Division Plane Orientation Defects Revealed by a Synthetic Double Mutant Phenotype

Abstract

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...

An overview of plant division‐plane orientation

“…Surprisingly, tan1air9 double mutants have unexpected interphase microtubule array organization defects leading to defects in cell elongation and aberrant root cell-file rotation. Although full-length TAN1-YFP rescued the double mutant, a TAN1-YFP protein lacking a domain required for its localization to the PPB (Rasmussen et al, 2011b) rescued everything but the cell-file rotation defect, potentially highlighting this domain's function in interphase microtubule array organization (Mir et al, 2018). TAN1 and AIR9 likely act in parallel pathways to promote division-plane maintenance and organize cortical microtubule arrays but the mechanisms are still unknown.…”

“…Maintenance: POK1 interacts with TAN1 (M€ uller et al, 2006;Walker et al, 2007;Rasmussen et al, 2011b), RAN-GAP (Xu et al, 2008) andPHGAP1 and2 (St€ ockle et al, 2016). tan1 air9 double mutants have a synthetic division-plane orientation defect suggesting genetic interaction (Mir et al, 2018). AIR9 physically interacts with KCBP (Buschmann et al, 2015).…”

“…AIR9's contribution to division-plane orientation remained elusive because air9 mutants have no obvious division-plane defects (Buschmann et al, 2015) similar to very minor division-plane defects in Arabidopsis thaliana tan1 mutants (Walker et al, 2007). Recently, a function in division-plane orientation was revealed for AIR9 using a tan1air9 double mutant (Mir et al, 2018). The double mutant displays a synthetic phenotype: short plants with division-plane defects, hypersensitivity to microtubuledepolymerizing drugs, and root cell-file rotation (Mir et al, 2018).…”

“…Recently, a function in division-plane orientation was revealed for AIR9 using a tan1air9 double mutant (Mir et al, 2018). The double mutant displays a synthetic phenotype: short plants with division-plane defects, hypersensitivity to microtubuledepolymerizing drugs, and root cell-file rotation (Mir et al, 2018). Around half of the divisions completed in a location different than the PPB, indicating a significant defect in divisionplane maintenance.…”

Cellular Dynamics: Cellular Systems in the Time Domain

“…Long-term time-lapse imaging and cross-correlation analyses of leaf epidermal pavement cell shape and microtubule organization were used to reveal unexpected temporal and spatial variability of cortical microtubules during the process of lobe formation in pavement cells (Belteton et al, 2018). This issue also contains important new discoveries on the genetic control of microtubule arrays that position the cell division plane (Mir et al, 2018) and a plant-specific Kinesin (KinG) that uses microtubule-based transport to affect intercellular transport of a developmental regulatory protein (Spiegelman et al, 2018). The actin cytoskeleton also is required for the growth of cells that employ either tip or diffuse growth mechanisms.…”