Project description:Floral organ identities are specified by few transcription factors which act as master regulators. Subsequently, specification of organ axes programs the distribution of distinct tissue types within the organs that themselves develop unique identities. The C-class, AGAMOUS-clade MADS box genes are primary promoters of the gynoecium which is divided into a distal style and a subtending ovary along the apical-basal axis. We show that members of a clade of B3-domain transcription factors, NGATHA1 to NGA4 (NGA1-4), are expressed distally in all lateral organs, and all four have a redundant and essential role in style development. Loss of all four genes results in gynoecia where style is replaced by valve-like projections and a reduction in style-specific SHATTERPROOF1 (SHP1) expression. In agreement, floral misexpression of NGA1 promotes ectopic style and SHP1 expression. STYLISH1, an auxin biosynthesis inducer, conditionally activated NGA genes, which in turn, promoted distal expression of other STY genes in a putative positive feed back loop. Inhibited auxin transport or lack of YABBY1 gene activities resulted in a basally expanded style domain and broader expression of NGA genes. We speculate that early gynoecium factors delimit NGA gene response to an auxin-based signal, elicited by STY gene activity, to restrict the activation of style program to a late and distal carpel domain. RNA was extracted from inflorescences of 30 day old plants using Qiagen RNAEasy kit. cRNA was synthesized and hybridized to Affymetrix ATH1 array according to manufacture recommendation. Seven repeats of wild type, two repeats of nga quadruple mutants and two repeats of 35S:amiR-NGA164a were collected. Signal values were obtained and normalized using the GC-RMA protocol, using R 2.7.2 (www.r-project.org) and Bioconductor 2.2 (www.bioconductor.org/). Average correlation values between repeats was 0.985, while the average correlation between wild type and mutant samples was only 0.972. As the average correlation between samples the nga quadruple mutants and 35S:amiR-NGA164a plants was 0.984, and given the identical phenotype, we grouped the two genotypes together.

Project description:Wild tobacco flowers wave rhythmically to facilitate specific pollinator interactions. This movement behavior is controlled by a regulatory network that involves the circadian clock- and auxin-signaling pathways. The plant hormone auxin, similarly to its function in tropic movements, acts as growth regulator in the circadian regulation of floral movement. Dorsoventral asymmetry in auxin levels and auxin transcriptional responses mediate the growth responses in the floral peduncle that make flowers move. Multiple components of the auxin-signaling pathway and auxin responses are under the control of circadian clock. However, it is unclear where these two pathways intersect and how collectively contribute to regulate specific rhythmic outputs. Here we found that the blue light photoreceptor and circadian clock component ZEITLUPE (ZTL) controls auxin responses through the regulation of the auxin-signaling pathway in a time-of-day and blue light specific manner. Abrogation of ZTL expression abolishes flower movement and the temporal gating of auxin-induced growth responses in the floral peduncle. ZTL regulates transcription and directly interacts with indole-3-acetic acid inducible 19 (IAA19), a circadian controlled gene that regulates development of curvature in moving organs. Indicating that ZTL modulates auxin sensitivity in part through the regulation of AUX/IAA transcriptional repressors. At night, growth responses in the peduncle to the synthetic auxin 2,4-D revealed that ZTL additionally controls auxin responses regulating auxin homeostasis. These results indicate that ZTL conveys temporal and environmental information, at multiple levels, into the auxin signaling-pathway and in this way sculpts the temporal gating of auxin responses that allow flowers to move. To gain further insight into the molecular basis of the movement of flowers we used a whole genome microarray as a discovery platform to identify genes differentially expressed in adaxial and abaxial sides of the floral peduncle at different stages of the movement.

