A novel Ca2+-binding protein that can rapidly transduce auxin responses during root growth

Signaling cross talks between auxin, a regulator of plant development, and Ca2+, a universal second messenger, have been proposed to modulate developmental plasticity in plants. However, the underlying molecular mechanisms are largely unknown. Here, we report that in Arabidopsis roots, auxin elicits specific Ca2+ signaling patterns that spatially coincide with the expression pattern of auxin-regulated genes. We have identified the single EF-hand Ca2+-binding protein Ca2+-dependent modulator of ICR1 (CMI1) as an interactor of the Rho of plants (ROP) effector interactor of constitutively active ROP (ICR1). CMI1 expression is directly up- regulated by auxin, whereas the loss of function of CMI1 associates with the repression of auxin-induced Ca2+ increases in the lateral root cap and vasculature, indicating that CMI1 represses early auxin responses. In agreement, cmi1 mutants display an increased auxin response including shorter primary roots, longer root hairs, longer hypocotyls, and altered lateral root formation. Binding to ICR1 affects subcellular localization of CMI1 and its function. The interaction between CMI1 and ICR1 is Ca2+-dependent and involves a conserved hydrophobic pocket in CMI1 and calmodulin binding-like domain in ICR1. Remarkably, CMI1 is monomeric in solution and in vitro changes its secondary structure at cellular resting Ca2+ concentrations ranging between 10−9 and 10−8 M. Hence, CMI1 is a Ca2+-dependent transducer of auxin-regulated gene expression, which can function in a cell-specific fashion at steady-state as well as at elevated cellular Ca2+ levels to regulate auxin responses

A Rho Scaffold Integrates the Secretory System with Feedback Mechanisms in Regulation of Auxin Distribution

In plants, auxin distribution and tissue patterning are coordinated via a feedback loop involving the auxin-regulated cell polarity factor ICR1 and the secretory machinery

Activation Status-Coupled Transient S Acylation Determines Membrane Partitioning of a Plant Rho-Related GTPase

ROPs or RACs are plant Rho-related GTPases implicated in the regulation of a multitude of signaling pathways that function at the plasma membrane by virtue of posttranslational lipid modifications. The relationship between ROP activation status and membrane localization has not been established. Here we demonstrate that endogenous ROPs, as well as a transgenic His(6)-green fluorescent protein (GFP)-AtROP6 fusion protein, were partitioned between Triton X-100-soluble and -insoluble membranes. In contrast, an activated His(6)-GFP-Atrop6(CA) mutant protein accumulated exclusively in detergent-resistant membranes. GDP induced accumulation of ROPs in Triton-soluble membranes, whereas GTPγS induced accumulation of ROPs in detergent-resistant membranes. Recombinant wild-type and constitutively active AtROP6 isoforms were purified from Arabidopsis plants, and their lipids were cleaved and analyzed by gas chromatography-coupled mass spectrometry. In Triton-soluble membranes, wild-type AtROP6 was only prenylated, primarily by geranylgeranyl. The activated AtROP6 that accumulated in detergent-resistant membranes was modified by prenyl and acyl lipids. The acyl lipids were identified as palmitic and stearic acids. In agreement, activated His(6)-GFP-Atrop6(CA)mS(156) in which cysteine(156) was mutated into serine accumulated in Triton-soluble membranes. These findings show that upon GTP binding and activation, AtROP6 and possibly other ROPs are transiently S acylated, which induces their partitioning into detergent-resistant membranes