Long-distance signaling in bypass1 mutants: bioassay development reveals the bps signal to be a metabolite.

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.