Journal of Experimental Botany,Vol.57,No.13,pp.3395–3403,2006 Major Themes in Flowering Rearch Special
Issue
*To whom correspondence should be addresd.E-mail:coupland@mpiz-koeln.mpg.de
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the control offlowering time(Mouradov et al.,2002; Yanovsky and Kay,2003;Boss et al.,2004;Jack,2004; Putterill et al.,2004;Searle and Coupland,2004).Speci-fically in the photoperiodic control offlowering,models have been propod in which circadian clock control of gene transcription and post-transcriptional regulation of protein stability by light can combine to confer control of a regulating pathway that mediates the induction offlower-ing by day length(Searle and Coupland,2004).Recently, veral papers have been published aimed at the iden-tification of the mobile signal acting downstream of leaf-induction and triggeringflowering at the SAM.Bad on molecular and genetic evidence,a product of the FLOWERING LOCUS T(FT)gene,particularly the mRNA, has been implicated as this signal and the advances are reviewed here.
Physiological approaches towards the
identification of thefloral stimulus
Early in the20th century,Chailakhyan propod that the floral stimulus is a universal,unique,and specific hormone called‘florigen’(Chailakhyan,1937)but,despite exten-sive studies,such a compound was never isolated.Later, alternative theories were propod and,using photoperi-odic species that c
an be induced toflower by exposure to a single inductive photoperiod such as Sinapis alba, Lolium temulentum,Pharbitis nil,and Xanthium struma-rium,physiological study of thefloral transition led to the identification of veral putativefloral signals such as sucro,cytokinins,gibberellins(GAs),and reduced N-compounds,that are translocated from leaves to the SAM in respon to exposure to appropriate daylengths.Inter-estingly,the compounds induce in the SAM some of the cellular and molecular events typical offloral evocation (reviewed in Bernier and Pe´rilleux,2005).However,all the signals do not act,or are not all of equal importance in all species studied.For example,despite GAs being a primary factor in Lolium(King et al.,2001),they are not involved in Sinapis alba(Corbesier et al.,2004).This supported a theory known as the‘multifactorial control hypothes’which propod that veral factors,promoters and inhibitors,belonging to the class of nutrients and hormones,are involved in the control of the SAMfloral transition and that genetic variation as well as past and prent growing conditions result in different factor(s) becoming limiting in different genotypes or in a given genotype in various environments(Bernier,1988). Genetic control offlowering time in Arabidopsis In addition to the physiological studies,the genetic approach developed more recently in Arabidopsis,a quan-titative LD and facultative vernalization-requiring plant,allowed the discovery of many genes that controlflowering time(reviewed in Boss et al.,2004;Searle and Coupland, 2004;Bernier and Pe´rilleux,2005;Corbesier and Coupland, 2005).To i
dentify genes that control thefloral transition, mutants that showed accelerated or delayedflowering under different conditions,commonly known asflowering-time mutants,have been isolated(Redei,1962;Koornneef et al., 1991).The mutants were grouped according to their re-spons to various physiological conditions and then in-tegrated into genetic pathways to explain the control of flowering time.Four main promotive pathways were identified in Arabidopsis:the‘photoperiodic’,‘autonomous’,‘vernalization’,and‘GA’pathways.In addition to the four main pathways,less dramatic changes in ambient conditions also strongly influenceflowering time.For example,exposure to lower temperatures(16°C)delays flowering compared with the effect of growing plants at typical growth temperatures of20–24°C,and exposure to high ratios of far-red to red light associated with shading conditions acceleratesflowering(Bla´zquez et al.,2003; Cerdan and Chory,2003).Interestingly,all the pathways appear to interact in a complex manner and converge to reg-ulate genes that are often referred to as‘floral integrators’, SUPPRESSOR OF OVEREXPRESSION OF CO1(SOC1) and FT,that act upstream of the genes involved infloral morphogenesis such as APETALA1(AP1)and LEAFY (LFY)(Moon et al.,2003;Pineiro et al.,2003;Takada and Goto,2003)(Fig.1).FT encodes a protein with similarity to the RAF kina inhibitors of animals(Kardailsky et al., 1999;Kobayashi et al.,1999)whereas SOC1encodes a MADS box transcription factor(Borner et al.,2000;Lee et al.,2000;Samach et al.,2000).
