Primary root protophloem differentiation requires balanced phosphatidylinositol-4,5-biphosphate levels and systemically affects root branching

Abstract

Protophloem is a specialized vascular tissue in growing plant organs, such as root meristems. In Arabidopsis mutants with impaired primary root protophloem differentiation, brevis radix (brx) and octopus (ops), meristematic activity and consequently overall root growth are strongly reduced. Second site mutation in the protophloem-specific presumed phosphoinositide 5-phosphatase COTYLEDON VASCULAR PATTERN 2 (CVP2), but not in its homolog CVP2-LIKE 1 (CVL1), partially rescues brx defects. Consistent with this finding, CVP2 hyperactivity in a wild-type background recreates a brx phenotype. Paradoxically, however, while cvp2 or cvl1 single mutants display no apparent root defects, the root phenotype of cvp2 cvl1 double mutants is similar to brx or ops, although, as expected, cvp2 cvl1 seedlings contain more phosphatidylinositol-4,5-biphosphate. Thus, tightly balanced phosphatidylinositol-4,5-biphosphate levels appear essential for proper protophloem differentiation. Genetically, OPS acts downstream of phosphatidylinositol-4,5-biphosphate levels, as cvp2 mutation cannot rescue ops defects, whereas increased OPS dose rescues cvp2 cvl1 defects. Finally, all three mutants display higher density and accelerated emergence of lateral roots, which correlates with increased auxin response in the root differentiation zone. This phenotype is also created by application of peptides that suppress protophloem differentiation, CLAVATA3/EMBRYO SURROUNDING REGION 26 (CLE26) and CLE45. Thus, local changes in the primary root protophloem systemically shape overall root system architecture.

Phosphoinositide signaling in plant development

“…In recent years, PIs have emerged as central regulators of plant function and development, and a number of important and severe plant phenotypes have been reported when plant PI metabolism is perturbed Nelson, 2009, 2004;Chen et al, 2008;Ercetin et al, 2008;Golani et al, 2013;Gunesekera et al, 2007;Ischebeck et al, 2008Ischebeck et al, , 2011Ischebeck et al, , 2013Kaye et al, 2011;Kusano et al, 2008;Löfke et al, 2008;Mei et al, 2012;Novakova et al, 2014;Parker et al, 2000;Preuss et al, 2006;Rodriguez-Villalon et al, 2015;Sousa et al, 2008;Stenzel et al, 2008;Tejos et al, 2014;Wang et al, 2012;Williams et al, 2005;Zhong et al, , 2005. The perturbation of PI signaling components in Arabidopsis, for example, gives rise to a variety of phenotypes affecting multiple tissue types and aspects of plant development (summarized in Table 1).…”

Phosphosite charge rather than shootward localization determines OCTOPUS activity in root protophloem

“…Therefore, they appear as distinct "gaps" in the protophloem differentiation zone (Fig. 1B), and the frequency of these so-called "gap cells" can serve as a quantitative indicator of phenotypic severity (18,(20)(21)(22).Mutants in which perception of the plant hormone brassinosteroid (BR) is knocked out also display protophloem differentiation defects; however, their phenotype is quantitatively less severe than Significance The evolution of plant vasculature was a key event in earth history because it enabled plants to effectively colonize land. The phloem vasculature was particularly important in this process.…”

“…1A) (12,15). In brx and ops loss-of-function mutants, sieve element differentiation is frequently impaired, which leads to discontinuous protophloem strands and various associated systemic effects, including strongly reduced root growth (12,15,18,19). Protophloem cells that fail to undergo sieve element differentiation in brx or ops can be visually identified by cell-wall staining in confocal microscopy because they not only fail to enucleate, but also fail to thicken their cell wall.…”

“…Therefore, they appear as distinct "gaps" in the protophloem differentiation zone (Fig. 1B), and the frequency of these so-called "gap cells" can serve as a quantitative indicator of phenotypic severity (18,(20)(21)(22).…”

The Effects of High Steady State Auxin Levels on Root Cell Elongation in Brachypodium

“…Likewise, Arabidopsis thaliana roots are shorter upon both a genetically imposed strong decrease as well as a strong increase in auxin production (Chen et al, 2014). Moreover, reduced mature cell length is typically accompanied by reduced meristem size and vice versa (Moubayidin et al, 2010;Scacchi et al, 2010;Rodriguez-Villalon et al, 2015), which makes it difficult to distinguish whether observed phenotypes are primarily caused by altered cell elongation or cell proliferation and/or differentiation. Thus, a paucity of clear-cut conditions and pertinent genetic material has prevented conclusive analyses of the Acid Growth Theory in Arabidopsis roots.…”