Strigolactones are common regulators in induction of stomatal closure in planta

ABSTRACT

Strigolactones (SLs) have been implicated in many plant biological processes, including growth and development and the acclimation to environmental stress. We recently reported that SLs intrinsically acted as prominent regulators in induction of stomatal closure. Here we present evidence that the effect of SLs on stotamal closure is not limited to Arabidopsis, and thus SLs could serve as common regulators in the modulation of stomatal apertures of various plant species. Nevertheless, TIS108, a SL-biosynthetic inhibitor, exerted no effect on stomatal apertures. In addition, the SL receptor mutant atd14-5, similar to SL-deficient and more axillary growth 2 (max2) mutants, exhibited hypersensitivity to drought stress. Altogether, these results reinforce the role of SLs as common regulators in stress resilience.

Strigolactones (SLs) are a group of sesquiterpene lactones, which were initially identified as seed germination stimulants for parasitic weeds Striga and Orobanche.^1,2 Their role as branching factor for arbuscular mycorrhizal (AM) fungi was subsequently uncovered. SLs could induce hyphal branching in AM fungi and therefore benefit root colonization.^3,4 Moreover, SLs have a positive role in non-parasitic Arabidopsis germination.⁵ Since SLs could stimulate seed germination at extremely low concentration, it was speculated to be a new class of plant hormones from the very beginning.⁶ It is greatly accepted now that SLs represent a novel class of root-derived hormones and long distance acropetal moving signaling molecules that inhibit shoot branching.^7,8

Except for their prominent roles in shoot architecture, SLs have been found to participate in a wide-range of plant developmental processes including root growth, leaf senescence, photomorphogenesis and flower development.^9,10 For instance, SL-deficient and SL-insensitive mutants exhibited higher density of lateral roots.^11,12 SLs could promote crown root growth,¹³ and inhibit adventitious root formation in rice, Arabidopsis, tomato, and pea.^11,14,15 Additionally, SLs were involved in the response to various external stimuli such as different biotic and abiotic stress.^10,16-18 For instance, SL-deficient and SL-response max mutants exhibited hypersensitivity to drought and salt stress,^19,20 implying a positive role of SLs in stress acclimatization.²¹ Further evidence suggested that SLs and abscisic acid (ABA) were integrated for the acclimatization response.^19,21 A comparative transcriptome analysis indicated that SL controls stress response not only in an ABA-dependent but also in an ABA-independent manner.^17,19

Plants have adopted various strategies to cope with abiotic stress such as increasing drought tolerance by decreasing water loss via stomatal closure. Recently we found that SLs could induce stomatal closure through enhancing hydrogen peroxide synthesis and nitric oxide production and activating SLOW ANION CHANNEL-ASSOCIATED 1 (SLAC1) anion channel,²² thus promoting plant resilience to environmental stress. Nevertheless, the underlying molecular mechanism(s) and intracellular processes which SLs initiate in guard cells require further investigation. In this study, we provide further evidence that SLs could serve as common regulators in stress resilience.

Like Arabidopsis, Vicia faba has long been used to investigate stomatal movement.²³ To determine whether SLs would influence the stomatal aperture in plants other than Arabidopsis, epidermal strips of Vicia faba were incubated with a range of concentrations of the synthetic SL analog GR24 as reported previously.²² We found that GR24 could induce stomatal closure in a dose-dependent manner in Vicia faba. The most significant effect was observed at a concentration of ≥ 1 μM (Fig. 1), which is consistent with the observation in Arabidopsis.²² Taken together, the exogenous application of SLs could trigger stomatal closure in both Arabidopsis and Vicia faba, indicating that SLs act as common regulators in stomatal closure.

Figure 1.

GR24 triggers stomatal closure in Vicia faba. (A) Representative stomata in the absence (left) and presence (right) of 2.5 μM GR24. (B) Stomatal apertures of mock-treated and GR24/ABA-treated Vicia faba for 2 hours. The data are presented as means±SE of three biological replicates. Means with different letters indicate statistically significant differences at P < 0.05.

