title: Analysis of calcium spiking using a cameleon calcium sensor reveals that nodulation gene expression is regulated by calcium spike number and the developmental status of the cell.
abstract: Rhizobium-made Nod factors induce rapid changes in both Ca(2+) and gene expression. Mutations and inhibitors that abolish Nod-factor-induced Ca(2+) spiking block gene induction, indicating a specific role for Ca(2+) spiking in signal transduction. We used transgenic Medicago truncatula expressing a "cameleon" Ca(2+) sensor to assess the relationship between Nod-factor-induced Ca(2+) spiking and the activation of downstream gene expression. In contrast to ENOD11 induction, Ca(2+) spiking is activated in all root-hair cells and in epidermal or pre-emergent root hairs cells in the root tip region. Furthermore, cortical cells immediately below the epidermal layer also show slow Ca(2+) spiking and these cells lack Nod-factor-induced ENOD11 expression. This indicates a specialization in nodulation gene induction downstream of Nod-factor perception and signal transduction. There was a gradient in the frequency of Ca(2+) spiking along the root, with younger root-hair cells having a longer period between spikes than older root hairs. Using a Ca(2+)-pump inhibitor to block Ca(2+) spiking at various times after addition of Nod factor, we conclude that about 36 consecutive Ca(2+) spikes are sufficient to induce ENOD11-GUS expression in root hairs. To determine if the length of time of Ca(2+) spiking or the number of Ca(2+) spikes is more critical for Nod-factor-induced ENOD11 expression, jasmonic acid (JA) was added to reduce the rate of Nod-factor-induced Ca(2+) spiking. This revealed that even when the period between Ca(2+) spikes was extended, an equivalent number of Ca(2+) spikes were required for the induction of ENOD11. However, this JA treatment did not affect the spatial patterning of ENOD11-GUS expression suggesting that although a minimal number of Ca(2+) spikes are required for Nod-factor-induced gene expression, other factors restrict the expression of ENOD11 to a subset of responding cells.
authors.affiliation: Department of Molecular Microbiology, John Innes Centre, Colney Lane, Norwich NR4 7UH, UK.
authors.forename: Hiroki
authors.initials: H
authors.lastname: Miwa
authors.forename: Jongho
authors.initials: J
authors.lastname: Sun
authors.forename: Giles E D
authors.initials: GE
authors.lastname: Oldroyd
authors.forename: J Allan
authors.initials: JA
authors.lastname: Downie
mesh_terms: D001426:Bacterial Proteins / Q000378:metabolism; D020013:Calcium Signaling / Q000187:drug effects; D018506:Gene Expression Regulation, Plant; D018546:Hypocotyl / Q000166:cytology / Q000378:metabolism; D007211:Indoles / Q000494:pharmacology; D008164:Luminescent Proteins; D046913:Medicago truncatula / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D009586:Nitrogen Fixation / Q000235:genetics*; D010940:Plant Proteins / Q000235:genetics; D018517:Plant Roots / Q000166:cytology / Q000235:genetics / Q000378:metabolism*; D011994:Recombinant Proteins; D016962:Sinorhizobium meliloti / Q000235:genetics* / Q000378:metabolism; D013559:Symbiosis / Q000235:genetics; D015533:Transcriptional Activation
chemical_list: D001426:Bacterial Proteins; C438018:ENOD11 protein, Medicago truncatula; D007211:Indoles; D008164:Luminescent Proteins; D010940:Plant Proteins; D011994:Recombinant Proteins; C507867:yellow cameleon 2.1; C000543:cyclopiazonic acid