Auxin at the Shoot Apical Meristem

15 Pages·2010·3.61 MB·English

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Auxin at the Shoot Apical Meristem
TevaVernoux,FabriceBesnard,andJanTraas
LaboratoiredeReproductionetDe´veloppementdesPlantes,ENS-Lyon,CNRS,INRA,UCBL,46Alle´ed’Italie,
69364Lyoncedex07,France
Correspondence:[email protected]
Plantscontinuouslygeneratenew tissues and organsthrough the activityofpopulationsof
undifferentiatedstemcells,calledmeristems.Here,wediscusstheso-calledshootapicalmer-
istem(SAM),whichgeneratesalltheaerialpartsoftheplant.Ithasbeenknownformanyyears
thatauxinplaysacentralroleinthefunctioningofthismeristem.Auxinisnothomogeneously
distributedattheSAManditisthoughtthatthisdistributionisinterpretedintermsofdifferential
geneexpressionandpatternedgrowth.Inthiscontext,auxintransportersofthePINandAUX
families,creatingauxinmaximaandminima,arecrucialregulators.However,auxintransport
isnottheonlyfactorinvolved.Auxinbiosynthesisgenesalsoshowspecific,patternedactivi-
ties,andlocalauxinsynthesisappearstobeessentialformeristemfunctionaswell.Inaddition,
auxinperceptionandsignaltransductiondefiningthecompetenceofcellstoreacttoauxin,
addfurthercomplexitytotheissue.TounravelthisintricatesignalingnetworkattheSAM,
systems biology approaches, involving not only molecular genetics but also live imaging
andcomputationalmodeling,havebecomeincreasinglyimportant.
Plantscontinuouslygeneratenewtissuesand The plant hormone auxin plays an instru-
organs through the activity of populations mentalroleinmeristembiologyandwediscuss
ofundifferentiatedstemcells,calledmeristems. hereitsroleinaparticularmeristem,theshoot
Becausemeristemscanmodulatetheiractivity, apical meristem (SAM) that generates all the
they provide the developmental flexibility that aerial organs including the floral meristems
allows plants to adapt their development in (Fig.1A–C).Inthiscontext,welimitourselves
reaction to the environment (reviews: Lyndon tothemeristemsinangiospermthathavebeen
1998;TraasandDoonan2001;AidaandTasaka studiedinmostdetail.
2006;Sablowski2007).
Distinctmeristemsexist.Apicalmeristems,
SHOOTAPICALMERISTEMSTRUCTURE
positionedatthetipoftheshootsandroots,ini-
ANDGENETICREGULATION
tiate aerial and underground organs, respec-
tively.Alongthestemsandroots,morediffuse AllSAMshaveanumberofbasiccharacteristics
secondary meristems are responsible for sec- in common (e.g., Lyndon 1998; Traas and
ondarythickeningofthesestructures. Doonan 2001). One of the most prominent
Editors:MarkEstelle,DolfWeijers,OttolineLeyser,andKarinLjung
AdditionalPerspectivesonAuxinSignalingavailableatwww.cshperspectives.org
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T.Vernoux,F.Besnard,andJ.Traas
A Inflorescences B D
Siliques
(fruits)
Lateral
Flower
meristem PZ CZ
L1
Primordia
L2
L3
Cauline C
leaf Stem cells
Rib
zone
Stem
Rosette
leaves
Stem
Figure1.TheshootapicalmeristemofArabidopsisthaliana.(A)Aerialpartofawild-typeplantoftheColumbia
ecotype(Col-0).TheSAMisresponsiblefortheproductionofrosetteleavesand,afterfloraltransition,forthe
productionofthestem,caulineleaves,lateralmeristems,andflowersoftheinflorescence.(B)Detailsofthetipof
theinflorescence,showingthehighlyorganizedpositioningofflowersaroundthemainaxis(aspiral).(C)A
dissected inflorescence meristem. Older flowers have been removed to expose the meristem surrounded by
young floral buds. (D) Longitudinal section of aninflorescence meristem showing the layered organization
(L1,L2,andL3celllayers).L1andL2arealsocalledthetunicaandL3tothecorpus.Thefunctionalzones
are also represented. At the meristem summit the central zone (CZ) contains the stem cells, whereas
primordiaareinitiatedintheperipheralzone(PZ).Theribzone(RZ)producestheinternalpartofthestem.
featuresisthepresenceofa surfacelayer,called resultingintheformationofarapidlyoutgrow-
the tunica (Fig. 1D). The tunica—itself often ingprimordium,delimitedbytheorganboun-
composedofseveralsublayers—coverstheinter- daryregion,wherecellexpansionisreduced.
