Phosphatidylcholine Mediates LET-607 Regulates DAF-16 Stress Response Pathways And LONGEVITY

Organism have complex stress response pathways to defend against numerous environmental and stresses,disturbance of stress response leads to compromised cellular homeostasis and organismal aging.Eukaryotic cells have evolved a membrane-enclosed subcellular structure that gives cells the ability to maintain a unique microenvironment,promotes the effective separation of various biological processes,and regulates the concentration,transport and diffusion of thousands of molecules required for proper cell function.This membrane closed system also allows cells to physically control their damage,thereby inducing chaperones and degradation mechanisms in specific subcellular structures,isolating misfolded proteins and abnormal molecules,so that other parts of the cell remain Normal operation.Therefore,each subcellular structure has evolved a unique stress ability.When pressure is exerted by the outside world,stress responses are typically activated in specific cellular compartments,allowing the cell to specifically cope with stresses or damages,while maintaining the normal functions of other parts of the cell.As a major site for protein folding and lipid biosynthesis,the endoplasmic reticulum is a key organelle for cellular stress responses,such as the unfolded protein response that restores proteostasis upon proteotoxic stress,also known as the ER stress.ER-resident transcription factors play crucial roles in ER stress responses.CREBH,which belongs to the CREB3 transcription factor family,is an important ERbound transcription factor.In response to ER stress,CREBH transits to the Golgi upon activation,where it is further cleaved by the proteases,and the cleaved N-terminal fragment translocates to the nucleus to activate downstream target genes,a process referred to as regulated intramembrane proteolysis.Activation of CREBH promotes the expression of inflammatory factors and iron-regulatory hormone hepcidin,thereby linking ER stress to the cellular defense responses.Here,we used the model organism C.elegans to study LET-607,the C.elegans ortholog of mammalian CREBH,a regulator of cellular defense response and proteostatic response.The main results are as follows:1.LET-607 regulates multiple stress responseslet-607 RNAi increased C.elegans survival in response to included Pseudomonas aeruginosa strain PA14-induced pathogen stress,tert-butyl hydroperoxide(TBHP)-induced xenobiotic stress,35 ?C heat stress,dithiothrei-tol(DTT)-induced ER stress and longevity.We showed that unlike mammalian CREBH,LET-607 locates in the nucleus but not on the ER.However,LET-607 could regulate cellular defense genes and UPR genes like CREBH,indicating their function may be conserved.2.LET-607 regulates stress resistance and longevity via DAF-16This identified 860 genes regulated by LET-607,of which 316 were upregulated and544 were downregulated Genes which were decreased by let-607 RNAi were classified as upregulated by LET-607.Functional analysis revealed these genes were enriched in immune and detoxification responses.These upregulated genes included C-type lectins and innate immunity genes,which are known to play a key role in pathogen defenses.These findings are again consistent with the function of mammalian CREBH.In addition,cytochrome P450,which mediates detoxification response,and peptidases,which are critical for cellular proteostasis,were also upregulated.Genetic analysis reveal that let-607 RNAi activates the nematode FOXO homologous protein DAF-16,and further promotes the enhancement of the nematode oxidative stress resistance and the extension of lifespan in a DAF-16-dependent manner.This result indicates that LET-607 is a novel aging regulator.3.LET-607 regulates PC metabolismWorking in C.elegans,knockdown of LET-607 induces a metabolic enzyme,called sphingomyelin synthase SMS-5,which metabolizes membrane lipid phosphatidylcholine to diacyl glycerol and sphingomyelin.The sms-5 RNAi can inhibit DAF-16 activation induced by let-607 RNAi,enhance oxidative stress resistance and extend lifespan.Through biochemical and genetic analysis,it is found that the decrease in the content of unsaturated phosphatidic acid choline caused by the activation of sms-5 caused by let-607 knockdown is the main reason for the activation of DAF-16.4.LET-607 regulates DAF-16 via calcium signalingPC is one of the most abundant membrane lipids,PC composition influences membrane properties that in turn alter the functions of the membrane proteins.The ERlocated calcium channel and calcium homeostasis are regulated by alterations in PC contents.Our data shows that SMS-5 consumes unsaturated phosphatidylcholine,the reduction of which signals to DAF-16 through ITR-1-dependent calcium signaling and calcium-sensitive kinase PKC-2.We identified the sphingomyelin synthase SMS-5 as a central mediator of the communication between LET-607 and DAF-16.Our findings reveal a novel crosstalk between LET-607 and the cytosolic DAF-16 stress pathways.Since suppression of LET-607 induces an adaptive activation of DAF-16,a master regulator of cytosolic stress responses in C.elegans,leading to improved animal health and longevity.Our data uncover the significance of crosstalk between different stress pathways in animal fitness,and identify LET-607/CREBH and specific PC as a regulator of DAF-16 and longevity.Collectively,our results delineate a cellular mechanism linking different stress response pathways and highlight its importance in animal longevity.