Goto, Koji

Homma, Takashi

Meyerowitz, Elliot M.

Oka, Atsuhiro

Kyoto University Research Information Repository

TitleRegulation ofArabidopsis thaliana Flower Development(MOLECULAR BIOLOGY AND INFORMATION -Molecular Biology)Author(s)Goto, Koji; Homma, Takashi; Meyerowitz, Elliot M.; Oka,AtsuhiroCitationICR annual report (1996), 2: 50-51Issue Date1996-03URL http://hdl.handle.net/2433/65698RightType ArticleTextversionpublisherKyoto University50 feR Annual Report, Vol. 2, 1995Regulation of Arabidopsis thalianaFlower DevelopmentKoji Goto, Takashi Homma, Elliot M. Meyerowitz* and Atsuhiro OkaAn Arabidopsis thaliana flower is composed of four organs, sepals, petals, stamens, and carpels.Development of these organs is determined by three classes of homeotic genes. One member genePISTILLATA (pn contributes to the formation of both petals and stamens. We have now cloned the PIgene with its promoter region and clarified the time and locations of PI expression during floralorganogenesis of the wild type plant and the flower mutant plants (pistillata, apetala3, and superman).Keywords: ABC model/Floral development/Homeotic gene/MADS box/Transcription factorOccurrence of cell divisions in higher plants suchas Arabidopsis thaliana is generally restricted to themeristems after embriogenesis has been finished.The apical meristem is organized tissues of pluripo-tent "stem" cells. Reproductive development inArabidopsis is controlled by the activities of the inflo-rescence and the floral meristems derived from theapical meristem. The inflorescence meristem dis-plays a pattern of indeterminate growth, whereas thefloral meristem shows a determinate pattern of celldivision and organogenesis, always resulting in theproduction of four whorls of floral organs (i.e., sepals,petals, stamens, and carpels). Genetic studies haveshown that the establishment of the floral meristemrequires several genes including LEAFY (LFY),APETALA1 (AP1), and APETALA2 (AP2) , and that*Division of Biology, California Institute of Technology,Pasadena, California 91125 USAthe floral organogenesis is directed by three classes oforgan identity genes (A, B, and C). The class A genesof Arabidopsis that are presently known are AP1 andAP2, the class B genes are PISTILLATA (PD andAPETALA3 (AP3) , and the only known class C geneis AGAMOUS (AG). Representative mutant flowersdefective in each class of the genes are shown inFigure (right half). How these homeotic genes specifythe four whorls is interpreted by the ABC model inwhich the organs of each whorl are determined bycombinations of the three class genes: sepals andcarpels are directed by the class A and the class Cgenes, respectively; petals and stamens are given bya combination between the class A and B genes andbetween the class Band C genes, respectively; andthe class A and C genes are mutually exclusive (seeFigure) [1].To date, the molecular cloning of PI, AG, AP1,AP2, and AP3 from Arabidopsis has been reported.MOLECULAR BIOLOGY AND INFORMATION -Molecular Biology-Scope of researchAttempts have been made to elucidate structure-function relationships of genetic materials and various geneproducts. The major subjects are mechanisms involved in signal transduction and regulation ofgene expres-sion responsive to environmental stimuli, development ofplant leaves and flowers, and plant-microbe interac-tion. As ofDecember 1995, study is being concentrated on (1) roles of homeo domain proteins and MADS boxproteins in developmental processes and transcriptional control in higher plants and (2) contribution ofpro-tein phosphorylation and dephosphorylation toward cell cycle control and signal transduction in plants andplant pathogens.OKA AOYAMA GOTO lMAJUKUProfessorOKA, Atsuhiro (D Sc)Associate ProfessorAOYAMA, Takashi (D Sc)InstructorGOTO, Koji (D Sc)Associate InstructorlMAJUKU, YoshiroStudentsAOKI, Mikio (DC); ITAHANA, Koji(DC); ISHIDA, Norihiro (DC); TSUKU-DA, Mayumi (DC); OHGISHI, Maki(MC); SAKAI, Hiroe (MC); HOMMA,Takashi (MC); UEDA, Yumi (RS)feR Annual Report, Vo!' 2, 1995 51Figure, Arrangement of floral organs in the wild-typeArabidopsis is shown in the left (1). Flowers in the rightindicate those of a (2), b (3), and c (4) homeotic mutants,respectively. Four rings correspond to the whorls (Se, se-pals; Pe, petals; St, stamens; and Ca, carpels), and A, B, andC within them indicate an expressing gene(s).All of these gene products except AP2, and also theirsnapdragon homologs, GLOBOSA (GLO), PLENA(PLE) , SQUAMOSA (SQUA), and DEFICIENS(DEF) , respectively, have been characterized by ahighly conserved 58-amino-acid DNA binding domaincalled MADS box. This name is come from yeastMCMl (minichromosome 1 maintenance),Arabidopsis AG, snapdragon IlEF, and humanSRF (serum response