nalcellsofthemeristem,collectivelycalledcor- Extensivegeneticscreensindifferentspecies
pus. The tunica layers are kept separate astheir haveidentifiedanetworkofregulatorsinvolved
cellsmostfrequentlydivideinanticlinalorienta- in meristem function, organ initiation, and
tions, causing daughter cells to remain in the outgrowth,whichisdescribedbrieflyinthefol-
samelayerastheirparent.Ifpresent,theinternal lowing paragraphs (for reviews: Traas and
tunicalayer—calledL2—usuallydisappearsdur- Doonan2001;AidaandTasaka2006;Sablowski
ingorganinitiation,whenL2cellsstarttodivide 2007;RastandSimon2008).Verysimilarmech-
inrandomdirections. anismsarefoundindifferentspecies,andhere
Superimposedonthislayeredorganization we discuss what is known on the best-studied
is a partitioning into zones (Fig. 1D). These model,Arabidopsisthaliana.
zones,characterizedbysometimessubtlecyto- A central role in shoot meristem main-
logicaldifferences,havewelldefinedfunctions. tenance is played by the transcription factor
Atthemeristemsummitasmallgroupofslowly WUSCHEL (WUS) as in corresponding mu-
dividing cells, called the central zone (CZ), tantsaSAMisinitiatedbutarrestsafterhaving
ensuresatruestemcellfunctionandisrespon- produced a few organs (Laux et al. 1996). Its
sible for meristem maintenance. The growth precise targets and regulators are largely
rates at the meristem summit usually differ unknown,butithasbeenwellestablished that
considerablyfromthoseattheperipherywhere it interferes with hormone signaling cascades,
cells accelerate their proliferation rates. It is in in particular cytokinins (Leibfried et al. 2005;
thisperipheralzone(PZ)thatnewlateralorgans Gordonetal.2009).WUSisinvolvedinaneg-
are initiated: a restricted number of founder ativefeedbackloopwiththeCLAVATAreceptor
cellsincreasetheirproliferationandgrowthrates kinase signaling cascade (CLV), whose activity
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AuxinattheShootApicalMeristem
A C
P1
Central zone y Primordium
P3 n dar
e Auxion ? Boun DR5rev
mptivntrati Auxine xpression
ue
sc
Precon Pin1
L1
P2 PID
CLV3 Auxin
B
CUC2 signaling
P1 (MP/ARF5)
WUS STM
P3
STM
ANT
P4 AS1 LFY
?
Auxin biosynthesis
(YUCCA genes) Organ Organ
outgrowth
identity
(CYCD)
and
patterning
P2
Figure2. From dynamic transport to patterning: Auxin and organogenesis at the shoot apical meristem of
Arabidopsis thaliana. (A) Immunodetection of PIN1 efflux carrier in the L1 (top view). The image was
obtained by confocal microscopy. Note the subcellular polarized localization of the auxin transporter in
mostcells.Thelocalizationsuggeststhatauxinaccumulatesintheseyoungorganprimordia(namedP1,P2,
andP3fromtheoldesttotheyoungestorgan).AdaptedfromdeReuilleetal.(2006).(B)Expressionofthe
syntheticDR5rev::GFPreporterintheinflorescencemeristem(topview).Projectionofserialopticalsections
obtainedbyconfocalmicroscopy.ThegreencorrespondstotheGFPandtheredtotheautofluorescenceof
meristematic cells. DR5 expression in young emerging primordia indicates activation of auxin induced-
genes.(C)Schematicrepresentationoftheroleofauxinduringorganogenesisintheinflorescencemeristem
ofArabidopsisthaliana.Seetextfordetails.
limitsthesizeofthepoolofstemcells(seeFig. STMinthedevelopingprimordia(Byrneetal.
2Cforexpressionpatternsofthegenes). 2000). The meristem either generates leaves
Attheperipheryofthemeristem,theinitial and lateral meristems or flowers. The identity
recruitmentoftheorganfoundercellsappears of the lateral organs produced depends on the
to be the result of two antagonistic processes. activityofLEAFY(LFY),which,byinteracting
First,thehomeodomainproteinSHOOTMER- with different transcription factors such as
ISTEMLESS (STM), in combinationwith sev- WUS or AGAMOUS (AG), plays a major role
eral members of the so-called CUP SHAPED in plant and flower architecture (Lenhard
COTYLEDON (CUC) family of transcription etal.2001;Lohmannetal.2001).
factors, defines meristematic identity, prevent- Thepatterningoftheflowermeristemitself
ing cells from being recruited by the young largely depends on transcriptional regulation
organs (e.g., Aida et al. 1999). Therefore, as a (for reviews: Krizek and Fletcher 2005; Bow-
neworganisinitiated,thesemeristematiciden- manandFloyd2008).Thisincludesgenesthat
tity genes are switched off in Arabidopsis. This control organ identity, organ number, organ
involves the transcription factor ASYMMET- boundaries, and symmetry. In Arabidopsis, for
RIC LEAVES1 (AS1), which maintains the example, APETALA1 (AP1), CAULIFLOWER
repressionofmeristemidentityfactorssuchas (CAL),PISTILLATA(PI)SEPALLATA1(SEP1),
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T.Vernoux,F.Besnard,andJ.Traas
and APETALA3 (AP3) direct the development homogeneously distributed at the SAM and it
of sepals and petals. Other relevant genes are is thought that this distribution is interpreted
upstream regulators of these identity genes in terms of differential gene expression and
(e.g., UNUSUAL FLORALORGANS and LFY) patterned growth. Because the distribution of
butalsogenesthatcontrolthenumberofsepals auxinislargelyunderthecontrolofauxintrans-
and petals formed in each flower (PERIAN- porters, we first discuss auxin transport at the
THIA, WIGGUM), genes required to establish shootapex.
organboundariesinallshootmeristems(CUC,
genes),andgenesthatparticipateinorganpat-
terning(KANADIs,YABBYs,andthePHABU- AUXINTRANSPORTANDSAMFUNCTION
LOSA/PHAVOLUTA/REVOLUTAgroup).
