Responses Of Cottonseed Kernel Development To Combined Elevated Temperature And Soil Drought During The Flowering And Boll Formation

Cottonseed is the basis and main by-product of cotton production.It is rich in highquality oil and protein as well as the fifth-largest vegetable oil and the second-largest protein sources in the world.Owing to global warming,a global average air temperature increase of 2.8±0.4℃ is predicted for 2100,increasing the risk and frequency of extreme weather events.The mean daytime air temperature of 32℃ was the critical temperature of warming effect,when it was higher than 35℃,cotton would suffer from high temperature stress.High temperature and drought occurred frequently and often accompanied during cotton flowering and boll formation,which is not conducive to the formation of cotton yield and quality.To explore the physiological mechanisms for the changes of cottonseed yield and quality under elevated temperature and soil drought,ponds experiments were carried out at the Pailou Experimental Station(32°02’ N,118°50’ E)in Nanjing,Jiangsu Province under two temperature regimes:ambient temperature(AT,31.0/26.4℃,mean daytime/night temperature)and elevated temperature(ET,33.4/28.9℃)and three levels of soil relative water content(SRWC)[(75±5)%,(60±5)%and(45±5)%]with two cultivars Sumian 15(heat-susceptible cultivar)and PHY370WR(heat-tolerant cultivar)during 2016-2018.We examined the co-effects of elevated temperature and drought on cottonseed yield,oil and protein quality,and physiological mechanisms of oil and protein synthesis in the cottonseed based on substance concentrations,enzymes activities,and genes expression.In addition,we analyzed the differences between the two cotton cultivars with different heat tolerance in response to the combined stress.The main results were as follows:1.Effects of combined elevated temperature and soil drought on cottonseed yield,oil and protein qualityElevated temperature(ET),drought and the combination of ET and drought had negative effects on cottonseed biomass accumulation and cottonseed yield formation.Both ET and drought significantly reduced the boll number,single seed weight,seed biomass and cottonseed yield.ET combined with SRWC(45±5)%caused the largest decrease in cottonseed yield in comparison with the control treatment(SRWC(75±5)%under AT,AT75).The cottonseed yield of Sumian 15 was decreased by 45.5%-54.9%;for PHY370WR,the decrease amplitude was 43.2%-49.2%across the three years.Drought tended to increase oil content and reduce protein content,whereas ET showed almost the opposite effects.Under the combination of ET and soil drought,oil content was still higher than under drought alone,although ET weakened the beneficial effects of drought.For protein,ET offset the negative impacts of mild drought on protein content,but protein content was not increased under SRWC(45±5)%.Both oil yield and protein yield were significantly decreased under ET,soil drought,and their combination in both cultivars.The negative effects of the combined stresses were more pronounced.Relative to control,oil yield and protein yield decreased by respectively 23.7%-58.2%and 27.3%-64.0%under ET combined with drought in Sumian 15 and decreased by 15.1%-45.8%and 19.0%-53.2%in PHY370WR across three years.ET,drought and the combined stress reduced unsaturated fatty acid(UFA)and increased saturated fatty acid(SFA),resulting in lower UFA/SFA.Essential amino acid(EAA)and non-essential amino acid(NAA)were increased under ET,but decreased significantly under drought.Under the combined stress,EAA and NAA were increased but EAA/NAA decreased significantly.Compared with PHY370WR,the sensitivity of Sumian 15 to the combined factors was evidenced in cottonseed yield components,oil and protein contents,fatty acid and amino acid compositions.2.Effects of combined elevated temperature and soil drought on the accumulation of kernel biomass,oil and protein of cottonseed kernelBoth ET and drought limited cottonseed kernel yield,and ET coupled with drought decreased seed weight,kernel weight,kernel percentage(kernel weight/seed weight)and kernel yield more significant than either single factor.ET enhanced the maximum accumulative rate(Vmax)of cotton kernel biomass,but significantly reduced the rapid accumulation duration(T),leading to a lower kernel biomass;drought and the combined stress significantly decreased the final kernel biomass by limiting Vmax and T,and the decrease of kernel biomass was greater than did exposure to the individual factors alone.Sucrose and hexose(glucose and fructose)are important carbohydrates affecting the development of cottonseed kernel.Sucrose content was increased with increasing temperature and decreased under soil drought and the combined ET and drought,and the greater decrease was detected by the combined factors in two cultivars.On the contrary,the hexose content(glucose and fructose)was increased significantly under drought and decreased significantly under ET or the coupling of ET and drought,but the decrease under the combined stress was lower than that under ET alone.Both ET and drought enhanced Vmax and reduced T of oil content accumulation,but the increase of Vmax under ET could not offset the adverse effect of the decrease of T value on oil accumulation,leading to lower final oil content decreased significantly.However,drought and the combined treatments were favorable to oil accumulation,but oil content was lower under the combined stress than that under drought alone.Compared with SRWC(75±5)%,protein content was less affected by SRWC(60±5)%;however,SRWC(45±5)%significantly reduced Vmax and T,resulting in lower protein content.ET was advantageous to protein accumulation by increasing Vmax significantly,but the effect of the combined stresses on protein accumulation depends on the level of soil relative water content.ET combined with SRWC(605)%increased protein content,but ET combined with SRWC(45±5)%was unfavorable for protein accumulation.Compared with PHY370WR,Sumian 15 was more sensitive to the combined factors on kernel yield,hexose content,oil and protein content.3.Effects of combined elevated temperature and soil drought on enzyme activities and gene expressions of key enzymes in cottonseed kernel developmentDrought increased activities of SuSy and Inv,but decreased SPS activity,inhibiting sucrose degradation and promoting hexose synthesis;simultaneously,activities of PEPCase,ACCase,PAP and DGAT were enhanced,which was conducive to triglycerides synthesis and oil accumulation.ET reduced the activities of SuSy and Inv,but increased SPS activity,resulted in lower hexose content;meanwhile,although PEPCase activity increased,ACCase,PAP and DGAT activities decreased significantly,thus oil content was decreased due to insufficient substrate and inhibited triglyceride synthesis.The combined treatments downregulated GhSuSy,GhInv and GhSPS,which inhibited sucrose synthesis and reduced hexose content;however,ET combined with drought decreased ACCase activity but up-regulated GhPEPC-1,GhDGAT-1 and enhanced the activities of PEPCase and DGAT,which was beneficial to the distribution of carbon skeleton to the direction of oil synthesis,triglyceride synthesis,and led a higher oil content than the control.For protein synthesis,ET increased GS and GOGAT activities through the up-regulation of GhGS and GhGOGAT to enhance protein accumulation.SRWC(60±5)%had no significant effect on protein accumulation.The variation trend of protein accumulation under ET combined with SRWC(60±5)%was consistent with that under ET.Whereas,SRWC(45±5)%and ET combined with SRWC(45±5)%down-regulated GhGS and GhGOGAT to limit protein synthesis.In conclusion,the synthesis and accumulation of oil in cottonseed kernel are mainly regulated by hexose content.SuSy,Inv,PEPCase and DGAT were the key enzymes,and GhSuSy,GhInv,GhPEPC-1 and GhDGAT-1 are the key enzyme genes sensitive to the combined factors of oil synthesis.The synthesis and accumulation of protein are mainly regulated by free amino acid content.GS and GOGAT are the key enzymes sensitive to the combination of ET and drought,and GhGS and GhGOGAT are the key enzyme genes.Compared with the heat-tolerant cultivar(PHY370WR),the heat-susceptible cultivar(Sumian 15)was more sensitive to combined stress on the enzyme activities and the expression levels of key enzyme genes of oil and protein synthesis.