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. 2015 Jul 28;169(1):371–378. doi: 10.1104/pp.15.00788

The VQ Motif-Containing Protein Family of Plant-Specific Transcriptional Regulators1

Yanjun Jing 1, Rongcheng Lin 1,*
PMCID: PMC4577417  PMID: 26220951

Recent advances highlight the roles and mechanisms of the plant-specific VQ-motif-containing protein family in regulating stress and developmental processes.

Abstract

The VQ motif-containing proteins (designated as VQ proteins) are a class of plant-specific proteins with a conserved and single short FxxhVQxhTG amino acid sequence motif. VQ proteins regulate diverse developmental processes, including responses to biotic and abiotic stresses, seed development, and photomorphogenesis. In this Update, we summarize and discuss recent advances in our understanding of the regulation and function of VQ proteins and the role of the VQ motif in mediating transcriptional regulation and protein-protein interactions in signaling pathways. Based on the accumulated evidence, we propose a general mechanism of action for the VQ protein family, which likely defines a novel class of transcriptional regulators specific to plants.

Transcriptional gene regulation is central to a pyramid of biological processes in all living organisms. As sessile organisms, plants have evolved many specific families of transcription factors that bind to the promoter regions of downstream genes and initiate their transcription (Shiu et al., 2005; Qu and Zhu, 2006; Yamasaki et al., 2013). Transcription regulators that do not bind DNA often work in concert with transcription factors via protein-protein interactions to fine-tune the regulatory machinery in response to surrounding environments, such as biotic stress and light stimuli (Wray et al., 2003; Yamaguchi-Shinozaki and Shinozaki, 2006; Jiao et al., 2007; Buscaill and Rivas 2014). A group of proteins designated as VQ motif-containing proteins (referred to here as VQ proteins) was recently identified (Xie et al., 2010; Cheng et al., 2012). Since the discovery of the first VQ protein, SIGMA FACTOR-BINDING PROTEIN1 (SIB1, also known as VQ23; Morikawa et al., 2002), nine VQ proteins have been functionally characterized. These factors play diverse roles in plant defense response, stress tolerance, and growth and development. Here, we review the developmental and physiological roles, regulation, and molecular mechanisms of VQ proteins and highlight the importance of the VQ motif in mediating protein-protein interactions and transcriptional activity.

STRUCTURAL FEATURES OF VQ PROTEINS

The short VQ motif possesses five conserved amino acids in its core sequence FxxhVQxhTG, where x represents any amino acid and h denotes a hydrophobic residue (Fig. 1; Pecher et al., 2014). Twenty-five, 39, and 34 VQ members were identified in a moss (Physcomitrella patens), rice (Oryza sativa), and Arabidopsis (Arabidopsis thaliana), respectively (Kim et al., 2013; Li et al., 2014). Interestingly, most VQ genes in higher plants do not have an intron, whereas 18 VQ genes contain at least one intron in moss. There are two splice variants for Arabidopsis VQ2, VQ4, VQ6, VQ14, VQ31, and VQ32. The short forms of VQ2 and VQ14 encode proteins that do not contain the VQ motif, whereas two splice forms of VQ4 and VQ31 generate proteins consisting of the VQ motif (Fig. 2). There is an intron in the promoter region of VQ6 and VQ32; thus, the two variants produce identical proteins. Furthermore, moss VQ proteins are relatively large, most being longer than 300 amino acids. By contrast, Arabidopsis and rice VQ proteins are small, with the majority consisting of 98 to 300 amino acid residues (Fig. 2; Li et al., 2014). Most Arabidopsis VQ proteins are predicted to localize to the nucleus, some localize to plastids, and a few localize to mitochondria solely from bioinformatic prediction (Fig. 2; Cheng et al., 2012). Experimental evidence is required to confirm the subcellular localization in future studies. Based on a phylogenetic analysis and structural features of the VQ domain or full-length sequences, the Arabidopsis VQ proteins were classified into seven or 10 groups, respectively (Kim et al., 2013; Pecher et al., 2014; Fig. 2). The primary structure of VQ proteins is highly diverse in regions other than the VQ motif, suggesting that VQ proteins have distinct interacting partners. Little significant similarity has been identified between VQ proteins of plants and sequenced or predicted proteins in other organisms, indicating that the VQ family is specific to plants.

