Transport of signaling molecules is of major importance for regulating plant growth, development, and responses to the environment. A prime example is the spatial-distribution of auxin, which is regulated via transporters to govern developmental patterning. A critical limitation in our ability to identify transporters by forward genetic screens is their potential functional redundancy. Here, we overcome part of this functional redundancy via a transportome, multi-targeted forward-genetic screen using artificial-microRNAs (amiRNAs). We generate a library of 3000 plant lines expressing 1777 amiRNAs, designed to target closely homologous genes within subclades of transporter families and identify, genotype and quantitatively phenotype, 80 lines showing reproducible shoot growth phenotypes. Within this population, we discover and characterize a strong redundant role for the unstudied ABCB6 and ABCB20 genes in auxin transport and response. The unique multi-targeted lines generated in this study could serve as a genetic resource that is expected to reveal additional transporters.

Plants have evolved a unique and conserved developmental program that enables the conversion of leaves into floral organs. Elegant genetic and molecular work has identified key regulators of flower meristem identity. However, further understanding of flower meristem specification has been hampered by redundancy and by pleiotropic effects. The KNOXI transcription factor SHOOT MERISTEMLESS (STM) is a well-characterized regulator of shoot apical meristem maintenance. loss-of-function mutants arrest shortly after germination; therefore, the knowledge on later roles of STM in later processes, including flower development, is limited. Here, we uncover a role for STM in the specification of flower meristem identity. Silencing in the () expression domain in the mutant background resulted in a leafy-flower phenotype, and an intermediate allele enhanced the flower meristem identity phenotype of Transcriptional profiling of perturbation suggested that STM activity affects multiple floral fate genes, among them the F-box protein-encoding gene (). In agreement with this notion, enhanced the floral fate phenotype, and ectopic expression rescued the leafy flowers in genetic backgrounds with compromised and activities. This work suggests a genetic mechanism that underlies the activity of in the specification of flower meristem identity.

Although strongly influenced by environmental conditions, lateral root (LR) positioning along the primary root appears to follow obediently an internal spacing mechanism dictated by auxin oscillations that prepattern the primary root, referred to as the root clock. Surprisingly, none of the hitherto characterized PIN-and ABCB-type auxin transporters seem to be involved in this LR prepatterning mechanism. Here, we characterize ABCB15, 16, 17,[18][19][20][21][22] as novel auxin transporting ABCBs. Knock-down and genome editing of this genetically linked group of ABCBs caused strongly reduced LR densities. These phenotypes were correlated with reduced amplitude, but not reduced frequency of the root clock oscillation. High-resolution auxin transport assays, and tissue-specific silencing revealed contributions of ABCB15-22 to shootward auxin transport in the lateral root cap (LRC) and epidermis, thereby explaining the reduced auxin oscillation. Jointly, these data support a model in which LRC-derived auxin contributes to the root clock amplitude.

Root branching is an important strategy to explore efficiently large volumes of soil. To economize this process, lateral roots (LR) are formed along the growing root at discrete positions that are instructed by oscillating auxin signals derived from the lateral root cap (LRC). This assumes that auxin moves from the LRC across multiple layers to accumulate in the pericycle. Here, we identified, using gene silencing and CRISPR based approaches, a group of five genetically linked, closely related ABCBs that control LR spacing by modulating the amplitude of the auxin oscillation. The transporters localize to the plasma membrane and reveal significant auxin export activity. These ABCBs are mainly expressed in the LRC and epidermis where they contribute to auxin transport towards the root oscillation zone. Our findings highlight the importance of auxin transport in the outer tissues of the root meristem to regulate LR spacing.

Clonal propagation of plants by induction of adventitious roots (ARs) from stem cuttings is a requisite step in breeding programs. Nevertheless, a major barrier exists for propagating valuable plants that naturally have low capacity to form ARs. Due to the central role of auxin in organogenesis, indole-3-butyric acid (IBA) is often utilized, yet many recalcitrant plants do not form ARs in response to such treatment. We describe the synthesis and screening of a focused library of synthetic auxin conjugates in Eucalyptus grandis cuttings, highlighting 4-chlorophenoxyacetic acid-L-tryptophan-OMe as a competent enhancer of adventitious rooting in a number of recalcitrant woody plants. Comprehensive metabolic and functional analyses revealed that this activity is engendered by prolonged auxin signaling due to initial fast uptake and slow release and clearance of the free auxin 4 chlorophenoxyacetic acid. This work highlights the utility of a slow-release strategy for bioactive compounds and provides an exemplar for further rational development of more effective plant-growth regulators for agriculture.