Long-Distance Signaling in bypass1 Mutants: Bioassay Development Reveals the bps Signal to Be a Metabolite

Abstract

Root-to-shoot signaling is used by plants to coordinate shoot development with the conditions experienced by the roots. A mobile and biologically active compound, the bps signal, is over-produced in roots of an Arabidopsis thaliana mutant called bypass1 (bps1), and might also be a normally produced signaling molecule in wild-type plants. Our goal is to identify the bps signal chemically, which will then allow us to assess its production in normal plants. To identify any signaling molecule, a bioassay is required, and here we describe the development of a robust, simple, and quantitative bioassay for the bps signal. The developed bioassay follows the growth-reducing activity of the bps signal using the pCYCB1;1::GUS cell cycle marker. Wild-type plants carrying this marker, and provided the bps signal through either grafts or metabolite extracts, showed reduced cell division. By contrast, control grafts and treatment with control extracts showed no change in pCYCB1;1::GUS expression. To determine the chemical nature of the bps signal, extracts were treated with RNase A, Proteinase K, or heat. None of these treatments diminished the activity of bps1 extracts, suggesting that the active molecule might be a metabolite. This bioassay will be useful for future biochemical fractionation and analysis directed toward bps signal identification.

Reporter Gene-Facilitated Detection of Compounds in Arabidopsis Leaf Extracts that Activate the Karrikin Signaling Pathway

“…The resulting reporter system allows the source-response relationship to be assayed readily. The approach adopted here mirrors that of Adhikari et al (2013), who used a reporter system to establish both the presence and chemical nature of the ‘bypass’ signal that controls shoot growth, although its identity is yet to be determined.…”

The Mobile bypass Signal Arrests Shoot Growth by Disrupting Shoot Apical Meristem Maintenance, Cytokinin Signaling, and WUS Transcription Factor Expression

“…Here, we extended the grafting approach to test whether the bps1 root was also sufficient to repress pWUS::GUS expression. We used a transient micrograft assay (Adhikari et al, 2013), which, by providing an aqueous cushion between the scion and rootstock (agarose in water, ;0.3 mm; Supplemental Fig. S2A), allows the transfer of hydrophilic molecules between scion and rootstock without the delay from tissue repair that occurs with traditional grafting.…”

“…Transient micrografts, a modification of the Arabidopsis hypocotyl graft, were described previously (Turnbull et al, 2002;Adhikari et al, 2013). Briefly, the rootstock-scion junction was stabilized inside a silicone sleeve (0.012-in.…”

“…We are pursuing characterization of this pathway to determine whether wild-type plants use the bps signal as part of a normal physiological response. For example, the bps signal might be used for growth arrest of salt-stressed plants, as both salt stress and the bps signal halt cell cycle progression (Burssens et al, 2000;Van Norman et al, 2011;Adhikari et al, 2013).…”

Editorial overview: Physiology and metabolism: Plant metabolism: globules to global, modules to models

“…The MEP pathway intermediate MEcPP, carotenoid derivative b-cyclotitral and products of tetrapyrrol metabolism are all implicated in this complex control system. There are still other terpenoid signalling molecules awaiting discovery [9][10][11].…”