Figure 1.

Figure 1.

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The consensus sequence of the VQ motif. The sequences of the 31 amino acids of the VQ motif (pfam05678; Marchler-Bauer et al., 2015) were extracted from 34 Arabidopsis, 39 rice, and 25 moss VQ proteins, and the graph was generated by WebLogo (http://weblogo.berkeley.edu/; Crooks et al., 2004). The overall height of the stack indicates the sequence conservation at that position, while the height of the symbols within the stack indicates the relative frequency of each amino acid at that position.

Figure 2.

Figure 2.

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Summary of 34 Arabidopsis VQ proteins. Protein lengtha, Based on data from The Arabidopsis Information Resource (www.arabidopsis.org). Localizationb, Predicted (adopted from Cheng et al. [2012]; the Subcellular Proteomic Database [http://suba.plantenergy.uwa.edu.au]) or supported by experiments. Transcriptional activityc, Based on the transient expression assay of Li et al. (2014). Groupd, On the basis of the whole sequence similarities, the VQ proteins are classified into 10 groups (Pecher et al., 2014). Blue boxes denote VQ motifs. AGI, Arabidopsis Genome Initiative; M, mitochondria; N, nucleus; P, plastid.

VQ PROTEINS FUNCTION IN THE PLANT DEFENSE RESPONSE

Plants have evolved effective defense systems to protect themselves from biotic stresses. Salicylic acid (SA) and jasmonic acid (JA) are two of the best-known defense signaling molecules that accumulate upon pathogen infection to trigger defense responses (Glazebrook, 2005). Increasing evidence demonstrates the involvement of VQ proteins in SA- and/or JA-mediated defense responses.

Plants with loss of function of VQ23/SIB1 have compromised resistance to both Botrytis cinerea and Pseudomonas syringae, whereas VQ23 overexpression lines showed reduced disease symptoms after infection with either pathogen. The increased resistance of VQ23 overexpression plants to B. cinerea is dependent on functional WRKY33 (Xie et al., 2010; Lai et al., 2011). Thus, VQ23 positively regulates the plant defense response against both necrotrophic and biotrophic pathogens likely via a WRKY-dependent signaling pathway. However, it was reported that VQ23 overexpression plants do not differ from the control with respect to their sensitivity to the virulent strain P. syringae pv tobacco (Pst) DC3000 (Narusaka et al., 2008). VQ16/SIB2 plays a redundant role in response to B. cinerea infection with VQ23 (Lai et al., 2011), consistent with their evolutionary relatedness in VQ group VI.

VQ21/MITOGEN-ACTIVATED PROTEIN KINASE4 SUBSTRATE1 (MKS1) was first discovered as a substrate of MITOGEN-ACTIVATED PROTEIN KINASE4 (MPK4) kinase in a yeast two-hybrid screen downstream of MPK4 (Andreasson et al., 2005; Petersen et al., 2010). VQ21 is required for full SA-dependent resistance to Pst DC3000 in mpk4 mutants, and VQ21 overexpression in wild-type plants is sufficient to activate SA-dependent resistance but does not interfere with the JA pathway (Andreasson et al., 2005). However, it was later found that overexpression of VQ21 leads to susceptibility to B. cinerea (Petersen et al., 2010; Fiil and Petersen, 2011). Moreover, vq21/mks1 knockout mutants exhibited increased susceptibility to bacterial strains of P. syringae and Hyaloperonospora arabidopsidis (Petersen et al., 2010). These studies suggest that VQ21 plays a positive role in SA-regulated defense against biotrophic pathogens but a negative role in JA-mediated immunity against necrotrophic pathogens. Consistently, MPK4 is required for both the repression of SA-dependent resistance and the activation of JA-dependent defense gene expression (Petersen et al., 2000). A recent study revealed that MPK3/6-TARGETED VQ MOTIF-CONTAINING PROTEIN1 (MVQ1, also known as VQ4) acts as a negative regulator of pathogen-associated molecular pattern-induced responses against Pst DC3000 (Pecher et al., 2014). Nine other MVQ proteins might have a similar role to VQ4.