Specifically in the photoperiodic control offlowering,a molecular hierarchy has been defined.Twoflowering-time genes specific to this pathway are GIGANTEA(GI)and CONSTANS(CO).The GI gene encodes a large protein that is prent in the nucleus and is highly conrved in Angiosperms and Gymnosperms but has no animal homo-logues(Fowler et al.,1999;Park et al.,1999)while CO encodes a B-box zincfinger protein that promotes tran-scription of downstreamflowering-time genes(Putterill et al.,1995;Robson et al.,2001).The biochemical function of GI is unknown,but gi mutations cau vere late flowering(Redei,1962),while overexpression of GI caus earlyflowering(Mizoguchi et al.,2005).GI regulates flowering time at least in part by the regulation of CO mRNA abundance;gi mutants contain less CO mRNA (Suarez-Lopez et al.,2001)while GI overexpressors show higher CO mRNA abundance.The abundance of GI and CO mRNAs is circadian clock regulated.Under LDs of16 h light,in which the genes promote earlyflowering,GI mRNA abundance peaks around10–12h after dawn, whereas CO mRNA abundance ris around12h after dawn and stays high throughout the night until the following dawn(Fowler et al.,1999;Park et al.,1999;Suarez-Lopez
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et al.,2001).CO mRNA abundance is therefore high when plants are expod to light at the end of a LD.CO ex-pression is also regulated at the post-transcriptional level, so that the cryptochrome and phytochrome A photo-receptors act at the end of the day to stabilize the CO protein(Valverde et al.,2004),whereas in darkness the protein is rapidly degraded,probably as a conquence of being ubiquitinated.Under SDs the CO mRNA is only expresd in the dark,and so the protein would be predicted never to accumulate.In agreement with the data,in wild-type plants FT is activated by CO under LDs,but not under SDs(Suarez-Lopez et al.,2001;Yanovsky and Kay,2002). Therefore,the combination of circadian clock-mediated regu-lation of CO mRNA abundance,and stabilization of CO protein by exposure to light can explain how CO promotes FT expression and,thus,flowering only under LDs.
The obrvation that CO is a major part of the molecular mechanism by which Arabidopsis discriminates between LDs and SDs suggests that CO is involved in the induction process and thus may act in the leaf to regulate the transition toflowering occurring at the apex.The CO mRNA is prent at very low abundance,but is expresd widely(Putterill et al.,1995;Simon et al.,1996;Takada and Goto,2003;An et al.,2004).Several recent obr-vations have shown that CO acts in the vascula
r tissue and not the meristem to promoteflowering.Specifically, triggering the expression of CO in the companion cells of the minor veins of the phloem of the mature leaves,using the promoter of a gene from melon encoding galactinol syntha,complemented the co-1mutation(Ayre and Turgeon,2004).Independently,An et al.(2004),using the phloem companion cell specific promoter of the Arabi-dopsis SUC2sucro-H+symporter gene(Truernit and Sauer,1995),obtained similar results and,in addition, showed that expression of CO from meristem-specific promoters had no effect onflowering.Therefore,CO appears to act specifically in the vascular tissue to regulate the synthesis or transport of a long-distance signal that initiatesfloral development at the apex.