AuxinInfluxandEffluxCarriersControlAuxin
Once the founder cell populations of the
DistributionattheSAM
organs have been identified, they grow out.
Our knowledge on the genetic regulation of Okada et al. (1991) identified an Arabidopsis
growthcontrolismorelimitedandthevarious mutantcalledpin-formed1(pin1)inreference
signalsthatintegrateandcoordinatecellprolif- to its needle-like inflorescence stem, unable to
erationandexpansionremainlargelyunknown. initiate flowers. Identification of the gene
Some of the connections between the cellular showed that it encoded atransmembrane pro-
processes and transcriptional regulation have tein (Galweiler et al. 1998) and there is now
beenidentified.AINTEGUMENTA(ANT),for overwhelming evidence that the PIN1 protein
instance,isstronglyexpressedintheoutgrowing isthefoundingmemberofafamilyofauxincar-
organs and acts at least partially via regulating riers that transport auxin across membranes
genesinvolvedincellproliferationlikeCYCLIN (for review: Petrasek and Friml 2009). More
D(MizukamiandFischer2000).Inpetals,sev- particularly,PIN1andcloselyrelatedfamilyme-
eralgenescontrolgrowthbyaffectingcellpro- mbers are involved in auxin efflux out of the
liferation and/or cell expansion (Anastasiou cells. PIN1 is strongly expressed in the SAM,
and Lenhard 2007). Some of these genes (i.e., whereitismainlyfoundinthetunicaandvascu-
JAGGED,ANT,andARGOS)affectpetalgrowth lartissues(Fig.2A).TheactivityofotherPIN-
bypositivelyregulatingcellproliferation(Mizu- membersattheSAMhasnotbeenanalyzedin
kamiandFischer2000;Huetal.2003;Dinneny detail. Expression studies show that at least
et al. 2004), whereas other genes (BIGBRO- PIN2and7areactiveintheshootapexbutsin-
THER, KLUH, and DA1) control final organ glemutationsinthesegenesdonotsignificantly
sizebynegativelyregulatingtheduration/period perturbmeristemfunction(Mulleretal.1998;
ofcellproliferation(Dischetal.2006;Anastasiou Frimletal.2003).Thepin1phenotypecanlo-
etal.2007;Lietal.2008).BIGPETAL,whichis callybecomplementedbylocal,externalappli-
regulateddownstreamoftheflowerorganiden- cationsofhighconcentrationsofauxin,which
titygenes,limitspetalgrowthbycontrollingcell areabletoinducetheformationofflowerbuds
expansion(Szecsietal.2006). (Reinhardtetal.2000).Theseapplicationsindi-
Inadditiontotranscriptionalcontrol,post- catethathighauxinconcentrationsarerequired
transcriptional regulation by microRNAs (mi- for the initiation of a new organ and, impor-
RNAs) plays an important role in meristem tantly, that PIN transporters are required for
function. These include, for example, the the creation of such auxin maxima. Direct
miR164 and miR156/157 families, which evidence demonstrating the existence of auxin
respectively target the CUC and SPL genes maximaattheperipheryofthemeristemarestill
(Peaucelleetal.2007;Wangetal.2009). lacking because of the absence of a true auxin
How is this complex regulatory network sensor.However,thisscenarioisfurthercorro-
coordinated?Ithasbeenknownformanyyears borated by the activation of the synthetic DR5
that auxin plays a central role in meristem auxin reporter in the young organs (Fig. 2B).
function and organ formation. Auxin is not DR5 is composed of a fusion of a synthetic
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AuxinattheShootApicalMeristem
promoter consisting of several auxin response etal.2003).Altogether,theavailabledataindi-
elements(AuxRE)andaminimal35Spromoter cate that the formation of local auxin maxima
withareportergene(Sabatinietal.1999).The mainlydependsontheactionofPINexporters
AuxREs corresponds to the DNA sequences atthemeristemsurface.AUXandLAXproteins
bound by the AUXIN RESPONSE FACTORS wouldfacilitateorganpositioning,probablyby
(ARFs;seelaterformoredetails)andexpression guaranteeingasufficientsupplyintheL1layer.
ofthereportergenethussuggestsactivationof
auxin-induced genes during the earlier stages
UsingModelingtoExplorePolarAuxin
of organ initiation (Benkova et al. 2003; de
TransportRegulation: Canalizationor
Reuilleetal.2006;Smithetal.2006).Inaddi-
Up-the-Gradient?
tion, the polarity of the PIN transporters has
beenanalyzedtoidentifytheputativedirection ThedistributionofPINatthemeristemsurface
of auxin fluxes. This has confirmed the hy- is complex and it is not obvious from simple
pothesis that auxin fluxes are directed toward visual inspection what the predicted auxin
theyoungprimordia(Benkovaetal.2003;Rein- fluxeswouldbe.Therefore,asetofcarefullocal-
hardtetal.2003). izationstudieswereperformedandtheproper-
Secondsetsoftransportersassociatedwith tiesofthecellulartransportnetworksanalyzed
auxin distribution at the SAM are the AUX/ using computer simulations (de Reuille et al.
