Molecular Basis Analysis Of Response To Elevated Carbon Dioxide During Growth And Development In Carrot

Carbon dioxide is a major contributor of the 'global greenhouse gases'.During the last 200 years,atmospheric CO2 concentration has increased from a level of 280 to 400?mol·mol-1 and still increasing over this century.CO2 enrichment has been shown to increase plant growth,development and yield of agricultural crops,and its response is a function of CO2 concentration and duration.Carrot(Daucus carota L.)is a biennial herb and belongs to the Daucus genus in the Apiaceae family.Carrot is an important vegetable and develops its taproot as the commonly eaten part.Under the elevated CO2 concentration and other abiotic stress treatments(high-salinity,drought,heat and cold),the growth and development have great changes in carrot.In this study,our objective was to investigate the effects of elevated carbon dioxide on morphological and anatomical characteristics,IAA and AsA accumulation in 'Kurodagosun' carrot.In order to further investigate the response mechanism in carrot under elevated carbon dioxide,the iTRAQ-based proteomic approach was used to analyze the different expression proteins(DEPs)of carrot taproots under elevated carbon dioxide.The DcPHD family transcription factors were investigated in'Kurodagosun' under abiotic stress treatmentes(high-salinity,drought,heat and cold)in this study.The main results are as follows:1.To explore the changes of IAA accumulation during the growth and the regulated mechanisms under elevated carbon dioxide in carrot,'Kurodagosun' samples from five stages(25,40,60,75 and 90 days)in three tissues(roots,leaf blades,petioles)were collected.The relative expression levels of 18 genes related to IAA biosynthesis and signaling were determined using qPCR.The results found that IAA levels underwent great changes at five successive stages,and IAA contents in the petioles and leaf blades were relatively higher than those in the roots.In leaf blades and petioles,IAA contents were peaked at 60 days,whereas the IAA levels were highest at 40 days in roots.Transcript levels of most genes were not well correlated with IAA accumulation.IAA may regulate carrot growth and development in tissue-specific and stage-dependent manner.These results indicated that IAA metabolic mechanism may be regulated through a complex network.Based on the above research,three carrot cultivars('Kurodagosun' 'Deep purple',and'QiTouHuang')(40 days)were subjected to 400 ?mol·mol-1(control)or 3000 ?mol mol-1(treated)CO2,respectively.The taproot samples of carrot were collected at 0,3,6 and 12 h under elevated CO2.In order to further investigate the affects of IAA contents under elevated carbon dioxide,the IAA contents were investigated.The study found that the IAA levels of three carrot cultivars were decreased at 3 h.The IAA contents were increased at 6 h and 12 h of 'Kurodagosun' and 'Deep purple',respectively.The IAA contents were successive decreased in 'QiTouHuang'.These results showed that the IAA contents of three carrot cultivars and the degree of response of elevated CO2 were different in three carrot cultivars.2.Two carrot cultivars('Kurodagosun' and 'Deep purple')were treated with ambient CO2(a[CO2],400 ?mol·mol-1)and elevated carbon dioxide(e[CO2],3000 ?mol·mol-1)concentrations in current study.The research discovered that under e[CO2]conditions,taproot and shoot fresh weights and the root/shoot ratio of carrot significantly decreased as compared with the control group.Elevated CO2 resulted in obvious changes in anatomy and ascorbic acid accumulation in carrot taproots.Compared with the control plants,the levels of AsA and T-AsA under e[CO2]slightly decreased in the two carrot cultivars.Moreover,the transcript profiles of 12 genes related to AsA biosynthesis and recycling were altered in response to e[CO2].These results showed that the e[CO2]possibly inhibited the growth and development and the root/shoot ratio of carrots.The xylem of carrot taproots were thought to be involved in e[CO2]stress tolerance mechanisms.The 'Kurodagosun' and 'Deep purple' carrots differed in sensitivity to e[CO2].3.The 'Kurodagosun' carrot was treated with control(a[CO2],400 ?mol·mol-1)and treated(e[CO2],3000 ?mol·mol-1)CO2 concentrations for 30 days in this research.Untreated seedlings(400 ?mol·mol-1 CO2)were used as the control.An iTRAQ based proteomic approach was used to investigate the proteomes of carrot taproots under e[CO2]stress treatment.In total,204 proteins with significantly altered abundances were identified.These differentially expressed proteins(DEPs)were from 38 biological processes,28 cellular processes and 14 molecular processes using GO enrichment.Moreover,protein-protein interaction analysis revealed that proteins involved in carbohydrate,energy metabolism,protein synthesis and redox homeostasis could be assigned to stress response networks under e[CO2].4.All nucleotide and protein sequences of DcPHD family TFs in carrot were obtained from CarrotDB,a database of genomic and transcriptomic sequence for carrot,which was built by our group.The leaf blades samples of carrot were collected at 0,1,2,4,8 and 12 h after abiotic stress treatments(high-salinity,drought,heat and cold).To confirm the subgroup classification and analyze the evolutionary relationship between carrot,Arabidopsis and Populus trichocarpa,we constructed the phylogenetic tree using PHD TFs sequences.The carrot PHD-finger family factors were divided into 11 subgroups(A-J,L).The multiple sequence alignment showed that the DcPHD proteins domain contained the C-X(1-2)-C-X(8-14)-C-X(2-8)-C-x(4-6)-H-X2-C-X(11-34)-C-X2-C(X represents any amino acid)motif.The PHD domain is a motif composed of seven cysteines and one histidine,in conjunction with zinc ions.The qPCR results showed that the relative expression levels of the four DcPHD genes(DcPHD-I-12?DcPHD-I-13?DcPHD-I-15 and DcPHD-I-24)were showed great changes under stress treatments(high-salinity,drought,heat and cold).