Dissection Of The Function And Molecular Mechanism Of Flowering Locus T In Potato Tuber Formation

Potato(Solanum tuberosum L.)is the most important global food crop after wheat and rice.Its underground tubers are the harvesting and reproductive organs.Because of its strong adaptability and comprehensive nutrition,potato is widely cultivated in more than150 countries around the world,and plays an important function in ensuring food security.Therefore,understanding how initiation and growth of potato tubers is regulated contributes to breeding innovations in seed potatoes,improvements in tuber yield potential and tuber numbers,and cultivation of potato in the hot tropics and areas with short growing seasons.In addition,tuber is a storage organ derived from the expansion of the sub-apical of underground stolon,the study of the formation of this metamorphic organ is also a fundamental question in plant developmental biology.Some plants have evolved the ability to develop their leaves,stems or roots into storage organs,these storage organs provide plants with additional survival strategies under continuously fluctuating or extreme climate conditions.Potato tuber formation is an adaptive biological trait formed in the long-term evolution process,showing the characteristics of "genetic-environment" interaction.Day length plays a key role in tuber formation,and overcoming the short-day dependence of tuber formation is the most important adaptation after landing in Europe from South America.Besides,the plant’s response to photoperiod determines its maturity.Previous studies have shown that the COFT pathway that controls floral transition of flowering plants,also regulates the formation of potato tubers.And a molecular genetic pathway for phytochrome to sense day-length cues and regulate the rhythm of St COL1 protein,so determine the expression of the tuberinducing signal St SP6 A has been established.However,the functions and mechanisms of tuber-forming signals are not fully understood.In this study,by using potato Solanum tuberosum(S.tub)and wild non-tuber-bearing species Solanum etuberosum(S.etub)and other plant materials,combined with methods such as grafting and functional genomics,the induction and molecular mechanism of potato tuber formation were studied.The results are as follows:1.By identifying the potato FT-like gene family,this study identified two additional FT-like genes in the reference genome DM1-3,named St SP5G-B and FLOWERING LOCUS T-like 1(St FTL1).Gene expression analysis showed that St FTL1 and florigen St SP3 D exhibit expression patterns analogous to tuberigen St SP6 A in leaves.Overexpression of St SP3 D or St FTL1 in the short-day potato E109 background promotes day-neutral tuberization.Grafting experiments showed that the protein and tuber induction effects of St SP3 Dox and St FTL1 ox are graft transmissible,and both St SP3 D and St FTL1 were involved in directing the expression of St SP6 A at the swelling-stolon.Collectively,these results suggest that St SP3 D and St FTL1 are long-range mobile signals for potato tuber-formation.2.The long-day Solanum wild species S.etub was used as scion to graft with E109 rootstock,and the grafts only flowered and tuberized under long-day conditions.Homologous cloning revealed that the deletion of full fourth exon of Se SP6 A,the homolog of St SP6 A,that is,S.etub is a natural loss-of-function mutant of SP6 A.St SP3 Doverexpressing S.etub transgenic shoots promotes tuber formation of St SP3Dox/E109 grafts under short-day conditions,while Se SP3 D knockout delays tuber formation of CRSe SP3D/E109 graft chimeras under long-day conditions.Further,knockout of Se SP5 G,Se SP3 D suppressor,promotes Se SP3 D expression and tuber formation of CRSe SP5G/E109 graft chimeras under short-day conditions,suggestive of the shoot-expressed SP6 A is dispensable for St SP3 D long-range activity.3.The protein interaction assay showed that St SP3 D and St FTL1 interact with the St SP6A-binding proteins.m RNA transcriptome data showed that overexpression of St SP3 D and St FTL1 induced similar transcriptional responses in non-swelling and swelling stolon tips.AGAMOUS-like gene,acting downstream gene of St SP6 A,was up-regulated at the non-swelling stolon tip of St SP3 Dox and St FTL1 ox stolons,while St SP5G-B and St SP6 A were only up-regulated in swelling stolons tips,suggesting that the possibility that the induction of AGAMOUS-like by the FT-like signals may precede that of St SP5G-B and St SP6 A during tuber induction.4.Overexpression of St SP3 D and St FTL1 promoted early flowering and accelerated flower bud maturation,but led to a simplified inflorescence,demonstrating its conserved florigen activity.Mutation of Se SP5 G and overexpressing of St FTL1 promoted flowering under low light intensity,indicating that the FT-like genes are also involved in light irradiance-controlled flowering.Additionally,the secondary tubers are formed directly in tuber sprouts of St SP3D-overexpressing tubers,suggesting that St SP3 Dox reprogrammed the meristem activity of tuber sprouts.Furthermore,St SP3 Dox plants also exhibit multiple morphological phenotypes such as attenuated leaf epidermal hair formation,stem thickness and branching,indicating the pleiotropic nature of St SP3 D.Since St SP3D-RNAi plants late flowered,thus,St SP3 D regulate potato flowering and tuberization in a dose-dependent manner.5.It was identified that ABI5 like 1(St ABL1),the core component of ABA signalling,binds to St SP3 D and St SP6 A through 14-3-3 to form a transcriptional complex,respectively.Overexpression of St ABL1 promote potato flowering and early tuberization,and exhibit accelerated leaf senescence and a short life cycle.Ch IP-seq and RNA-seq analysis showed that the AGAMOUS-like and St GA2ox1 genes were directed by St ABL1.Hormone profiling showed that the gibberellin levels change caused by St ABL1 overexpression was consistent with that induced by St GA2ox1 overexpression.In addition,St ABL1-overexpressing plants were less sensitive to gibberellins treatment in terms of tuberization.Taken together,these results indicated that transcription factor St ABI5 like 1 binding to FLOWERING LOCUS T homologs promotes early maturity in potato.In conclusion,this study identify St SP3 D and St FTL1 as long-range tuber-inducing signals,and uncover the molecular mechanism that the transcription factor St ABL1 interacts with St SP3 D and St SP6 A to direct gibberellin homeostasis and thereby promote synchronization of potato maturity syndrome.These results suggest that short-day induced multiple long-distance FT-like tuber-inducing signals promote the robustness of sensitivity to short-day tuberization.This study advances our understanding of potato tuber formation and also provides cues that the mobility and relative transport of this protein to the apical meristem and stolons,as well as a dose-dependent effect on flowering and tuberization,presumably mediating pleiotropic effects of this FT ortholog.