LAXinfluxcarriers.AUX1,thefoundingmem- 2006). This confirmed that PIN directs auxin
berof the gene family (Bennett et al. 1996), is to the sites where young primordia are being
expressed in the L1 surface layer of the shoot formed. The same computer simulations also
apical meristem (Reinhardt et al. 2003). The identifiedadditionalpropertiesofthetransport
protein seems to be evenly localized over all network,andinparticularastillundefinedrole
membranes, indicating that the protein is not for the meristem summit in auxin redistri-
involved in the creation of hormone fluxes. butionwasproposed(deReuilleetal.2006).
Instead, it might rather concentrate auxin at Whatcoordinatesauxinfluxesinthemeri-
the meristem surface. Somewhat surprisingly, stem? The complex and dynamic patterns of
its loss of function has only a relatively mild PINdistributioncouldsuggestveryintricatere-
phenotype in the shoot, including perturba- gulatorymechanisms.Computermodels,how-
tionsinphyllotacticpatterns.TheAUX1sequ- ever, have shown that the actual basis of the
ence shows a high degree of similarity with observedtransportpatternscouldbeverysim-
three other sequences in the Arabidopsis gen- ple. Jo¨nsson et al. (2006) and Smith et al.
ome, named LAX1, LAX2, and LAX3 (Like (2006) proposed models where cells check the
AUX1) (Swarup and Bennett 2003), but even auxin concentrations in their direct environ-
thequadruplemutantisstillabletoproducea mentandsubsequentlypumpauxintowardnei-
viable, fertile plant albeit with important ghborswithahigherconcentration,i.e.,against
changes in its architecture (Bainbridge et al. thegradient.Becausethesepatterningprocesses
2008).Itisthereforeconceivablethatotherpro- require the interaction of hundreds of cells, it
teins,suchascertainP-glycoproteintransport- is impossible to estimate on a purely intuitive
ers(PGPs),whichhavealsobeenassociatedwith basisifaparticularscenarioisplausibleornot.
auxinimport,sharearedundantfunctionwith Therefore, computational modeling was used
themembersoftheLAXfamilyinthemeristem asapowerfulmeanstotestthishypothesis.Int-
(Forreview:Boutteetal.2007).Anotherclueon erestingly, these models showed that transport
thepreciseroleofAUX1comesfromadouble “againstthegradient”issufficienttoreproduce
pin1/aux1 mutant. Auxin applications do not realisticPIN1patternsandtogeneratedifferent
inducesingleflowersontheapexofsuchadou- types of phyllotactic patterns (Jonsson et al.
blemutant,butratherverylarge,fusedorgans, 2006;Smithetal.2006).
suggestingthatsomehowAUX1isrequiredfor It should be noted that, although these
therestrictionoforganboundaries(Reinhardt models show that the mechanism is plausible,
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T.Vernoux,F.Besnard,andJ.Traas
they by no means provide absolute proof. processes. Indeed, although the exact mecha-
Indeed, other models are able to explain the nismsthatcontrolthedynamicsofPIN1polar
distributionofauxintransporters.Anotherhy- localization are still unknown, accumulating
pothesis,forinstance,wasbasedonthepioneer- evidence indicatesthat it depends on multiple
ingworkofSachswhoproposedtheexistenceof processes, such as membrane traffic and cyto-
apositivefeedbackbetweenfluxandtransport skeletonorganization(forreview:Kleine-Vehn
(Sachs 1969). It was subsequently shown that and Friml 2008). A striking set of data also
this mechanism is able to amplifysmall fluxes points at the importance of phosphorylation
and can potentially create canals of auxin bet- and the PINOID (PID) serine-threonine pro-
ween hormone sources and sinks. A range of teinkinase(Christensenetal.2000;Benjamins
experimentssupportsthecanalizationhypoth- et al. 2001). Arabidopsis mutants perturbed in
esis, at least in the inner tissues of the plant thePIDgenehavepin-likeinflorescences.Inter-
whereitcanaccountfortheformationofvenat- estingly,thisgoesalongwithanapical-to-basal
ion patterns (Scarpella et al. 2004; Saueret al. shift in PIN1 polarity (Friml et al. 2004).