TIS108 was reported as a novel SL-biosynthetic inhibitor which could reduce the level of SLs in various plant species.^24-26 TIS108-treated Arabidopsis and rice exhibited SL-deficient phenotypes such as more branches and root hair inhibition.^25,26 We next investigated the effect of TIS108 on stomatal closure. Unexpectedly, the exogenous application of TIS108 at different concentrations exerted no effect on stomatal apertures (Fig. 2A), even though it was applied at a high concentration of 50 μM (Fig. 2B). It was reported that TIS108 specifically reduced the level of 2’-epi-5-deoxystrigol (epi-5DS).^24-26 However, it was suggested that unknown molecules in guard cells such as SL-like compound(s) or SL intermediates, rather than epi-5DS, were implicated in the stomatal movement.^22,27 Therefore it is possible that TIS108 could not inhibit the biosynthesis of those molecules in guard cells. Alternatively, TIS108 may fulfill its function in an organ-specific or in a physiological-specific manner.

Figure 2.

Exogenous application of TIS108 exerted no effect on stomatal apertures in Arabidopsis. (A) Stomatal apertures of mock-treated and TIS108-treated Arabidopsis plants under different concentrations. (B) Stomatal apertures of mock-treated and 50 µM TIS108-treated Arabidopsis plants. The data in (A) and (B) are presented as means±SE of three biological replicates at each time point.

The α/β-hydrolase protein DWARF14 (D14) was characterized as a non-canonical SL receptor,²⁸ which was subjected to a conformational change after SL hydrolysis that facilitates interaction with MAX2 to stimulate downstream signaling.^28-31 Previous studies indicated that max2 and SL-deficient mutants showed reduced drought resistance,^17,19,20,27 probably due to wider stomatal apertures of those mutants under drought.^19,20,22,27 Combined with the fact that D14 is involved in the stomatal closure,²² it thereby raised the question of whether D14 contributes to the drought resistance. To this aim, the drought response of atd14-5 mutants, as well as max1-1 and max2-1 mutants using as controls, was examined. We found that atd14-5 showed hypersensitive to drought stress similar to max1-1 and max2-1 (Fig. 3). This observation is in line with a very recent publication that showing the loss-of-function of D14 exhibited reduced drought acclimatization.³² Accordingly, D14-mediated SL signaling is crucial for plant drought response and therefore helps plant to improve stress acclimatization and resistance.

Figure 3.

atd14-5 is hypersensitive to drought stress as max2-1 and max1-1. Two-week-old plants (upper panel) were subjected to drought conditions by withholding water for additional 2 weeks (lower panel).

We conclude that SLs are common regulators in induction of stomatal closure in plants. It thus implies that genetic manipulation of SLs content and signaling could be potentially used for stress tolerance. In agreement with the latest review by Cardinale and colleagues,²¹ the finding of SLs in induction of stomatal closure advances our research on SL biology, and thus raising the demand for unraveling detailed molecular mechanism underlying the SL-mediated stomatal closure. In this regard, the identification of downstream component(s) that mediates the SL signaling in guard cells remains a major subject for further studies. Particularly, to benefit crop protection in realistic fields, an organ-specific dynamics of SL synthesis and perception (especially in guard cells) need to be addressed experimentally in the future.

Funding Statement

National Natural Science Foundation of China (31200902, 31771556, 31271575); the 100-Talent Program of Shaanxi Province; Fundamental Research Funds for the Central Universities (GK201702016); Natural Science Foundation of Hubei Provincial Department of Education (Q20172103); and Initial Project for Post-Graduates of Hubei University of Medicine (2016QDJZR14).

Abbreviation

SLs

Strigolactones

Disclosure of potential conflicts of interest

No potential conflicts of interest were disclosed.

Acknowledgments

Our research is supported by the National Natural Science Foundation of China (31200902, 31771556 and 31271575 to G.W.), by the 100-Talent Program of Shaanxi Province (to G.W.), by the Fundamental Research Funds for the Central Universities (GK201702016 to G.W.), by the Initial Project for Post-Graduates of Hubei University of Medicine (2016QDJZR14 to Y.Z.), and by the Natural Science Foundation of Hubei Provincial Department of Education (Q20172103 to Y.Z.).