Identification Of MicroRNAs Upon Cold And Heat Stresses In Cotton And Role Of GhKCS13 In Regulation Of Cold Sensitivity

Temperature is a key environmental factor affecting the geographical distribution of plants,which also effects on plant' growing season.Plants gradually form the complex regulatory mechanisms in response to cold and heat stresses.Cotton is sensitive to temperature stress.Hence,it is necessary to understand the response mechanisms and explore additional genetic resources for the environmental adaption to temperature stress in cotton.It is important to the improvement of fiber quality and yields,which has important guiding significance for cotton production in China.1.GhKCS13 regulates cold response in cotton by modulating lipid remodeling and jasmonate biosynthesisCold stress is an environmental factor limiting plant development and productivity.The plasma membrane is the first sensor of low temperature,and alterations of membrane lipids can be induced by cold stress from plastid to ER,where the lipids synthesis and elongation as well as galactolipids and sphingolipids biosynthesis.Understanding plant responses to cold stress mediated by lipids is a fundamentally important question,as mechanisms remain quite unclear;and is also important for crop yield enhancement.Very-long-chain fatty acids(VLCFAs)are important component of membrane lipid bilayer likely play a key role in adaptation to cold stress.Here,we report the physiological function of a ?-ketoacyl-CoA synthase gene GhKCS13 in VLCFAs elongation in the defense response to cold stress in cotton.? GhKCS13 overexpressing transgenic cotton plants(KO)show increased sensitivity to cold stress,demonstrating an important role of GhKCS13 in regulating the response to cold stress.ESI-MS/MS analysis shows that overexpressing GhKCS13 leads to modifications of sphingolipid and glycerolipid compositions in leaves,which may alter the fluidity of the cell membrane by the unsaturation degree of lipids and changes of the proportion of non-bilayer lipids under cold conditions.? Particularly,the lipid signal molecules,including unsaturated sphingolipids and phosphatidic acid(PA),which might be players in regulating cotton cold tolerance,were significantly declined in the GhKCS13-overexpression lines and accumulated in the GhKCS13-RNAi lines.? Our findings also show GhKCS13 may participate in response to cold stress via regulating biosynthesis of the oxylipin jasmonic acid(JA),and increased accumulation of JA in GhKCS13 RNAi cottons,and lower JA levels in overexpression lines,may mediate cold stress response together with lysophospholipids.Our results show that overexpressing GhKC.S13 likely causes remodeling of lipids in the ER and oxylipin JA biosynthesis in chloroplasts,which may account for the increased sensitivity to cold stress in the transgenic cotton.Complex interactions between lipid components,lipid signaling,and JA determine the response to cold stress in cotton.2.Deep sequencing exposits small RNA-mediated responses to cotton cold and heat stressCold and heat stresses affect plants' growth and development,and there are complex reactions and regulatory mechanisms responding to stresses performed in plants.The different expression genes involved in stress response can be regulated by transcription factors and non-coding RNAs.One class of endogenous?19-25 nucleotide non-coding RNAs-miRNAs are post-transcriptional gene regulatory molecules associated with stress response.In our study,miRNAs in cotton seedlings treated with different temperature conditions(4,12,25,35,and 42?)are identified by small RNA sequencing.There are 63 known miRNAs and 105 novel miRNAs identified as the stress-responsive miRNAs.216 target genes for 32 known and 31 novel miRNAs targets are analyzed by degradome sequencing.Most of the targets for stress-responsive miRNAs may take part in the response to temperature stress by regulating genes involved in growth homeostasis,chloroplast function,oxidation-reduction reaction and plant hormone signal transduction pathways.Expression patterns between miRNAs and the targets showed perfect negative relationships,which suggested that cotton stress responses were controled by diverse downstream signaling pathways mediated by diverse miRNA regulated target degradation.This research should enrich our understanding of miRNA-mediated regulatory network in cotton tolerance to different temperature stresses.