Two subtypes of GTPase-activating proteins coordinate tip growth and cell size regulation in Physcomitrium patens.

The establishment of cell polarity is a prerequisite for many developmental processes. However, how it is achieved during tip growth in plants remains elusive. Here, we show that the RHO OF PLANTs (ROPs), ROP GUANINE NUCLEOTIDE EXCHANGE FACTORs (RopGEFs), and ROP GTPASE-ACTIVATING PROTEINs (RopGAPs) assemble into membrane domains in tip-growing cells of the moss Physcomitrium patens. The confinement of membrane domains requires redundant global inactivation of ROPs by PpRopGAPs and the PLECKSTRIN HOMOLOGY (PH) domain-containing RenGAP PpREN. Unexpectedly, PpRopGAPs and PpREN exert opposing effects on domain size and cell width upon overexpression. Biochemical and functional analyses indicate that PpRopGAPs are recruited to the membrane by active ROPs to restrict domain size through clustering, whereas PpREN rapidly inactivates ROPs and inhibits PpRopGAP-induced clustering. We propose that the activity- and clustering-based domain organization by RopGAPs and RenGAPs is a general mechanism for coordinating polarized cell growth and cell size regulation in plants.