JASMONATE-ASSOCIATED VQ MOTIF GENE1 (JAV1, also known as VQ22) was identified in a forward genetic screen for components that specifically regulate jasmonate-mediated plant defense. Silencing of VQ22 enhanced resistance to necrotrophic pathogens and herbivorous insects, whereas transgenic lines constitutively expressing VQ22 were highly susceptible to these biotic stresses (Hu et al., 2013a), indicating that VQ22 functions as a negative regulator of JA-mediated plant defense. Therefore, members of the VQ family play either positive or negative roles in SA- and/or JA-mediated plant immune responses, possibly by interacting with different WRKY transcription factors and mediating distinct activities (see below).

VQ PROTEINS IN ABIOTIC STRESS RESPONSES

VQ15 and VQ9 were reported to modulate abiotic stress tolerance (Perruc et al., 2004; Hu et al., 2013b). A transient increase in cytosolic free calcium is one of the earliest events in response to abiotic stresses (Knight, 2000). Calmodulin (CaM) couples Ca2+ signals to changes in the activity of downstream effectors by interacting with other binding proteins. Perruc et al. (2004) identified Arabidopsis CaM-binding protein of 25 kD (AtCAMBP25), encoded by VQ15, as a partner of CaM in response to osmotic stress. VQ15 localizes to the nucleus and interacts with CaM in a calcium-dependent manner. Transgenic plants overexpressing VQ15 display increased sensitivity to both NaCl- and mannitol-induced osmotic stresses during seed germination and seedling growth. By contrast, plants harboring antisense VQ15 exhibit increased tolerance to osmotic stress (Perruc et al., 2004). This inverse relationship between VQ15 transcript level and the stress tolerance indicates that VQ15 acts as a negative regulator in osmotic stress. It is assumed that VQ24, its closely related protein in VQ group X, might have a similar function.

The WRKY8 transcription factor plays a positive role in salt stress tolerance (Hu et al., 2013b). VQ9 was identified as an interacting protein of WRKY8 in a yeast two-hybrid system, and the VQ9-GFP fusions were targeted exclusively to the nucleus. However, mutation of VQ9 exhibited an opposite response to that of WRKY8 (Hu et al., 2013b). Thus, VQ9 may act as a transcriptional repressor that antagonizes WRKY8 during salt stress.

VQ PROTEINS IN PLANT GROWTH AND DEVELOPMENT

Several studies have revealed that a number of VQ proteins control diverse plant growth and developmental processes. VQ14/HAIKU1 (IKU1) regulates the expression of MINISEED3 (MINI3; encoding a WRKY transcription factor) and IKU2 (encoding a Leu-rich repeat kinase), which are important for seed development (Garcia et al., 2003; Luo et al., 2005). The finding that the recessive vq14/iku1 mutant displays a small-seed phenotype indicates that VQ14 positively regulates endosperm growth and seed development. In line with this, VQ14 is expressed preferentially in the early endosperm and localizes to the nuclei of the syncytial endosperm (Wang et al., 2010).

Our laboratory recently reported the functional characterization of VQ29, which negatively regulates seedling photomorphogenesis (Li et al., 2014). Overexpression of VQ29 results in the hyposensitivity of hypocotyl growth to far-red light and low-light conditions, whereas the vq29 loss-of-function mutant exhibits decreased hypocotyl elongation under low-intensity far-red light and white light (Li et al., 2014). Thus, the VQ family is also involved in regulating the light-mediated response and light signaling pathway.

A loss-of-function mutant harboring a transposon tag in vq8 displayed a pale-green and stunted growth phenotype throughout its life cycle (Cheng et al., 2012). It is likely that VQ8 regulates chloroplast development or photosystem assembly, consistent with its predicted localization to plastids. Interestingly, a recent study reported that the heterologous expression of Arabidopsis VQ21 in Kalanchoe blossfeldiana and Petunia hybrida resulted in dwarfed phenotypes and delayed flowering in both species. The transgenic K. blossfeldiana flowers had increased anthocyanin content, and the length of the inflorescence stem was decreased (Gargul et al., 2015).

