptors,three Dvl protein kinase,glycogen synthesis and four?-catenin RNAi,found the cwn-2?cfz-2?dsh-1?gsk-3 and bar-1 participate in the avoidance about PA14.The mutant of cwn-2?cfz-2?dsh-1and bar-1make the avoidance index is higher than wild tipe,but the mutant of gsk-3 make the avoidance index is lower than wild tipe.The experimental results of the mutant strain were consistent with the results of RNAi.Therefore,it is determined that Wnt pathway may be involved in regulating the avoidance behavior in C.elegans.In the mutants cfz-2?ok1201?and gsk-3?tm1273?,the two genes,cfz-2 and gsk-3,can be fully responded to the defect phenotypes of cfz-2?ok1201?and gsk-3?tm1273?.Moreover,cfz-2?ok1201?;gsk-3?tm1273?were determined in the same way through the hybridization of cfz-2?ok1201?and gsk-3?tm1273?,and gsk-3?tm1273?played a role in the downstream of cfz-2?ok1201?.2.Wnt pathway controls the avoidance behavior in the downstream of NPR-1.For npr-1?ur89?,gsk-3?tm1273?;npr-1?ur89?was obtained by hybridization between gsk-3?tm1273?and npr-1?ur89?.npr-1?ur89?,compared to gsk-3?tm1273?,npr-1?ur89?and gsk-3?tm1273?;npr-1?ur89?avoidance in the pathogenic bacteria,it is found that gsk-3?tm1273?and npr-1?ur89?have a same way to control the escape behavior of nematodes against the pathogenic bacteria,and gsk-3?tm1273?plays a role in the downstream of npr-1?ur89?.3.In ALM neurons,Wnt pathway may be involved in the avoidance of PA14 by pseudomonas aeruginosa.In order to study whether a certain neuron participated in the process of avoidance behavior involved in Wnt pathway,the two fluorescence positioning vectors Pdsh-1:m-cherry and Pcfz-2::GFP were constructed.Fluorescence microscopy revealed that both fluorescence can be localized in a single neuron in the head.The position identification is the ALM neuron,and the follow-up work needs to be further confirmed.The innovation of this paper:This is novel that the Wnt pathway was found to participate in the avoidance behavior of the pathogenic bacteria PA14 in the Caenorhabditis elegans.