2006; Scarpella et al. 2006). The existence of Accordingly, in the root, ectopic expression of
canalization would imply the coexistence of PID induces a basal-to-apical shift in PIN2
two radically different mechanisms for PIN andPIN4polarities.Theresultssofarindicate
allocationtothemembrane,onebasedonflux that the PID kinase controls PIN localization
sensing(intheinner tissues)andtheotheron via direct phosphorylation of the transporter
local concentration sensing (at the meristem (Michniewiczetal.2007).Interestingly,theki-
surface). Therefore, Stoma and colleagues nasedoesnotseemtoaffectthepolarizedlocal-
(2008)testedwhethercanalizationcouldpoten- ization per se, but rather decides on what
tially also account for the behavior of auxin particular membrane the carrier will be local-
transporters at the shoot apical meristem sur- ized. Lee and Cho (2006) also showed that
face. Using a dedicated computer simulation PID may facilitate trafficking of PIN to the
tool,theyshowedthatthiswasindeedthecase, plasma membrane, suggesting that it may not
thusprovidingaunifyingconceptforthecontrol only affect the polarity of PIN1 but also the
ofauxindistributionintheplant (Stomaetal. dynamics of polarity changes in meristematic
2008).Morerecently,Bayerandcoworkerstested cells(LeeandCho2006).Antagonistregulation
the alternative scenario where both modes of ofPIN1byPP2Aphosphatasehasbeenshown
auxintransport(i.e.,up-the-gradientandcanal- (Michniewicz et al. 2007) and PID activity it-
ization) coexist. One or the other mechanism selfdependsonitsautophosphorylationstatus
wouldprevail,dependingontheabsoluteauxin (Zegzouti et al. 2006). Precise modulation of
concentrations (Bayer et al. 2009). This model both PIN1 and PID phosphorylation status is
also reproduced realistic distributions of PIN thusanessentialregulationlevelofPIN1local-
proteins.Furtherexperimentsarenowrequired ization in the meristem. It was recentlyshown
to distinguish between flux-based polarization that PINs are initially delivered to the plasma
and other hypotheses. Although these models membraneinanonpolarmannerandthattheir
remaintobetested,notablythroughtheidenti- polarityisestablishedbysubsequentendocytic
ficationofthemolecularmechanismcontrolling recycling (Dhonukshe et al. 2008). It is thus
PIN1polarity,theyshow thatverysimplelocal possiblethatPIDcouldbeinvolvedindirecting
behaviors of individual cells can generate the PIN1tospecificmembranes,thusparticipating
complexoverallpatternsweobserve. in controlling the dynamics of the PIN1 net-
workinthemeristem.
Other genes are involved in the PID path-
OrientingAuxinTransportattheSAM:
way,likeNPY1/ENP/MAB4,aNPH3-likepro-
TheRoleofPID
teinfirstidentifiedasanenhancerofPIDduring
Concepts like canalization are still relatively cotyledon development (Treml et al. 2005;
abstract and could represent a combination of Furutani et al. 2007). Mutations in NPY1
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AuxinattheShootApicalMeristem
induce pin-like inflorescence in the yuc1 yuc4 We will therefore focus mainly on its impor-
(yucca) double mutant affected in auxin bio- tanceinshootapicalmeristemfunction.
synthesis ([Cheng et al. 2007b]; see part 4 for
discussion on the role of the YUCCA genes).
LocalAuxinBiosynthesisisEssentialforShoot
NPY1 is expressed in the L1 at the meristem
ApicalMeristemFunction
and accumulates in the young organs (Cheng
etal.2007b;Furutanietal.2007).Theinactiva- IAAbiosynthesisoccursmostlythroughatryp-
tion of NPY1 together with two of its closest tophan (Trp)-dependent pathwayandinvolves
homologsalsocompletelyabolishesorganini- four parallel and partly interdependent path-
tiation at the SAM (Cheng et al. 2008). ways named after the main intermediates: the
AlthoughtheexactroleofNPY1anditshomo- indole-3-pyruvicacid(IPA),tryptamine(TAM),
logs still needs to be established, these results indole-3-acetaldoxime (IAOx), and indole-3-
suggestthattheNPYproteinmediatesorfacil- acetamide(IAM)pathways(WoodwardandBar-
itatesPIDactiononPIN1localization. tel2005).TherecentidentificationoftheYUCCA
ItisinterestingtonotethatPIDisexpressed (YUC)familyofauxinbiosyntheticgenesencod-
atahigherlevelinthefrontiersthatseparateorg- ing flavin monooxygenases regulating the TAM
ansfromthemeristem(Furutanietal.2004).It pathwayhasbeeninstrumentalindemonstrating
ispreciselyattheseboundariesthatPINunder- that auxin biosynthesis is not homogenous in
goes important relocalizations (Heisler et al. agiventissueandthatafinecontrolofauxinbio-
2005). PID could therefore play a decisive role synthesisintheshootapexplaysakeyroleinthe
inorganseparation. regulationoftheactivityoftheshootandfloral
meristems.
