Hormonal Regulatory Mechanisms Of Helianthus Tuberosus Dwarfism

Crop height is an important agronomic trait regulated by various phytohormones.Semi-dwarf strains can improve the resistance and yield of crops such as rice and wheat,making an important contribution to increasing world food production.Helianthus tuberosus is one of the most important bioenergy crops but existing cultivars of H.tuberosus are tall and prone to collapse,resulting in lower tuber yields and lower quality.H.tuberosus is a hexaploid species and it is difficult to obtain cultivars of ideal stature using traditional breeding methods.LZJ015 is a naturally dwarf H.tuberosus cultivar obtained by our group through germplasm collection.In this thesis,the molecular regulation mechanism of dwarfing in LZJ015 was investigated using phenotypic analysis,metabolomic and transcriptomic methods to analyse the synthesis and metabolic changes of various phytohormones using normal height H.tuberosus LZJ119as a control.Here are the main results:1.Phenotypic characteristics such as plant height and basal diameter and leaf area of dwarf and normal H.tuberosus grown in field and indoor pots were measured to compare the phenotypic differences between the two cultivars of H.tuberosus.The results showed that dwarf H.tuberosus had shorter nodes,more main stems,denser root systems and reddish tubers compared to normal H.tuberosus.Compared to normal H.tuberosus in field and indoor cultivation,dwarf H.tuberosus:reduced plant height by88.44%and 66.80%respectively;reduced the length of the corresponding stem nodes by more than 50%except for the top stem node;reduced the number of main stems in field cultivation by more than 2 compared to 1 in normal H.tuberosus;dwarf H.tuberosus had purplish-red tubers compared to normal H.tuberosus,which were yellowish in appearance;in addition,the number of flowers,lateral branches,and stem nodes of dwarf H.tuberosus was lower than that of normal H.tuberosus under field cultivation conditions,where the number of flowers was reduced by 97.40%,the number of branches at the base of the main stem by 51.52%and the number of main stem nodes by 74.60%compared to normal H.tuberosus.2.Using a targeted metabolomic approach,the levels of eight plant hormones（auxin,cytokinin,gibberellin,abscisic acid,ethylene,jasmonate,salicylic acid and strigolactone）in the stem tips of the two cultivars were analysed comparatively.The results showed that compared with normal H.tuberosus,the dwarf H.tuberosus had:76.42%less auixn and 74.97%less synthetic precursor（mainly tryptophan）,while the metabolite content increased by 4.53 times,featuring faster auxin synthesis and metabolism;7.45 times more cytokinin and 3.86 times more metabolites than normal H.tuberosus;the content of strigolactone decreased by 78.69%,while the precursor of synthesis increased by 1.41 times,with the possible defect in the synthesis pathway of strigolactone;jasmonate increased by 1.32 times and its synthetic precursor content increased by 1.46 times,while the metabolite content decreased by 69.90%;gibberellin decreased by 37.12%,abscisic acid decreased by 43.85%,the content of ACC that is the synthetic precursor of ethylene increased by 54.39%and salicylic acid by 66.14%.The above results tentatively establish that dwarfing of LZJ015 may be associated with a reduction in auxin,strigolactone and gibberellin etc.3.To investigate the mechanism of the defective synthesis of strigolactone in dwarf H.tuberosus,the carotenoid content of both H.tuberosus was analyzed using a targeted metabolomic approach.The results showed a 19.57%reduction in total carotenoid content and a 24.55%reduction inβ-carotene content in the stem tips of dwarf H.tuberosus compared to the control H.tuberosus,and the detection of three violaxanthin derivatives not found in normal H.tuberosus.Using a colorimetric method to determine the total carotenoid content in different organs of the two cultivars,the total carotenoid content in stem tips was consistent with the metabolomic results;however,the carotenoid content of dwarf H.tuberosus roots and tubers increased by 24.74%and58.78%,respectively,compared to the normal cultivar,indicating that carotenoid metabolism is regulated by opposite mechanisms in the stem tips and tubers dwarf H.tuberosus.4.The regulatory mechanisms of hormone synthesis and metabolism in dwarf H.tuberosus were further investigated using a comparative transcriptomic approach.The results showed that compared with normal H.tuberosus,in dwarf H.tuberosus:Total expression of TAA1（L-tryptophan-pyruvate aminotransferase）,a key gene in the indole pyruvate（IPA）pathway of auixn,increased by 31.36%,while total expression of DDC（L-tryptophan decarboxylase）,a key gene in the tryptamine（TAM）pathway,decreased by 39.05%;the expression of CKX genes related to cytokinin metabolism decreased by70.08%;the expression of GA₂₀ox that is a key gene for gibberellin synthesis decreased by 96.25%;the expression levels of 13 genes involved in multiple steps of carotenoid synthesis and metabolism were significantly down-regulated,such as PDS,ZDS,NCED,etc.Among them,the enzyme encoded by NCED is involved in the isomerization of violaxanthin,which is a key gene for the synthesis of abscisic acid and strigolactone.Combined with the metabolomic analysis results,this study showed that the transition of auxin synthesis from the TAM pathway to the IPA pathway in dwarf H.tuberosus led to an acceleration of auxin synthesis,the inhibition of CKX expression that resulted in the increase of cytokinin content,the extreme reduction of GA₂₀ox expression caused a decrease of gibberellin content,and the reduction of carotenoid synthesis and the decline of NCED expression contributed to the reduction of strigolactone and abscisic acid synthesis.In summary,this thesis found that phytohormones directly related to plant height,such as auxin,gibberellin,and strigolactone,are defective in synthesis or metabolism in dwarf H.tuberosus.Key genes related to phytohormone synthesis and metabolism and plant height regulation,such as TAA1,DDC,GA₂₀ox,CKX and NCED,were identified,providing an important theoretical basis for the selection and creation of semi-dwarf H.tuberosus cultivars.