The mechanism by which CO acts to promoteflowering in the phloem partially involves the FT gene.In wild-type plants,FT is expresd in the phloem,as detected using FT::GUS reporter constructs.Furthermore,FT expression is incread in the earlyflowering terminalflower2(tfl2) mutant,and in particular is expresd at higher levels in the vascular tissue,suggesting that CO may activate its target gene directly in the tissues(Takada and Goto,2003).In the phloem of SUC2::CO plants,FT mRNA abundance was incread in the phloem and ft mutations strongly suppresd the earlyflowering of SUC2::CO(An et al., 2004).Overexpression of CO in a ft-10mutant resulted in a lateflowering phenotype,similar to that of the co mutant under long photoperiod,suggesting that inacti
vation of FT suppress almost completely the signalling from CO and that FT is the major downstream target of CO(Yoo et al., 2005).Furthermore,expression of FT in the phloem from the SUC2promoter complemented the co mutation. However,in contrast to CO,FT promotedflowering when expresd in the meristem and the epidermal layer,as well as the phloem(An et al.,2004).Interestingly,among the 2000genes activated or represd in Arabidopsis leaves within an8h period after exposure to a single16h LD, only three genes responded differentially between WT and the co mutant and only one,FT,does not respond at all to the LD suggesting that FT is the major primary target of CO in leaves(Wigge et al.,2005).This is in agreement with the suppression of the earlyflowering phenotype of CO-overexpressors carrying an almost-null allele of FT whereas mutation of SOC1only partially suppresd early flowering(Yoo et al.,2005).
The data indicated that a major role of CO inflowering control is to activate FT in the leaf,and the obrvation that FT activatesflowering when expresd in the leaf or the SAM suggested that a product of FT might be transferred to the SAM to activateflowering.However,the data are also consistent with FT activating synthesis of afloral promoting compound in the leaf or SAM(An et al.,2004). Mode of action of FT
FT interacts with the bZIP transcription factor FD in yeast (Abe et al.,2005;Wigge et al.,2005).Mutatio
ns in FD cau lateflowering and the FD mRNA is detected in the shoot apex and FD expression incread with age in both SD-and LD-grown plants.FD appears restricted to
the
nucleus while FT is detected both in the nucleus and the cytoplasm(Abe et al.,2005).Mutation of FD
strongly suppresd the earlyflowering phenotype of35S::FT suggesting that FD and FT might interact in plants.The fusion of the VP16activation domain to FT supports this proposal:35S::FT-VP16induced extreme earlyflower-ing in a ft tfl1double mutant.The plants also had an incread expression level of AP1(Wigge et al.,2005).All the data support a model in which FT acts in the nucleus as part of a transcriptional complex with FD to activate the expression of the MADS-box transcription factor AP1in floral meristems(Abe et al.,2005;Wigge et al.,2005).In agreement with this hypothesis,the AP1mRNA level is reduced in the fd lfy double mutant,which also exhibits an inflorescence phenotype indistinguishable from the ft lfy double mutant suggesting that FD and FT are together involved in the up-regulation of AP1redundantly with LFY (Ruiz-Garcia et al.,1997;Abe et al.,2005).Independently, Wigge et al.(2005)reached the same conclusion and mapped a FD-respon element in the AP1promoter at the same location as the LFY binding site.Interestingly,AP1 expression is found in the vascular-rich region where FT is known to be expresd,is incread in35S::FD and this is FT dependent since it is abolished in a ft mutant. However,AP1is unlikely to be the FT target in the SAM involved inflowering control becau ap1mutants are not lateflowering(Page et al.,1999).On the contrary, mutation in SOC1results in lateflowering and the up-regulation of that gene in the SAM is one of the earliest events characteristic of thefloral transition(Borner et al., 2000;Samach et al.,2000).Mutation in FT delays strongly the expression of S
OC1in the SAM,even in plants overexpressing CO,and the direct expression of SOC1in the SAM is able to promoteflowering even in the abnce of CO or FT indicating that SOC1acts downstream of FT in the SAM(Searle et al.,2006).The u of plants overexpressing FT and carrying a SOC1::GUS reporter gene support this hypothesis(Yoo et al.,2005).A high GUS signal was obrved in the apex and only a weak increa was en in the vascular bundle of the cotyledons suggesting that SOC1is indeed downstream of FT,but the effect of FT on its activation ems restricted to the SAM.Finally,the activation of SOC1through FT appears to be FD-dependent since a mutation of either FD or FT reduced and delayed SOC1expression in the SAM(Searle et al.,2006).
