Arabidopsis KASH Proteins SINE1 and SINE2 Are Involved in Microtubule Reorganization During ABA-Induced Stomatal Closure

Abstract

Abscisic acid (ABA) induces stomatal closure by utilizing complex signaling mechanisms, allowing for sessile plants to respond rapidly to ever-changing environmental conditions. ABA regulates the activity of plasma membrane ion channels and calcium-dependent protein kinases, Ca2+ oscillations, and reactive oxygen species (ROS) concentrations. Throughout ABA-induced stomatal closure, the cytoskeleton undergoes dramatic changes that appear important for efficient closure. However, the precise role of this cytoskeletal reorganization in stomatal closure and the nature of its regulation are unknown. We have recently shown that the plant KASH proteins SINE1 and SINE2 are connected to actin organization during ABA-induced stomatal closure but their role in microtubule (MT) organization remains to be investigated. We show here that depolymerizing MTs using oryzalin can restore ABA-induced stomatal closure deficits in sine1-1 and sine2-1 mutants. GFP-MAP4-visualized MT organization is compromised in sine1-1 and sine2-1 mutants during ABA-induced stomatal closure. Loss of SINE1 or SINE2 results in loss of radially organized MT patterning in open guard cells, aberrant MT organization during stomatal closure, and an overall decrease in the number of MT filaments or bundles. Thus, SINE1 and SINE2 are necessary for establishing MT patterning and mediating changes in MT rearrangement, which is required for ABA-induced stomatal closure.

Controlling the Gate: The Functions of the Cytoskeleton in Stomatal Movement

“…During light-induced stomatal opening, oryzalin treatment blocks stomatal opening (Eisinger et al, 2012a;Qu et al, 2017), while stabilization of microtubules by paclitaxel accelerates stomatal opening in a dose-dependent manner (Qu et al, 2017). ABA-, darkness-, and NO-induced stomatal closure was markedly inhibited by cotreatment with paclitaxel, but no significant changes were observed when oryzalin was applied (Zhang et al, 2008;Eisinger et al, 2012a;Qu et al, 2017;Biel et al, 2020). It has also been reported that treatment with oryzalin alone affects stomatal closure (Khanna et al, 2014).…”

“…Live-cell imaging revealed that the number and arrangement pattern of MTs in guard cells are correlated with the stomatal aperture. In open stomata, MTs radiate from the ventral side to the dorsal side in a more parallel, straighter and denser fashion relative to AF organization ( Eisinger et al, 2012a ; Qu et al, 2017 ; Biel et al, 2020 ). Following stomatal closing, MT structures decrease in number and become diffused.…”

“…Following stomatal closing, MT structures decrease in number and become diffused. Some studies have shown that MTs completely depolymerized ( Qu et al, 2017 ; Yu et al, 2020 ), while other studies have indicated that MTs are still present, but with reduced density, in closed stomata ( Khanna et al, 2014 ; Biel et al, 2020 ; Yu et al, 2020 ; Dou et al, 2021 ). In the latter case, observable MTs tend to have a longitudinal arrangement and become crisscrossed or randomly patterned near the ventral side ( Fukuda et al, 1998 ; Lahav et al, 2004 ; Zhang et al, 2008 ; Eisinger et al, 2012a ; Biel et al, 2020 ).…”

“…Some studies have shown that MTs completely depolymerized ( Qu et al, 2017 ; Yu et al, 2020 ), while other studies have indicated that MTs are still present, but with reduced density, in closed stomata ( Khanna et al, 2014 ; Biel et al, 2020 ; Yu et al, 2020 ; Dou et al, 2021 ). In the latter case, observable MTs tend to have a longitudinal arrangement and become crisscrossed or randomly patterned near the ventral side ( Fukuda et al, 1998 ; Lahav et al, 2004 ; Zhang et al, 2008 ; Eisinger et al, 2012a ; Biel et al, 2020 ). By using end-binding protein 1 (EB1) to label the growing plus ends of microtubules, it was observed that there were no significant changes in the number of growing ends or the growth velocity rate of microtubules during stomatal closure.…”

“…SINE1 and SINE2 (SUN-INTERACTING NUCLEAR ENVELOPE PROTEIN 1 AND2), which are two components of the plant LINC (LINKER OF NUCLEOSKELETON AND CYTOSKELETON) complex, are involved in regulating the reorganization of MTs in guard cells during stomatal movement. The loss of function of either SINE1 or SINE2 results in a disordered MT organization in open stomata ( Biel et al, 2020 ). There are fewer MT filaments or bundles in sine 1 or sine 2 mutants compared to the wild type during stomatal closure, leading to insensitivity to ABA-induced stomatal closure ( Biel et al, 2020 ).…”