A Preliminary Study Of CAMP Signaling-mediated Regulation Of Growth And Development In Upland Cotton

CyclicAMP(cAMP)is the first discovered “second messenger”,and it participates in the regulation of numerous life activities such as cell metabolism,secretion,signal transduction and gene transcription.cAMP is generated by adenylyl cyclase(AC)directly catalyzing ATP,while it can be hydrolyzed by cyclic nucleotide phosphodiesterase(PDE)to produce adenosine monophosphate(AMP).These two enzymes act through different regulatory mechanisms and together determine the intracellular cAMP levels.While cAMP is a ubiquitous signaling molecule in the biological world,our understanding of the regulatory roles of cAMP signaling in plants remain very limited until now.A few studies indicate that cAMP is involved in the regulation of plant growth and development and many physiological processes,including hormone responses,seed germination,stomatal movement,light and temperature responses,biotic and abiotic stresses responses.Currently,rare documentation was found with the knowledge of cAMP in cotton.This work conducted preliminary studies on the regulatory roles of cAMP signaling in upland cotton,and the results were as follows:1.Using cell permeable cAMP analogue(8-Br-cAMP)and AC specific agonist(forskolin,FSK)and inhibitor(2’,3’-dideoxyadenosine,DDA)to treat cotton seedlings,it was found that DDA significantly inhibited plant height,root length and lateral root development,and the degree of inhibition was significantly increased depending on the elevation of DDA concentration,suggesting that the endogenous production of cAMP is essential for cotton growth and development.However,no significant phenotypic effects were observed with the exogenous application of8-Br-cAMP and FSK.2.Using cotton ovule culture under the treatments of cAMP signaling-related drugs,it was found that DDA treatment significantly inhibited fiber development with the degree of inhibition apparently relying on the DDA concentration,suggesting a critical role of cAMP in cotton fiber development.However,the effects of FSK were not visible under the condition of low concentration(10 μM),and a higher concentration(50 μM)seemed to show a marginal inhibition;8-Br-cAMP had no impact on the fiber development under the tested conditions.3.RNA-seq analysis was performed using cotton ovule culture under the treatments of cAMP signaling-related drugs,significantly differential expression genes(DEGs)were identified and subjected to GO and KEGG enrichment analyses.Our results indicated: under the conditions of DDA treatment,the up-regulated were mainly related to atrichoblast differentiation(GO:0010055),auxin-activated signaling pathway(GO:0009734),developmental process(GO:0032502),regulation of response to nutrient levels(GO:0032107),cell recognition(GO:0008037);down-regulated DEGs were functionally related to cell wall organization or biogenesis(GO:0071554),microtubule-based process(GO:0007017),developmental process(GO:0032502),immune system process(GO:0002376),signaling(GO:0023052),cell division(GO:0051301);these DEGs were mainly associated with KEGG pathways including plant hormone signal transduction(ko04075),biosynthesis of secondary metabolites(ko01110),plant MAPK signaling pathway(ko04016)and phenylpropanoid biosynthesis(ko00940).Under the conditions of FSK treatment,the up-regulated DEGs were mainly related to immune system process(GO:0002376),atrichoblast differentiation(GO:0010055),regulation of phenylpropanoid metabolic process(GO:2000762)and cell recognition(GO:0008037);down-regulated DEGs were mainly related to polysaccharide metabolic process(GO:0005976),sucrose transport(GO:0015770),regulation of jasmonic acid biosynthetic process(GO:0080141)and seed trichome elongation(GO:0090378);they were associated with KEGG pathways mainly including biosynthesis of secondary metabolites(ko01110),phenylpropanoid biosynthesis(ko00940),plant MAPK signaling pathway(ko04016),plant hormone signal transduction(ko04075),and pentose and glucuronate interconversions(ko00040).Under the conditions of 8-Br-cAMP treatment,the up-regulated DEGs were functionally related to biological processes such as cellular anion homeostasis(GO:0030002),response to hydrogen peroxide(GO:0042542)and COP9 signalosome assembly(GO:0010387),while down-regulated DEGs were mainly related to cell wall organization(GO:0071555),regulation of jasmonic acid biosynthetic process(GO:0080141)and response to ozone(GO:0010193);they were mainly associated with KEGG pathways including pentose and glucuronate interconversions(ko00040),phenylpropanoid biosynthesis(ko00940)and plant hormone signal transduction(ko04075).4.By comparison of the DEGs lists obtained above,421,125 and 13 common DEGs were found with different concentrations of DDA,FSK and 8-Br-cAMP treatments,respectively.Among them,there were 3 common DEGs(Gh＿D07G0502;Gh＿D13G2156;Gh＿D12G2181)shared between DDA and FSK,two common DEGs(Gh＿A02G0682;Gh＿D08G0593)shared between FSK and 8-Br-cAMP,but no common DEGs between DDA,FSK and 8-Br-cAMP were found.These DEGs are closely associated with cAMP signaling,providing key targets for further research in future.