Aux/IAA Proteins Are Phosphorylated by Phytochrome in Vitro

Abstract

Auxin/indole-3-acetic acid (Aux/IAA) genes encode short-lived transcription factors that are induced as a primary response to the plant growth hormone IAA or auxin. Gain-of-function mutations in Arabidopsis genes, SHY2/IAA3, AXR3/IAA17, and AXR2/IAA7 cause pleiotropic phenotypes consistent with enhanced auxin responses, possibly by increasing Aux/IAA protein stability. Semidominant mutations shy2-1D, shy2-2, axr3-1, and axr2-1 induce ectopic light responses in dark-grown seedlings. Because genetic studies suggest that the shy2-1D and shy2-2 mutations bypass phytochrome requirement for certain aspects of photomorphogenesis, we tested whether SHY2/IAA3 and related Aux/IAA proteins interact directly with phytochrome and whether they are substrates for its protein kinase activity. Here we show that recombinant Aux/IAA proteins from Arabidopsis and pea (Pisum sativum) interact in vitro with recombinant phytochrome A from oat (Avena sativa). We further show that recombinant SHY2/IAA3, AXR3/IAA17, IAA1, IAA9, and Ps-IAA4 are phosphorylated by recombinant oat phytochrome A in vitro. Deletion analysis of Ps-IAA4 indicates that phytochrome A phosphorylation occurs on the N-terminal half of the protein. Metabolic labeling and immunoprecipitation studies with affinity-purified antibodies to IAA3 demonstrate increased in vivo steady-state levels of mutant IAA3 in shy2-2 plants and phosphorylation of the SHY2-2 protein in vivo. Phytochrome-dependent phosphorylation of Aux/IAA proteins is proposed to provide one molecular mechanism for integrating auxin and light signaling in plant development.

Phytochrome RNAi enhances major fibre quality and agronomic traits of the cotton Gossypium hirsutum L

“…Interactions of auxin with PHYA through phytochromedependent phosphorylation of auxin-related proteins, increases in auxin biosynthesis or regulation of auxin distribution 26 could explain the fibre length improvement in our PHYA RNAi plants 6 . Further, we previously observed that photomorphogenesis-related factors (for example, PHYC, SPA1, FAR1, COP1, COP9, CIP7, CIP8 and RTP2) were targeted by putative ovule-derived short interfering RNAs during the initiation and elongation phases of fibre development 28 , thus indirectly supporting a role for light signalling in fibre traits.…”

“…As cotton lacks the sophisticated molecular and genomic 'tool kit' of the model plant Arabidopsis, including facile Agrobacterium transformation 24 and comprehensive collections of knockout mutants 25 , it is not possible at this point to ascribe the fibre phenotypes directly to the suppression of PHYA1 or, alternatively, to the indirect effects of the increased levels of PHYA2, PHYC and PHYE. In either case, enhancement of fibre length could occur because of a change in phytochrome-mediated plant hormone signalling 1,26,27 . Auxin (indole-3-acetic acid, IAA), abscisic acid, gibberellic acid, brassinosteroids, ethylene and cytokinin are known factors associated with fibre development 6 .…”

Auxin signaling and regulated protein degradation

“…In another study, mutations in the PINOID Ser/Thr kinase result in defects in auxin response [49]. Phytochrome A is known to phosphorylate the N-terminal half of some AUX/IAA proteins in vitro [50], an event that might regulate auxin-induced AUX/IAA degradation. However, results from several laboratories [11,41] indicate that phosphorylation is not directly involved in regulating AUX/IAA degradation.…”

Auxin-Binding Protein 1 is a negative regulator of the SCFTIR1/AFB pathway

“…An attractive hypothesis is that the ABP1 pathway would confer auxin conditionality to the interaction between TIR1/AFBs and AUX/IAAs. Such conditionality might result from a change in subcellular or nuclei protein localization of AUX/IAA and/or TIR1/AFB proteins 22,33 , from the presence or absence of another protein or peptide interacting or competing with one of the component of the transcriptional regulatory module [34][35][36][37] or from post-translational modification affecting the relative affinity between TIR1/AFBs 38 and AUX/ IAAs 28,39 . A future challenge will be to elucidate the molecular basis of the action of ABP1 on the SCF TIR1/AFB pathway and to identify ABP1 downstream signalling elements filling the gap between ABP1 and the nuclear-localized SCF TIR1/AFB pathway.…”