A Live-imaging, Heat Shock-inducible System to Measure Aux/IAA Degradation Rates in Planta.

An emerging theme in biology is the importance of cellular signaling dynamics. In addition to monitoring changes in absolute abundance of signaling molecules, many signal transduction pathways are sensitive to changes in temporal properties of signaling components (Purvis and Lahav, 2013). The phytohormone auxin regulates myriad processes in plant development. Many of these require the nuclear auxin signaling pathway, in which degradation of the Aux/IAA repressor proteins allows for transcription of auxin-responsive genes (Korasick et al., 2015). Using a heterologous yeast system, we found that Aux/IAAs exhibit a range of auxin-induced degradation rates when co-expressed in isolation with F-box proteins (Havens et al., 2012). Subsequent studies connecting signaling dynamics to plant growth and development confirmed that Aux/IAAs show similar differences in plants (Guseman et al., 2015; Moss et al., 2015). Here, we describe in detail the use of a heat-shock-inducible fluorescence degradation system to capture Aux/IAA degradation in real time in live plant roots. By employing this method, we were able to obtain high Aux/IAA expression and avoid the dampening long term effects of turnover, feedback and silencing. Degradation was dependent on the presence of an Aux/IAA degron and rates increased in response to exogenous auxin.