A model that emerges from all the results can be summarized as follows.Photoperiodic induction occurs in the leaves and activates CO that stimulates FT expression. FT expression is not detected in the SAM but only in the vascular tissue suggesting that the FT mRNA or protein or both move to the SAM where FT interacts with FD to up-regulate SOC1within hours offloral induction.Later FD/FT act redundantly with LFY to activate AP1.Another possibility concerning thefirst events occurring in the leaves in respon to induction is that the activation of FT in the leaves results in the production of a condary signal that moves to the meristem where it induces FT expression.This multi-step regulation/signalling process might be required tofine-tune theflowering signal at the apex to preventflowering in non-optimal conditions.
FT as thefloral stimulus
FT is a direct target of CO that is expresd in the leaves and not in the SAM,but FT acts in the meristem to regulate gene expression suggesting that FT mRNA or protein moves to the SAM.Recent work suggests that FT mRNA might be the moving signal.Using a heat-shock inducible promoter fud to FT(Hsp::FT),Huang et al.(2005) showed that a single burst of FT expression in a single leaf of SD grown plants was able to triggerflowering in Arabidopsis,strongly supporting the major role played by FT in the leaf in the control offlowering time. Interestingly,the treated leaf could be removed from the plant7h after the heat treatment suggesting that the FT-dependent signal had left the leaf within this time window. This result gives some indication of the timing of the move-ment of thefloral stimulus out of the leaf in Arabidopsis and is compatible with the results of Corbesier et al.(1996) showing that the slowest component of the stimulus started to be exported from the leaves between8h and12h after the shift from SD to LD.Interestingly,in Lolium temulentum,King et al.(2006)also obrved a dramatic increa in the level of LtFT mRNA in the leaves within16 h of exposure to the critical daylength forflowering suggesting that LtFT RNA or protein could be part of the floral stimulus together with GAs in that species. Later,Huang et al.(2005)detected Hsp::FT mRNA in the apex of Arabidopsis plants(8-fold induction in the apex compared with2000-fold in the l
eaf)and this increa occurred24h after the treatment while the incread Hsp::FT mRNA level stopped after3h in the heat shock-treated leaf.In addition,9h after the end of the Hsp::FT mRNA burst,a strong increa of the endogenous FT mRNA occurred in the leaf and,at the same time in the apex.This9h lag pha suggests that FT does not directly stimulate its own transcription but intermediary factors might be implicated.Interestingly,the obrved continu-ous increa in endogenous FT mRNA would suggest that Arabidopsis,like some other species,once induced continues to produce theflowering signals which might be reprented here by FT mRNA.In Perilla,for example, once induced,a single leaf stably produces the stimulus, and can induceflowering in multiple shoots;repeated grafting of a single induced Perilla leaf quentially triggeredflowering in ven shoots over a period of97d (Zeevaart,1985).In Xanthium,Silene armeria,and Bryophyllum daigrementianum shoots induced toflower by grafting to donor shoots can themlves act as donors in
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subquent grafts(Zeevaart,1976).This suggests that the floral stimulus can act in the leaves of the species to trigger its own synthesis.However,this phenomenon may not be widespread,since other species,such as Perilla,do not exhibit indirect induction offlowering.On the other hand,in Arabidopsis,the fact that leaves can be removed from plants once the stimulus has been emitted(Corbesier et al.,1996;Huang et al.,2005)suggests that this species does not need the continuous synthesis of the stimulus and thus,it can be hypothesized that the increa of endogenous FT mRNA expression found by Huang et al. (2005)late after the end of the heat treatment may provide a mechanism for maintaining the induced state.