Project description:Wild tobacco flowers wave rhythmically to facilitate specific pollinator interactions. This movement behavior is controlled by a regulatory network that involves the circadian clock- and auxin-signaling pathways. The plant hormone auxin, similarly to its function in tropic movements, acts as growth regulator in the circadian regulation of floral movement. Dorsoventral asymmetry in auxin levels and auxin transcriptional responses mediate the growth responses in the floral peduncle that make flowers move. Multiple components of the auxin-signaling pathway and auxin responses are under the control of circadian clock. However, it is unclear where these two pathways intersect and how collectively contribute to regulate specific rhythmic outputs. Here we found that the blue light photoreceptor and circadian clock component ZEITLUPE (ZTL) controls auxin responses through the regulation of the auxin-signaling pathway in a time-of-day and blue light specific manner. Abrogation of ZTL expression abolishes flower movement and the temporal gating of auxin-induced growth responses in the floral peduncle. ZTL regulates transcription and directly interacts with indole-3-acetic acid inducible 19 (IAA19), a circadian controlled gene that regulates development of curvature in moving organs. Indicating that ZTL modulates auxin sensitivity in part through the regulation of AUX/IAA transcriptional repressors. At night, growth responses in the peduncle to the synthetic auxin 2,4-D revealed that ZTL additionally controls auxin responses regulating auxin homeostasis. These results indicate that ZTL conveys temporal and environmental information, at multiple levels, into the auxin signaling-pathway and in this way sculpts the temporal gating of auxin responses that allow flowers to move. To gain further insight into the molecular basis of temporal regulation of the movement of flowers we used a whole genome microarray as a discovery platform to identify genes differentially expressed in a RNAi knockdown line silenced in the expression of the circadian clock component ZEITLUPE (irZTL-314).

Project description:The emerging picture of transcriptional regulation is one of unexpected complexity. It is now clear that single transcription factors control hundreds, if not thousands, of direct targets by binding their genomic loci, but it is not understood how many of these are major players and how many are supporting cast. To address this, we leverage a well-characterized developmental network in Arabidopsis and map genome-wide binding of related proteins in multiple tissues. The transcription factor APETALA2 (AP2) has numerous functions, including roles in floral organ identity, seed development and stem cell maintenance. We focus on the role of AP2 in the floral transition and map direct targets on a genome-wide scale. We show that ap2 mutants flower early in long and short days, and that AP2 binds to many loci, most prominently floral pathway integrators, microRNAs and floral organ identity genes, many of which exhibit AP2-dependent transcription. Opposing, logical effects are evident in AP2 binding to two developmental microRNA genes that control AP2 expression, with AP2 positively regulating miR156 and negatively regulating miR172, forming a complex direct feedback loop, which also included all but one of the AP2-like miR172 target clade members. We also seek conserved targets by comparing the genome-wide direct target repertoire of AP2 with that of SCHLAFMÜTZE (SMZ), another member of the AP2-like miR172 target clade that shares partial redundancy, as evidenced by a hexuple mutant for the entire clade that flowered extremely early. Clear similarities and divergence are exposed in the AP2 and SMZ direct target repertoires. Finally, using an inducible expression system, we demonstrate that AP2 has dual molecular roles. It functions both as a transcriptional activator and repressor, directly inducing the expression of the floral repressor AGAMOUS-LIKE 15 (AGL15), and directly repressing the transcription of floral activators like SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1). ChIP-Seq of two biological replicates for ATH-AP2 and respective control samples

Project description:The emerging picture of transcriptional regulation is one of unexpected complexity. It is now clear that single transcription factors control hundreds, if not thousands, of direct targets by binding their genomic loci, but it is not understood how many of these are major players and how many are supporting cast. To address this, we leverage a well-characterized developmental network in Arabidopsis and map genome-wide binding of related proteins in multiple tissues. The transcription factor APETALA2 (AP2) has numerous functions, including roles in floral organ identity, seed development and stem cell maintenance. We focus on the role of AP2 in the floral transition and map direct targets on a genome-wide scale. We show that ap2 mutants flower early in long and short days, and that AP2 binds to many loci, most prominently floral pathway integrators, microRNAs and floral organ identity genes, many of which exhibit AP2-dependent transcription. Opposing, logical effects are evident in AP2 binding to two developmental microRNA genes that control AP2 expression, with AP2 positively regulating miR156 and negatively regulating miR172, forming a complex direct feedback loop, which also included all but one of the AP2-like miR172 target clade members. We also seek conserved targets by comparing the genome-wide direct target repertoire of AP2 with that of SCHLAFMÜTZE (SMZ), another member of the AP2-like miR172 target clade that shares partial redundancy, as evidenced by a hexuple mutant for the entire clade that flowered extremely early. Clear similarities and divergence are exposed in the AP2 and SMZ direct target repertoires. Finally, using an inducible expression system, we demonstrate that AP2 has dual molecular roles. It functions both as a transcriptional activator and repressor, directly inducing the expression of the floral repressor AGAMOUS-LIKE 15 (AGL15), and directly repressing the transcription of floral activators like SUPPRESSOR OF OVEREXPRESSION OF CONSTANS 1 (SOC1).

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Ngatha mutant inflorescences

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