TheYUCfamilyhas11membersandYUC1,
itsfoundingmember,wasisolatedbasedonan
THEROLEOFAUXINMETABOLISMATTHE
increase in hypocotyl length in an activation-
SAM
tagging screen (Zhao et al. 2001). The yuc1-D
Althoughtheroleofpolarauxintransportdur- gain-of-functionmutantdisplayedtypicalaux-
ing developmental processes, notably in the inoverproductionphenotype,suchasepinastic
shoot meristem, has been extensively studied, cotyledons and long hypocotyl, short roots
littleattentionhasbeenpaidtotheroleofauxin and increased apical dominance. Based on in
metabolism until recent years. However, local vitro biochemical assays and on resistance of
concentrationsofauxinintheshootmeristem the mutant to the toxic tryptophan analog,
areexpectedtobeunderthecontrolofthecom- 5-methyl-tryptophan, it was proposed that
binedactionofpolarauxintransportandauxin YUC1 catalyzed the oxidization of tryptamine
biosynthesis, but also of auxin catabolism and to N-hydroxyl-tryptamine, a rate-limiting step
conjugation. Auxin metabolism is still largely inthetryptophan-dependentauxinbiosynthe-
underinvestigationanditscomplexityprovides sistryptaminepathway(Zhaoetal.2001).The
one historical reason for focusing primarily importanceofYUC-dependentlocalauxinbio-
on polar auxin transport. Nevertheless, recent synthesisinorganinitiationwasrevealedbythe
studies have identified mutants in key biosyn- analysisofmultipleloss-of-functionyuccamu-
thetic enzymes that highlight the importance tants in Arabidopsis (Cheng et al. 2006; Cheng
of local auxin biosynthesis in the shoot meri- etal.2007a).Althoughnoneofthesinglemu-
stem and underline the need to consider this tantsshowedobviousphenotypes,severaldou-
parameter when studying the role of auxin in ble and triple mutant combinations and the
theshootmeristemandmoregenerallyinplant quadruple mutant of yuc1, yuc2, yuc4, and
development. yuc6 displayed strong inflorescence and floral
Severalexcellentreviewshavedescribedour developmental defects as well as alterations of
current knowledge on auxin metabolism (e.g., leafvasculature(Chengetal.2006).Thesefloral
Ljungetal.2002;WoodwardandBartel2005). and vascular phenotypes are reminiscent of
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T.Vernoux,F.Besnard,andJ.Traas
those observed in the pin1, pid, and ettin/arf3 be involved in the local regulation auxin
mutants (Okada et al. 1991; Bennett et al. biosynthesis.
1995; Sessions et al. 1997). In the quadruple At least two nonredundant auxin biosyn-
yuc mutant, pin-like structures were also thesispathwaysarethusimplicatedinorganini-
observed,indicatingthatthesefourYUCgenes tiation at the shoot or floral meristem. The
areinvolvedinfloraldevelopmentfromtheini- phenotype obtained on inactivation of these
tiationofthefloweronwards.Theanalysisofthe pathways indicates that they are necessary to
expressionpatternoftheYUCgenesfurthersug- control locally auxin homeostasis at the shoot
gested that they regulate flower initiation and meristem andthat auxin transportis probably
developmentthroughthespatialandtemporal nottheonlylimitingmechanismforgenerating
control of auxin biosynthesis. Direct evidence the spatial variations in auxin concentration
that the developmental defects in the multiple implicatedinlateralorganinitiation.
yuc mutant were caused by a local decrease in
auxinbiosynthesiswasshownbycomplement-
IntegratingAuxinMetabolismandTransport
ingtheyuc1yuc4doublemutantwithabacterial
attheShootMeristem:AChallengeforthe
auxinbiosynthesisgeneunderthecontrolofthe
Future
YUC1 promoter (Cheng et al. 2006). In addi-
tion, Cheng et al. (2007) showed through the Theresultsdiscussedinthepreviousparagraphs
analysis of combinations of mutants in yuc1, suggestthattheintegratedactionofbothauxin
yuc4, yuc10, and yuc11 that auxin synthesized transport and metabolism define where and
by the YUC proteins is also necessary for leaf when auxin will accumulate. Ljung et al.
development. (2001) showed the existence of a feedback
ThetemporalandspatialregulationofYUC mechanismbetweenbiosynthesisandtransport
genesthusappearstoprovideamechanismfor bymeasuringauxinconcentrationandsynthe-
fine-tuning the concentration of auxin during sisratesontreatmentofyoungArabidopsisroots
organ initiation and development at the shoot with NPA (Ljung et al. 2001). Cheng et al.
apex. Mutants in YUC homologs of petunia, (2007)suggestthatsuchafeedbackmechanism
maize,andrice(Tobena-Santamariaetal.2002; is also active in the shoot meristem because
Fujinoetal.2008;Gallavottietal.2008)exhib- combinations of the pin1 mutation with yuc1
ited similar developmental defects, indicating andyuc4enhancespin1defects,blockingalmost
thattheregulationoftheTAMpathwayforauxin completelyleafandflowerinitiation.Asimilar
biosynthesisisaconservedmechanismformod- phenotype is obtained by combining aux1
ulatingauxinintheangiosperms.Theidentifica- with yuc1, yuc2, yuc4, and yuc6 (Cheng et al.
tionofTRYPTOPHANAMINOTRANSFERASE 2007a).Cross-regulationsbetweenauxintrans-
of ARABIDOPSIS 1 (TAA1), a long-predicted port and biosynthesis could be a key feature
key enzyme in the indole-3-pyruvic acid (IPA) explaining the robustness of auxin gradients
pathway,anditsparalogs(TRYPTOPHANAMI- (Weijersetal.2005)anditwouldbeparticularly
NOTRANSFERASE RELATED or TAR) has re- instructivetoanalyzetheeffectofincorporating
cently shown that spatial regulation of auxin auxin production and inactivation on the
biosynthesisthroughthissecondTrp-dependent stability of phyllotaxis models. Further work
pathway might also play a similar role during on the spatial control of auxin biosynthesis
development (Stepanova et al. 2008; Tao et al. willprobablybeessentialinthiscontext.
