SMAX1-LIKE7 signals from the nucleus to regulate shoot development in Arabidopsis via partially EAR motif-independent mechanisms

Abstract

Strigolactones (SLs) are hormonal signals that regulate multiple aspects of shoot architecture, including shoot branching. Like many plant hormonal signaling systems, SLs act by promoting ubiquitination of target proteins and their subsequent proteasome-mediated degradation. Recently, SMXL6, SMXL7, and SMXL8, members of the SMAX1-LIKE (SMXL) family of chaperonin-like proteins, have been identified as proteolytic targets of SL signaling in Arabidopsis thaliana. However, the mechanisms by which these proteins regulate downstream events remain largely unclear. Here, we show that SMXL7 functions in the nucleus, as does the SL receptor, DWARF14 (D14). We show that nucleus-localized D14 can physically interact with both SMXL7 and the MAX2 F-box protein in a SL-dependent manner and that disruption of specific conserved domains in SMXL7 affects its localization, SL-induced degradation, and activity. By expressing and overexpressing these SMXL7 protein variants, we show that shoot tissues are broadly sensitive to SMXL7 activity, but degradation normally buffers the effect of increasing SMXL7 expression. SMXL7 contains a well-conserved EAR (ETHYLENE-RESPONSE FACTOR Amphiphilic Repression) motif, which contributes to, but is not essential for, SMXL7 functionality. Intriguingly, different developmental processes show differential sensitivity to the loss of the EAR motif, raising the possibility that there may be several distinct mechanisms at play downstream of SMXL7. INTRODUCTIONShoot system architectural characteristics strongly influence the productivity of many crop species, and architectural traits have been selected in both historical and contemporary breeding schemes. Understanding the mechanisms that regulate shoot architecture, and its environmental responsiveness, is therefore an important goal for plant research. It is well established that long-distance hormonal signals, including auxin, cytokinin, and strigolactone (SL), are key regulators of shoot architecture and allow communication both within the shoot system and between the shoot and root . For instance, cytokinin produced in the root system in response to the availability of nitrate ions is systemically transported to the shoot, where it promotes branching (Kiba et al., 2011; Müller et al., 2015). Similarly, root-derived SL plays a key role in negatively regulating branching in response to low phosphate availability in the rhizosphere (Kohlen et al., 2011). However, our understanding of the molecular mechanisms that act downstream of these hormones to alter developmental processes in the shoot is currently limited. This is particularly true of SLs. Analysis of the phenotypes of SL biosynthesis and signaling mutants has revealed roles for SLs in the regulation of shoot branching, branching angle, plant height, stem thickness, and leaf blade elongation . The role of SLs in regulating shoot branching has been intensively studied, resulting in two contrasting, nonexclusive models for their mode of action. In the first, SLs are proposed to act locally in axill...

Strigolactone regulates shoot development through a core signalling pathway

“…Remarkably, D53 is a homologue of SMAX1, suggesting that as with KAI2 and D14, different members of the same protein family mediate separable SL and KL signalling activities. Recent studies in Arabidopsis have shown that the co-orthologues of D53, SMAX1-LIKE6 (SMXL6), SMXL7 and SMXL8, have conserved roles as SL targets in the regulation of development (Soundappan et al, 2015; Wang et al, 2015; Liang et al, 2016). This suggests the attractive hypothesis that the SL signalling pathway evolved through duplication and diversification of proteins both upstream and downstream of MAX2.…”

From little things big things grow: karrikins and new directions in plant development

“…D53 is another member of the SMXL family, all of which share similarity to the ClpB/HEAT SHOCK PROTEIN 100 (HSP100) class of heat shock proteins. In Arabidopsis, three D53 homologues (SMXL6, SMXL7 and SMXL8) act redundantly to mediate all tested strigolactonerelated components of MAX2 function (Soundappan et al 2015;Liang et al 2016;Bennett et al 2016). This redundancy might explain why suppressor screens for the increased shoot branching phenotype of max2 had not uncovered this protein family.…”

“…Hydrolysis might also serve as a mechanism for destroying the strigolactone ligand and deactivating the signal. At least in vitro, the hydrolysis product is slowly released, suggesting (Wang et al 2015;Liang et al 2016), but a physical association between KAI2 and SMAX1/SMXL2 is only inferred genetically. In this model, the specific ligand-substrate-repressor associations explain the observed physiological outcomes.…”

Shapeshifters: How Strigolactone Signaling Helps Shape the Shoot

“…Using a powerful mechanistic approach, Liang et al (2016) provide direct, in planta evidence in support of this D14-mediated SL signaling pathway. First, they assessed the subcellular localization of SMXL7 by fusing it to a fluorescent reporter gene and expressing it in Nicotiana benthamiana epidermal cells, finding that it strongly localized to the nucleus.…”

“…Adult shoot morphology in smxl max2 (left) and max2 (right) mutants untransformed (upper panels) and overexpressing SMXL7 (lower panels). (Reprinted from Liang et al [2016] nanometers of each other, a distance sufficiently close for molecular interactions to occur. SMXL7 localized to speckles in the nucleus, and D14 appeared to be recruited to these speckles in response to treatment with a synthetic SL analog.…”