A molecular rheostat adjusts auxin flux to promote root protophloem differentiation

Abstract

Auxin influences plant development through several distinct concentration-dependent effects . In the Arabidopsis root tip, polar auxin transport by PIN-FORMED (PIN) proteins creates a local auxin accumulation that is required for the maintenance of the stem-cell niche. Proximally, stem-cell daughter cells divide repeatedly before they eventually differentiate. This developmental gradient is accompanied by a gradual decrease in auxin levels as cells divide, and subsequently by a gradual increase as the cells differentiate. However, the timing of differentiation is not uniform across cell files. For instance, developing protophloem sieve elements (PPSEs) differentiate as neighbouring cells still divide. Here we show that PPSE differentiation involves local steepening of the post-meristematic auxin gradient. BREVIS RADIX (BRX) and PROTEIN KINASE ASSOCIATED WITH BRX (PAX) are interacting plasma-membrane-associated, polarly localized proteins that co-localize with PIN proteins at the rootward end of developing PPSEs. Both brx and pax mutants display impaired PPSE differentiation. Similar to other AGC-family kinases, PAX activates PIN-mediated auxin efflux, whereas BRX strongly dampens this stimulation. Efficient BRX plasma-membrane localization depends on PAX, but auxin negatively regulates BRX plasma-membrane association and promotes PAX activity. Thus, our data support a model in which BRX and PAX are elements of a molecular rheostat that modulates auxin flux through developing PPSEs, thereby timing PPSE differentiation.

A Plant-Specific Polarity Module Establishes Cell Fate Asymmetry in the Arabidopsis Stomatal Lineage

“…Here we identify members of the BREVIS RADIX (BRX) protein family as such components. The family's founding member, BRX, previously linked to cell proliferation and elongation in the root, has been hypothesized to integrate auxin and brassinosteroid (BR) hormone signaling and was recently incorporated into a molecular rheostat to modulate auxin flux during protophloem development in the root (Marhava et al, 2018). We demonstrate that in the context of the asymmetric cell divisions of the stomatal lineage, however, BRX and its paralogs are partners of BASL.…”

“…1C and Adrian et al, 2015). To confirm the transcriptomic data, we generated transcriptional reporters for each of the five Arabidopsis BRX family genes (hereafter, designated BRXf) ( Fig BRXf genes are redundantly required for asymmetric division in the stomatal lineage BRX has a well-established role in root growth and protophloem differentiation (Rodriguez-Villalon et al, 2014;Marhava et al, 2018). Although a leaf growth phenotype has also been described in brx mutants (Beuchat et al, 2010b), this was not investigated at the level of stomatal production or pattern, and no such phenotypes have been reported for other BRXf members in the literature (Mouchel et al, 2004;Briggs et al, 2006).…”

“…To begin addressing the mechanisms by which BRXf proteins function collectively in stomatal development, we generated translational reporters to monitor protein dynamics in the leaf epidermis. We chose members of two distinct family subgroups: BRX, because it is has been most extensively characterized (Beuchat et al, 2010b;Marhava et al, 2018;Mouchel et al, 2006;Sankar et al, 2011;Scacchi et al, 2009) and BRXL2 because of its stomatal lineage enrichment and non-redundancy with BRX in the root context (Beuchat et al, 2010a;Briggs et al, 2006). Like the transcriptional reporter, BRXL2p:BRXL2-YFP is detected in MMCs and SLGCs.…”

OBERON3 and SUPPRESSOR OF MAX2 1-LIKE proteins form a regulatory module specifying phloem identity

“…A similar trend was observed for brx;smxl5 double mutants. Like OPS, the BRX gene ensures continues SE formation in this case, however, by downregulation of BAM3 transcription and steepening the auxin gradient in developing phloem cells (Scacchi et al 2010;Depuydt et al 2013;Marhava et al 2018;Marhava et al 2019). Similar to ops;smxl5 double mutants, brx;smxl5 double mutants developed shorter roots than brx and smxl5 single mutants and largely failed to differentiate SE ( Figure 3A, Supplementary Figure 3).…”

“…On the genetic level, smxl4;obe3, smxl5;obe3 and the smxl4;smxl5 double mutants share the same phloem defects meaning that they are deprived of protophloem 15 formation within the RAM. Other reported phloem mutants have either problems with completing phloem differentiation in general, which is the case in mutants of the ALTERED PHLOEM DEVELOPMENT (APL) gene (Bonke et al 2003;Truernit et al 2008;Kondo et al 2016), or they develop 'gap cells' in which SE-differentiation is disturbed as in ops or brx single or in cvp2;cvp2-like1 (cvl1) double mutants (Depuydt et al 2013;Anne et al 2015;Rodriguez-Villalon et al 2015;Marhava et al 2018;Breda et al 2019). In contrast, smxl5;obe3, smxl4;obe3 and multiple mutants of the SMXL3/4/5 genes show absence of all morphological hallmarks of phloem formation within the RAM.…”