factor). MCMl and SRF areknown to function as homodimers to regulate geneexpression in response to extracellular signals. SincePI has been expected to be a member of the MADSbox genes on the analogy of other organ identitygenes, we have cloned it by using PCR-amplified GLOcDNA as a probe. A flower cDNA library wasscreened under the low stringency conditions. Eightpositive clones obtained were all derived from thesame RNA species [2]. Also a genomic clone corre-sponding to these cDNAs was isolated. The clonedDNA actually corresponds to the PI gene was con-firmed by RFLP mapping and by sequencing of threePI mutant alleles (pi-i, pi-2, andpi-3). The PI cDNAhas a single protein coding frame of 208-amino-acidresidues. This protein includes a MADS box domainand a K box domain at the amino-terminal and thecentral portions, respectively. The latter domainseems to be liable to coiled-coil formation because ofthe resemblance to the amphipathic a-helical regionof keratin proteins [2].To see the precise spatiotemporal expression pat-tern of PI, in situ hybridization was done with crosssections prepared from various developmental stagesof flowers. Hybridization signals of PI mRNA appearat stage 3 flowers, the same stage when AG and AP3mRNA's are first detected. The signals are all locatedcentripetally in a region of floral meristem, cells atwhich will later contribute the formation of not onlythe petal and stamen but also ovary. However, sig-nals gradually become limited to the second and thirdwhorl cells, and remain at high levels until stages 10and 11 where differentiation of the cells in each organtype is finished. This PI expression pattern is differ-ent from that of another class B gene AP3: signals ofAP3 mRNA are always restricted in the cells formingthe second and third whorls and never detected in thecenter (fourth whorl) of developing flowers [2].PI expression was analyzed in the ap3-3 and pi-imutant backgrounds. PI expression in ap3-3 flowersis normal in early stages but not in later stages, sug-gesting a role for AP3 in maintenance, but not estab-lishment, of PI expression. PI mRNA in pi-i flowersseems to accumulate normally at stages 3-6, beforethe primordia of the second and third whorl organsbegin to differentiate obviously, but then to decrease.The weaker mutant allele pi-2 shows a less severeeffect on the reduction of PI mRNA level. Thus, PI islikely to be necessary for maintaining its own expres-sion [2]. This was confirmed by transient expressionexperiments in which the GUS reporter gene placeddownstream of the PI promoter was introduced witha particle delivery procedure into the leaves of thewild-type plant and of a transgenic plant constitu-tively expressing PI [3]. The superman mutant dis-plays the phenotype of reduction or elimination of theovary, concomitant with appearance of many extrastamens in the forth whorl [1]. This phenotype isattributed to PI (and presumably AP3, too) expres-sion at the forth whorl. Effects of overexpression ofPI and AP3 were tested with transgenic plants carry-ing the cauliflower mosaic virus 35S promoter-drivenPI and AP3 [3]. Plants overexpressing either geneare expected to show the normal phenotype, whilethose overexpressing both genes simultaneously arethought to convert homeotically sepal-to-petal andcarpel-to-stamen, provided no other gene is requiredfor the class B function. The latter gives the expectedresults, indicating PI and AP3 being sufficient for theclass B function. However, the former phenotypedoes not become normal: sepal-to-petal conversionoccurs in the 35S::PI plants, whereas carpel-to-sta-men conversion does in the 35S::AP3 plants. Theseresults suggest that PI induces AP3 expression at thefirst whorl but not the forth whorl and that AP3induces PI expression at the forth whorl but not thefirst whorl. Although there is no direct biochemicalevidence of Arabidopsis homeotic gene products beingtranscription factors at present, these experimentalresults suggest the existence of complicated regulato-ry network in control of flower morphogenesis.Assignment of DNA elements responsive to each fac-tor on the PI promoter region is now in progress.References1. Weigel D and Meyerowitz EM, Cell, 78, 203-209(1994).2. Goto K and Meyerowitz EM, Genes & Develop., 8,1548-1560 (1994).3. Goto K, Homma T and Oka A, unpublished results.Sf>SeC4lCa.(2) ,\ Inntant(:.q b ulutantAACCC C'aB'e StB+C StC ('aPt'StCaAA'BB-CC'Y.~..~.~.

Regulation ofArabidopsis thaliana Flower Development (MOLECULAR BIOLOGY AND INFORMATION - Molecular Biology)

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Regulation ofArabidopsis thaliana Flower Development (MOLECULAR BIOLOGY AND INFORMATION - Molecular Biology)

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