PMC

Flowchart representation of the strategy and main findings. The experimental parts of the work are shown in light green boxes. The connections between the different parts of the work are represented by directed arrows.

Structure the auxin signalling network. (A) Visual representation of the Aux/IAA-ARF interactome using Cytoscape (http://www.cytoscape.org). The proteins are grouped according to their biological identity as indicated. Note the global differences in connectivity of the three biological groups (B–D) Connectivity graph and clusters identified by the MixNet algorithm. The probabilities associated with the connectivity structure for the global network are indicated in (B). The three clusters are mainly composed of Aux/IAA (I), ARF activators (II) and ARF repressors (III) as indicated in brackets in (B). The identity of the proteins in these clusters for both the global network (C) and the SAM-specific network (D) is shown. The proteins are ordered from the most to the least central in each cluster based on the distance of the protein to the cluster. (E) The topology of the network relies on stereotypic interaction capacities for the different classes of proteins as represented. ARF+: ARF activators; ARF−: ARF repressors.

Spatial control and dynamics of auxin signalling at the inflorescence meristem. (A) Schematic representation of signalling parameters monitored by DII-VENUS as compared with DR5::VENUS. (B–D) Expression of DII-VENUS (B), mDII-VENUS (C) and DR5::VENUS (D) visualized by confocal microscopy. Insets: overlay of the VENUS signal (green) with the autofluorescence signal (red). In (B, D) the three first primordia (P) are indicated and numbered from the youngest to the oldest. Two initia (I) are indicated and numbered from the oldest to the youngest following standard nomenclature. (E) Auxin-dependent binding of IAA28 domain II to TIR1/AFB auxin co-receptors. Anti-FLAG immunoblots of IAA28 domain II peptide pull-down assay with TIR1-FLAG, AFB1-FLAG or AFB5-FLAG. IAA treatments are as indicated. (F, G) Time course of DII-VENUS (F) and DR5::VENUS (G) expression followed by confocal microscopy (VENUS fluorescence in green). Images were taken at indicated time after t0. The initia (I1–3) and the localization of the centre of the meristem (C) are indicated. Scale is the same in all images. Scale bar: 50 μm.

Spatial regulation of Aux/IAA-ARF signalling in the inflorescence. (A–D). Expression patterns of TIR1/AFB F-box co-receptors. Expression was analysed using GUS translational fusions for TIR1, AFB1 and AFB3 and in situ hybridization for AFB5. The relative levels of the protein are indicated for AFB1 and TIR1, as revealed by GUS activity detection time (+for TIR1: 48h; +++for AFB1: 8 h). (E–ZC) Expression patterns of ARFs and Aux/IAAs revealed by RNA in situ hybridization. (ZD) Detection of Aux/IAA and ARF expression by RT–qPCR in the inflorescence meristem. The analysis was done in duplicate on meristem mRNAs. Error bars represent the range of values. (ZE) Schematic representation of the Aux/IAAs and ARFs distribution in the meristem. The meristem is represented as a dome (PZ, peripheral zone; CZ, central zone, in grey; SC, stem cells; OC, organizing centre). Global tendency in expression levels are indicated by the size of the+sign. A dashed line was drawn between the upper and lower part of the centre to indicate differences in signalling capacities since ARF4 and IAA18 are expressed in the inner core. The primordia (P) have been delineated by a dashed line in the PZ to indicate that several Aux/IAAs and ARFs show an even higher accumulation in the organ primordia. Several Aux/IAAs and ARFs are also more specifically associated with vasculature (V; see main text). Median or near-median sections are shown. Scale bar: 50 μm.

Mathematical model of the auxin signalling network. (A) Reaction scheme considered for the model. The numbers in brackets indicate the five populations of molecule described by ODEs in the model. (B) Effect of the level of ARF activators (middle panel) and ARF repressors (bottom panel) on target gene induction capacity upon an increase in auxin (upper panel). (C) 3D representation of the induction capacity as a function of ARF levels and ARF activator to ARF repressor ratio. The surface has been obtained by calculating the transcription fold change, that is, the ratio of the mRNA levels at equilibrium before and after a step increase in auxin. A colour map representing the parameter values is shown. (D) Effect of the level of ARF activators (middle panel) and ARF repressors (lower panel) on the stability of target gene induction upon varying auxin level. Transcription in response to sinusoidal changes in auxin levels (upper panel) has been studied. Two situations, corresponding to the centre (CZ) or the periphery (PZ), were considered. The effect of increasing ARF activators was tested for the first situation (CZ + ARF⁺) and of decreasing ARF repressors for the second (PZ − ARF⁻). For simplicity, mRNA levels are shown in (B, D) for only the higher and the lower concentration of the variable parameter used in the simulation. See and of for the full range of values.