Differential Interaction between RAC/ROP-GTPases and RIC-Effectors: A Network Hub for Broader Signal Transduction in Pollen Tubes

Abstract

ABSTRACTTo date knowledge about plant RAC/ROP-GTPase effectors and downstream targets is still limited. This work aims on elucidation of related signal transduction networks involved in pollen tube growth. Yeast two-hybrid and Pull Down methodology were used to identify and characterize hitherto unknown components of RAC-related protein complexes fromNicotiana tabacum. Nt-RIC11pt specifically interacts with diverse active tobacco RAC-GTPases, and it is particularly significant, that their binding affinity is differential, thus implicating a multifaceted role in an interconnected RIC-RAC network. Moreover, Y2H-screening for Nt-RIC11pt targets identified Nt-CAR4, which is phylogenetically assigned to a multifaceted family of novel unusualGTPaseactivatingproteins (GAP). It is argued that scaffold Nt-RIC11pt connects active Nt-RAC3 with membrane-bound Nt-CAR4, thus relaying GAP-activity. Quantitative RT-PCR demonstrates Nt-RIC11pt is primarily expressed in pollen and YFP-fusion proteins show homogeneous cytoplasmic localization in growing tubes, what builds the prerequisite for a proposed role in broader signal transduction. By synoptically integrating experimental data, bioinformatic sequence comparison, phylogenetic analyses, and detailed literature review, this study hypothesizes a concept in which RIC-effectors collectively constitute a multifaceted network hub linking diverse GTPase-dependent processes.