MODE OF ACTION OF VQ PROTEINS

Accumulating studies indicate that many VQ proteins interact with WRKY transcription factors (Chi et al., 2013). VQ23 and VQ16 recognize the C-terminal WRKY domain and stimulate the DNA-binding activity (but not the transcription-activating activity) of WRKY33 (Fig. 3A). Consistently, VQ23 coprecipitated with the WRKY33/DNA complex in an electrophoretic mobility shift assay, and VQ23 and VQ16 exhibit transcriptional activation properties (Lai et al., 2011; Li et al., 2014). By contrast, the interaction between VQ9 and WRKY8 represses the DNA-binding activity of WRKY8 (Fig. 3B; Hu et al., 2013b). Moreover, a bimolecular fluorescence complementation assay demonstrated that VQ14 interacts with the WRKY transcription factor MINI3 and that their interaction likely regulates the expression of the downstream gene IKU2 (Fig. 3C; Wang et al., 2010). In addition, VQ22 may interact with WRKY28 and WRKY51 to negatively control JA-mediated defense (Hu et al., 2013a). Two systematic yeast two-hybrid assays showed that a large proportion of VQ proteins associate physically with members of group I or group IIc WRKY transcription factors (Table I; Rushton et al., 2010; Cheng et al., 2012; Pecher et al., 2014). This interaction with WRKY members was further validated for VQ4 in Arabidopsis protoplasts (Weyhe et al., 2014). The combinatorial complexity of VQ-WRKY interactions might allow sophisticated transcriptional regulation of downstream targets and, consequently, diverse physiological responses. WRKY proteins play crucial roles in plant defense responses (Pandey and Somssich, 2009). Other uncharacterized VQ proteins that interact with WRKYs might also be involved in mediating stress responses or even growth and development. Consistent with this interpretation, the growth of F1 crosses that overexpress both VQ10 and either WRKY25 or WRKY33 was greatly retarded, indicating that VQ10 coordinates with WRKY proteins to repress plant growth (Cheng et al., 2012).

Figure 3.

Figure 3.

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Transcriptional regulatory mode of VQ proteins. A, VQ23 interacts with WRKY33 and stimulates its binding to the promoter region of defense-responsive genes. B, The interaction between VQ9 and WRKY8 inhibits WRKY8 from binding to the promoter regions of its target genes during salt stress tolerance. C, VQ14 and MINI3 interact and coregulate the abundance of mRNAs encoding proteins involved in seed development. D, The interaction between VQ4 and WRKYs inhibits WRKYs from binding to their target genes in the plant defense response. MPK3/6 phosphorylate VQ4 and induce its degradation. E, After pathogen infection, MPK4 is activated and phosphorylates VQ21, triggering the release of VQ21 and WRKY33, which binds to the promoter of defense-responsive genes, such as PAD3, and activates their expression. VQ21 may optimize the transcriptional activation of WRKY33. In D and E, p denotes phosphorylated residues at multiple sites. F, VQ29 and PIF1 interact directly to regulate XTR7 expression and cell elongation.

Table I. The interacting factors and physiological functions of Arabidopsis VQ proteins.

VQ proteins with characterized developmental and physiological roles are listed. Interacting partners shown in boldface were verified by pull-down, coimmunoprecipitation, or bimolecular fluorescence complementation assays in addition to yeast two-hybrid assay.