On the other hand,FT is a small protein of23kDa (Kardailsky et al.,1999;Kobayashi et al.,1999)and thus is below the size exclusion limit of plasmodesmata(Imlau, 1999).This small size suggests that the FT protein may move freely through plant tissues.Symplastic downloading of proteins from the sieve elements into the sink tissues of the apex through plasmodesmata has been propod (Ruiz-Medrano et al.,2001),suggesting that FT may move directly by this mechanism into apical cells and induce flowering.However,so far,the movement of FT protein during thefloral transition has not been demonstrated,but FT protein was found in the proteome of the phloem sap of Brassica napus collected from the inflorescences of plants 1week after theflower buds become visible by eye (Giaval
isco et al.,2006).Bad on high resolution2-D gel electrophoresis,they were able to detect600spots among which140could be reliably identified by MALDI-MS peptidefingerprints or by partial quence determination by mass spectrometry.Among the140proteins,they identified both the FT and the homolog of FT,TWIN SISTER OF FT(TSF),proteins.This paper is thefirst published work showing that the FT protein can indeed be detected in the phloem sap.However,the movement of the FT protein was not linked to thefloral transition since the sap was collected on plants which had already formed floral buds.The proteome analyd is most probably linked with the process of inflorescence/flower development.This is supported by the prence of latex proteins in the sap, some of which are involved in fruit ripening in melon (Aggelis et al.,1997)High-resolution phloem protein profiling is difficult to establish in Arabidopsis becau of its rotte habit and should be ud in other species able to be induced by a photoperiodic treatment allowing a synchronous shift from the vegetative stage toflowering. Such ideal species would posss a stem at the vegetative stage and belong to the same family as Arabidopsis to render possible the quence-bad identification of proteins.Although the genetic study of theflowering process is still incomplete in this species,Sinapis could be considered a species of choice for this type of work since the movement of thefloral stimulus out of the leaves has been precily timed in this caulescent species(Bernier et al.,1993).So far,whether the FT mRNA,the FT protein
or both move in wild-type plants remains to be established,
移动镜头as does the requirement of any movement forflowering.
In agreement with the latter hypothesis,Lifschitz et al. (2006)recently identified the tomato FT orthologue as SINGLE-FLOWER TRUSS(SFT),a gene regulating pri-
mary shootflowering time,sympodial habit,andflower morphology(Carmel-Goren et al.2003).sft mutants showed
lateflowering,indeterminate vegetative inflorescence
shoots with fewflowers each with a single enlarged pal. Constitutive expression of SFT under the35S promoter induced extreme earlyflowering in day-neutral tobacco
and tomato.In addition,when sft receptor shoots were grafted on35S::SFT donors,the receptor shoots produced
normalflowers,normal inflorescences and normal sympo-
沈阳周边
dial architecture suggesting that graft transmissible signals initiated by the SFT gene rescuedflowering time and morphogenesis defects in sft mutants.Interestingly,graft-transmissible SFT signals also substituted for the long-day
stimuli in Arabidopsis when expresd under the control of
a leaf-specific promoter,short-day stimuli in Maryland Mammoth tobacco,and light-do requirements in tomato
uniflora mutant plants.In tomato,SFT is expresd in the
leaf veins,shoot apices,stem,but not in roots,nor in the
SAM itlf where thefloral transition takes place.A G-box
factor called SPGB was also detected in the leaves of tomato
and the clost homologue of that gene in Arabidopsis is
FD suggesting a role for SFT in the leaves directly,poten-扫描频率
跖疣是怎么引起的
tially making it unnecessary for SFT RNA to travel toward
its interacting partners as implied for Arabidopsis.The localization of SFT RNA was performed on grafted
flowering plants and while they were able to detect the
RNA in the donor leaves,the authors could not detect it in
the receiver apices suggesting thatflorigen-like messages
in tomato are part of a downstream pathway triggered by
cell-autonomous SFT RNA transcripts.This is in contrast
to the conclusion that FT mRNA compris a transmis-
sible signal in Arabidopsis.经典名曲
Besides its role in the SAM,FT also regulates gene expression in the leaves
要求While it is clear that FT acts,together with FD,in the SAM
to up-regulate SOC1and AP1,Teper-Bamnolker and Samach(2005)give some support for a role of FT directly
捉萤火虫in the leaves.Normally,earlyflowering is associated with
small leaves.An association between ontogenetic changes
in vegetative metamers(heteroblasty)during plant de-velopment and the transition to the reproductive stage was
noted and documented a century ago,but the link between
the two process remained unclear(Goebel,1900;Jones, 1999).In Arabidopsis,LD reduce bothflowering time and
rotte leaf size.Most late-flowering mutants,including
ft and fd loss-of-function mutants,have larger rotte leaves.In early-flowering ecotypes expod to LDs,the
The quest forflorigen3399
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