2008). Stepanova et al. (2008) notably found
that the double taa1/tar2 mutant displays a
FROMAUXINSIGNALTRANSDUCTIONTO
severe reduction of growth of the aerial tissues
MORPHOGENESISATTHESHOOTAPEX
andalterationsofflowerdevelopment,suggest-
ing a role in floral organ initiation. As for the The action of auxin does not only depend on
YUC genes, TAA1 expression is restricted to theregulationofitssynthesisortransport,but
specific parts of the flowerand the gene might thecompetencetoreacttoauxinseemsalsoto
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AuxinattheShootApicalMeristem
becontrolledintimeandspace.Wenowdiscuss BecauseMP/ARF5isonlyexpressedatthemeri-
the regulation of auxin signal transduction in stemperiphery(HardtkeandBerleth1998),the
the shoot meristem before considering how competencefororgan initiation at the periph-
auxininteractswiththegenenetworksunderly- ery of the meristem thus depends, at least in
ingorganoutgrowth. part, on a spatial modulation of auxin signal
transduction.Despiteanextensivegeneticanal-
ysisoftheARFandAux/IAAfamilies(Okush-
SpatialModulationofAuxinSignal
ima et al. 2005; Overvoorde et al. 2005), no
TransductionisInvolvedinMeristem
other ARF and no Aux/IAA has been impli-
Function
cated in shoot meristem function, although
Bothbiochemicalandgeneticapproacheshave indirect evidence suggests that several ARFs
identified the members of the Aux/IAA and couldbeinvolved(Hardtke2004;Pekker2005;
ARF family of transcriptional regulators as Mallory 2005). The lack of genetic evidence
major effectors of auxin signal transduction could be because of the demonstrated redun-
(for review: Leyser 2006). There are 29 Aux/ dancy between these transcription regulators
IAA genes that encode mostly short-lived (Ellis 2005; Pekker 2005; Okushima 2005). It
repressors of auxin-inducible genes. The 23 willthereforebeimportanttogeneratemultiple
ARF proteins can be either activators (Q-rich mutantcombinationsforARFsaswellasdom-
ARFs)orrepressorsoftranscription.Aux/IAA inantnegativemutantinallAux/IAAmembers.
andARFproteinsareabletoformhomo-and Togetherwithanexplorationoftheexpression
heterodimersbothwithinandbetweenthefam- patternsofthesetwogenefamilies,thisshould
ilies.TheinstabilityoftheAux/IAAsisintrinsic give valuable information of the role of signal
to the so-called domain II, which interacts transduction modulation in the dynamics of
directlywithSCF-likeubiquitinproteinligases auxinresponsesinthemeristem.
harboring the TIR1 F-box protein or one of
the three Auxin-related F-box (AFB) proteins
InductionoftheMorphogeneticProgramby
(Dharmasiri et al. 2005a; Dharmasiri et al.
AuxinintheShootMeristem
2005b; Kepinski and Leyser 2005; Tan et al.
