Handouts

During plant embryogenesis, the formation of an axial-radial pattern
ultimately determines the developmental fates of the embryo (Capron et
al. 2009). In Arabidopsis, it starts with the asymmetrical zygotic division
that generates a small apical cell (purple) and a basal cell (grey). After two
rounds of longitudinal divisions and one round of transverse division, the
apical cell converts into an octant stage embryo (8-celled).

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Arabidopsis early embryo development

The naturel auxin molecule is an indole-derived compound, indole-3-acetic acid, IAA. It is produced either from, or independently of, Tryptophan (Trp). Illustrated here are the known pathways of Trp-dependent IAA production in plants.  

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Auxin Biosynthesis

The R2D2 auxin reporter system has been developed from the DII reporter (see the handout sheet “Auxin reporter: DII”) (Brunoud et al., 2012). The DII-Venus reporter is a chimeric protein composed of the fusion of the domain II of IAA28 to a nuclear targeted Venus fluorescent protein. The absence of the fluorescence, due to the targeted degradation of DII by auxin, marks the accumulation of auxin in the cell. 

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Auxin reporter:R2D2

The DII auxin reporter has been developed as a sensor to map the auxin response and distribution (Brunoud et al, 2012). This sensor is based on the characteristics of the auxin signaling pathway (A). The auxin signaling pathway relies on a ubiquitin- and proteasome-dependent degradation of the Aux/IAA repressor proteins, catalyzed by the SCF-type E3 ubiquitin-ligase complexes SCFTIR1/AFB1-5 (TIR1, ASK1, Cullin, RBX1, E2). TIR1, AFB1 to AFB5 are F-box proteins, receptors of the molecule auxin. Auxin (IAA) promotes the interaction between TIR1/AFBs and the Aux/IAA proteins.

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Auxin reporter:DII

A small sequence of 6 nucleotides (TGTCTC, Auxin response element AuxRE) has been identified on promoter of the GH3 genes from soybean, responsive to the hormone auxin (Ulmasov et al, 1997). This sequence was used to create a synthetic auxin response promoter referred to as DR5. The DR5 promoter consists in 7 repeats of a 11-nucleotide sequence, including the TGTCTC element and a minimal 35S promoter (Sabatini et al, 1999; Ulmasov et al, 1997). 

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Auxin reporters: DR5, DR5rev and DR5v2