ORCID IDs: 0000-0003-1506-4683 (R.M.); 0000-0003-4881-6343 (V.H.M.); 0000-0002-4354-6295 (C.G.R.).
TANGLED1 (TAN1) and AUXIN-INDUCED-IN-ROOTS9(AIR9) are microtubule-binding proteins that localize to the division site in plants. Their function in Arabidopsis (Arabidopsis thaliana) remained unclear because neither tan1 nor air9 single mutants have a strong phenotype. We show that tan1 air9 double mutants have a synthetic phenotype consisting of short, twisted roots with disordered cortical microtubule arrays that are hypersensitive to a microtubule-depolymerizing drug. The tan1 air9 double mutants have significant defects in division plane orientation due to failures in placing the new cell wall at the correct division site. Full-length TAN1 fused to yellow fluorescent protein, TAN1-YFP, and several deletion constructs were transformed into the double mutant to assess which regions of TAN1 are required for its function in root growth, root twisting, and division plane orientation. TAN1-YFP expressed in tan1 air9 significantly rescued the double mutant phenotype in all three respects. Interestingly, TAN1 missing the first 126 amino acids, TAN1-DI-YFP, failed to rescue the double mutant phenotype, while TAN1 missing a conserved middle region, TAN1-DII-YFP, significantly rescued the mutant phenotype in terms of root growth and division plane orientation but not root twisting. We use the tan1 air9 double mutant to discover new functions for TAN1 and AIR9 during phragmoplast guidance and root morphogenesis.Plant cells are typically constrained by cell walls (Cosgrove, 2005) that are also connected via plasmodesmata (Brunkard and Zambryski, 2017), and they do not migrate relative to each other. In the absence of significant cell migration, the entire plant body must be built through elegant coordination between the division, expansion, and differentiation of cells. Therefore, division plane orientation, or the spatial control of cytokinesis, has important roles in plant development and growth (Pickett-Heaps et al., 1999).The key steps of plant division plane orientation occur during interphase, G2, and mitosis. During interphase, the cortical microtubule array typically aligns perpendicular to the cell expansion axis (Baskin, 2001), and during G2 (Gunning and Wick, 1985), this promotes the formation of a land plant-specific microtubule and microfilament array called the preprophase band (PPB;Pickett-Heaps and Northcote, 1966). Although the location of the PPB accurately predicts the future division site (Gunning et al., 1978;Van Damme et al., 2007;Rasmussen et al., 2013;Lipka et al., 2014;Martinez et al., 2017), its role in division plane establishment has recently been called into question by mutants that do not form obvious PPBs but manage to produce relatively orderly roots. These plants have mutations in the tonneau1a locus (Zhang et al., 2016) and mutations in three related tonneau1 recruiting motif (trm) loci (Schaefer et al., 2017). Whether the PPB establishes the future division site or is the signpost of an ear...