2007).TIR1andtheAFBsactdirectlyasauxin Theanalysisofcellidentityinthemeristemof
receptors and their activation leads to auxin- thepin1mutantgavethefirstcluesconcerning
dependentdegradationofAux/IAAs.Thecur- the role of auxin in the control of cell identity
rent simplified model for auxin transduction atthemeristem(Vernouxetal.2000).Although
is that Aux/IAAs dimerize with the Q-rich meristem structure and maintenance were not
ARFs.Thesecomplexesbindtoauxin-inducible severely affected in the mutant, major altera-
genes, thus preventing transcription. By pro- tionsoccurredattheperipheryofthepin1mer-
moting the degradation of the Aux/IAAs by istem, where organ initiation should occur. A
theSCF,auxinwouldallowtheARFstoactivate ring-like domain expressing twoearly markers
transcription of target genes by binding to the of organ initiation, LFYand ANT, but also the
AuxREspresentintheirpromoter. boundary marker CUC2 was observed around
Whenpin1meristemsaretreatedwithexog- the central zone. This showed that the zone at
enousauxin,allthecellsattheperipheryofthe the meristem periphery has a hybrid identity
meristemarecompetenttorespondtoauxinto in pin1. These results also implied that auxin
initiateorgans(Reinhardtetal.2000).However, levels control the identities of the cells at the
auxinisnotabletoinduceorgansatthecentral peripheryofthemeristem,thusimpactingorgan
zone of the meristem. Mutation in MONOP- separation,positioning,andoutgrowth.Analy-
TEROS/ARF5 induces a phenotype similar to sis of the genetic interactions between PIN1
pin1andblockstheabilityofthemeristematic and MP/ARF5 further suggested a differential
cells to respond to exogenous auxin (Hardtke regulation of the CUC genes by PIN1 and MP
and Berleth 1998; Reinhardt et al. 2003). duringembryogenesis(Aidaetal.2002).Taken
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T.Vernoux,F.Besnard,andJ.Traas
together,thesedatasuggestamodelwhereaux- lateralorgans(BowmanandFloyd2008).Mu-
in, through PIN1 and MP, could regulate pat- tationsinARF3/ETTINwereisolatedassecond
terning at the meristem partially through the site suppressors of ectopic KAN expression
controlofCUCgeneexpression(Fig.2C). (Pekkeretal.2005).Furthergeneticandexpres-
More recently, live-imaging of Arabidopsis sion analysis indicated that ARF3/ETT and
inflorescencemeristemsalloweddetailedanaly- ARF4 mediate the KAN pathway but that
sisofthesequenceofgeneactivationinrelation KAN does not regulate ARF3 and ARF4 acti-
toauxintransportandresponses(Heisleretal. vation in lateral organs. ARF3 and ARF4 are
2005). These authors used PIN1-GFP trans- expressedduringtheearlystagesoforganinitia-
lational fusion in conjunction with the auxin- tion (Sessions et al. 1997; Pekker et al. 2005),
responsive DR5::VENUS marker, to show that indicating that modulation of auxin activity is
the convergence of PIN1 proteins toward the likely cooperatively involved with KAN in set-
site of the new organ is likely the first event ting upadaxial/abaxialorgan polarity(Pekker
of organogenesis and is immediately followed etal.2005).
byauxin-inducedtranscription. Bycombining
multiple GFP-protein fusions, their work fur-
INTEGRATIONOFHORMONESIGNALING
ther suggests that the KNOX gene STM and
ATTHESHOOTAPEX
CUC2 are down-regulated in the early pri-
mordia, whereas they are up-regulated in the As in other tissues, SAM function relies on
frontiers in response to specific auxin concen- the combinatorial action of several hormones
trationand/orresponses.Thiselegantanalysis (Shani et al. 2006). We review here briefly the
notonlylinkedauxinpatternsatthemeristem best-characterizedhormoneinteractionsinvol-
todevelopmentalpatterningbutalsosupported vingauxin.
the proposed role of PIN1 in CUC regulation Studies on plant tissue regeneration have
discussed earlier. In addition to the effects on shownthatahighcytokinin(CK)toauxinratio
STM,Hayetal.(2006)demonstratethatBRE- can trigger the initiation of shoot meristems
VIPEDICELLUS (BP), another KNOX gene, is fromundifferentiated callus butthe molecular
ectopically expressed in the leaves of a pin1 basisforsuchinteractionsintheshootarejust
mutant, whereasthe bp mutation slightlysup- startingtoemerge(Gordonetal.2007;Gordon
presses both pin1 and pid mutants (Hay et al. et al. 2009). In the last few years, it has been
2006). These datasupport a scenarioinwhich shown that high CK signaling is essential for
theexpressionofKNOXgenesisfirstrepressed maintenance of the meristem through a direct
in the incipient primordium by an auxin- effectonstemcellactivity.However,theidenti-
dependentmechanism,andislatermaintained fication and characterization of the aberrant
inarepressedstate viatheactionofregulators pyllotaxy1 (abphyl1) mutant of maize suggest
suchasAS1(Fig.2C). that CKs contributes to organ initiation toge-
Auxin is not only involved in organ initia- ther with auxin. This mutant shows decussate
tion but is also associated with the establish- phyllotaxis, whereas distichous arrangement are
ment of symmetry. In vivo imaging suggests usuallyobservedinwildtypeplants(Jacksonand
that auxin maxima precede and might control Hake 1999). ABPHYL1 encodes a cytokinin-
theestablishmentofadaxial/abaxialsymmetry inducibleRESPONSEREGULATOR,aprimary
byFILAMENTOUSFLOWERandREVOLUTA cytokininresponsegenelikelytoactinanegative
(Heisleretal.2005).Sucharolewasfurthersup- feedbacklooptolowerCKsignalingintheleaf
ported by genetic studies, identifying ETTIN/ primordia(Giulinietal.2004).Inabphyl1mu-
ARF3andARF4asregulatorsoforganasymme- tants,auxincontentsarelowerthaninwildtype
try (Pekkeret al. 2005). Another link between and zmPIN1 expression in the incipient leaf
auxin and organ symmetry involves members primordia is also greatly reduced (Lee et al.
oftheKANADI(KAN)genefamily,whichreg- 2009). Unexpectedly, zmPIN1 is rapidly ind-
ulate abaxial identity and laminar growth of uced in the meristem on exogenous cytokinin
10 AdvancedOnlineArticle.CitethisarticleasColdSpringHarbPerspectBioldoi:10.1101/cshperspect.a001487