Genetic screen for factors mediating PIN polarization in gravistimulated Arabidopsis thaliana hypocotyls

Abstract

Summary Gravitropism is an adaptive response that orients plant growth parallel to the gravity vector. Asymmetric distribution of the phytohormone auxin is a necessary prerequisite to the tropic bending both in roots and shoots. During hypocotyl gravitropic response, the PIN3 auxin transporter polarizes within gravity‐sensing cells to redirect intercellular auxin fluxes. First gravity‐induced PIN3 polarization to the bottom cell membranes leads to the auxin accumulation at the lower side of the organ, initiating bending and, later, auxin feedback‐mediated repolarization restores symmetric auxin distribution to terminate bending. Here, we performed a forward genetic screen to identify regulators of both PIN3 polarization events during gravitropic response. We searched for mutants with defective PIN3 polarizations based on easy‐to‐score morphological outputs of decreased or increased gravity‐induced hypocotyl bending. We identified the number of hypocotyl reduced bending (hrb) and hypocotyl hyperbending (hhb) mutants, revealing that reduced bending correlated typically with defective gravity‐induced PIN3 relocation whereas all analyzed hhb mutants showed defects in the second, auxin‐mediated PIN3 relocation. Next‐generation sequencing‐aided mutation mapping identified several candidate genes, including SCARECROW and ACTIN2, revealing roles of endodermis specification and actin cytoskeleton in the respective gravity‐ and auxin‐induced PIN polarization events. The hypocotyl gravitropism screen thus promises to provide novel insights into mechanisms underlying cell polarity and plant adaptive development.

SCFTIR1/AFB Auxin Signaling for Bending Termination during Shoot Gravitropism

“…Exogenous auxin application also induces PIN3 inner-lateralization, similarly as observed at later stages of gravitropic response. As shown previously (Rakusová et al, 2016(Rakusová et al, , 2019, PIN3-GFP relocated to inner side of endodermal cells after 4 hours of auxin (NAA) treatment ( Fig. 2A, B, H).…”

“…Bending termination involves the reestablishment of auxin-induced symmetrical PIN3 subcellular distribution at later stages (Supplemental Fig. S1A; Rakusová et al, 2016Rakusová et al, , 2019. Therefore, we investigated these processes under conditions of compromised TIR1/AFB auxin signaling.…”

“…We then investigated the involvement of TIR1/AFB pathway in the PIN3 repolarization at later stages of gravitropic response (Rakusová et al, 2016). After 24 hours of gravity stimulation, PIN3-GFP repolarized to inner side of endodermal cells at the bottom side of the wild-type hypocotyl (Figure 1D, G;Rakusová et al, 2016Rakusová et al, , 2019. By contrast, when the www.plantphysiol.org on October 1, 2020 -Published by Downloaded from Copyright © 2020 American Society of Plant Biologists.…”

Switching the Direction of Stem Gravitropism by Altering Two Amino Acids in AtLAZY1

“…Then, in response to the increased auxin concentration, PIN3 moves to the opposite cell side to dissipate the auxin gradient, terminate gravitropic bending, and produce a straightening phase (Rakusová et al, 2016). Genetic and pharmacological results indicate that the cytoskeleton plays a role in the PIN repositioning and possible proprioception required for the straightening response (Rakusová et al, 2011(Rakusová et al, , 2019Okamoto et al, 2015). The region II modification (L92A/I94A) may have impaired the first phase and/or strengthened the second phase, resulting in an overall reverse response (Fig.…”

Directional auxin fluxes in plants by intramolecular domain–domain coevolution of PIN auxin transporters

“…The plant hormone auxin and its graded distribution between cells govern almost all aspects of plant growth and development (Benkov a et al, 2003;Gray, 2004;Vanneste & Friml, 2009;Fendrych et al, 2016;Lavy & Estelle, 2016;Robert et al, 2018), as well as plant adaptive growth in response to environmental cues (Ding et al, 2011;Su et al, 2017;Grones et al, 2018;Rakusov a et al, 2019). Unlike other phytohormones, auxin-dependent plant morphogenesis and adaptabilities rely on auxin concentration gradients that are, besides the contribution of local auxin biosynthesis (Brumos et al, 2018), established by an active, directional cell-to-cell polar auxin transport mediated by auxin transporters, among which, the PIN-FORMED (PIN) auxin exporters play a plethora of developmental roles (Swarup et al, 2001;Blakeslee et al, 2005;Adamowski & Friml, 2015;Park et al, 2017).…”