Arabidopsis Genome Initiative No. Name Other Name Interacting Partners Functions References
AT1G28280 VQ4 MVQ1 MPK3/6/10, WRKY33, WRKY68 Defense (P. syringae) Pecher et al. (2014); Weyhe et al. (2014)
AT1G78310 VQ9 MVQ10 WRKY8, WRKY20, MPK3/4/6/10/11 Salinity stress tolerance Hu et al. (2013b); Pecher et al. (2014)
AT1G78410 VQ10 WRKY25/26/33 Vegetative growth Cheng et al. (2012)
AT2G35230 VQ14 IKU1/ MVQ9 MINI3 Seed size Wang et al. (2010)
AT2G41010 VQ15 CAMBP25 CaM, WRKY25, WRKY51 Osmotic stress tolerance Perruc et al. (2004); Cheng et al. (2012)
AT2G41180 VQ16 SIB2 WRKY33, WRKY25, MPK1-18/20 Defense (B. cinerea) Lai et al. (2011); Pecher et al. (2014)
AT3G18690 VQ21 MKS1 MPK4, MPK11, WRKY25, WRKY33 Defense (B. cinerea and P. syringae) Andreasson et al. (2005); Petersen et al. (2010); Pecher et al. (2014)
AT3G22160 VQ22 JAV1 WRKY28, WRKY51 JA-mediated defense Hu et al. (2013a)
AT3G56710 VQ23 SIB1 WRKY33, SIG1, MPK6/14/16, WRKY3/4/20/25 SA- and JA-mediated defense Morikawa et al. (2002); Narusaka et al. (2008); Xie et al. (2010); Lai et al. (2011); Pecher et al. (2014)
AT4G37710 VQ29 PIF1 Photomorphogenesis Li et al. (2014)
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The VQ-WRKY partnership becomes even more complicated with additional signaling components. For instance, the VQ4-WRKY interaction inhibits WRKY function, and this effect is released by MPK3/6-triggered phosphorylation of VQ4 and its degradation (Fig. 3D; Pecher et al., 2014). It was proposed alternatively that VQ4 has a positive regulatory effect on gene expression that can be antagonized by WRKYs (Pecher et al., 2014). Similar to VQ4, VQ21 acts as an adaptor that links MPK4 kinase to WRKY33 by interacting with both proteins. However, activated MPK4 phosphorylates VQ21, leading to the release of VQ21 and WRKY33 from MKP4. Consequently, WRKY33 binds to the promoter of PHYTOALEXIN DEFICIENT3 (PAD3), which encodes an enzyme required for the synthesis of antimicrobial camalexin (Andreasson et al., 2005; Qiu et al., 2008). This mechanism may ensure that VQ21 facilitates the recruitment of the kinase to WRKY factors to activate the MPK4-regulated signaling pathway (Fig. 3E). Hence, a triangular relationship between VQ proteins, WRKYs, and mitogen-activated protein kinases was proposed (Weyhe et al., 2014). Such an intricate network consisting of tripartite interacting factors can provide more differential, precise, and effective regulatory mechanisms for plants in response to environmental stimuli. Similarly, VQ15 associates with CaM, WRKY25, and WRKY51 and likely connects Ca2+ signals to the transcriptional regulation of downstream targets in the osmotic stress signaling pathway (Perruc et al., 2004; Cheng et al., 2012). It is speculated that VQ proteins with putative localization in plastid or mitochondria possibly mediate transcription regulation by associating with other components in these organelles. Intriguingly, a previous pull-down assay revealed that VQ23 interacts physically with SIGMA FACTOR1, which is a component of the plastid-encoded RNA polymerase complexes and plays essential roles in regulating transcription in plastids (Morikawa et al., 2002).

There is additional evidence of VQ proteins interacting with transcription factors that function in the nucleus other than those of the WRKY family. For example, VQ29 interacts physically with the basic helix-loop-helix transcription factor PHYTOCHROME-INTERACTING FACTOR1 (PIF1), and these two proteins associate with the promoter of a cell elongation-related gene, XYLOGLUCAN ENDOTRANSGLYCOSYLASE7 (XTR7), to coactivate its expression in response to light environmental signals (Fig. 3F; Li et al., 2014). It is anticipated that some members of the VQ family might be recruited by other transcription factors to modulate distinct responses.

The interaction between VQ proteins and transcription factors suggests that VQ proteins might possess transcriptional activity. A transient expression assay using Arabidopsis mesophyll protoplasts reveals that, except for VQ1, VQ10, VQ22, VQ25, and VQ27, all members of the Arabidopsis VQ family facilitate either transcriptional activation or repression (Li et al., 2014), likely through the binding of specific transcription factors to cis-elements or other coregulators. In agreement with this, genome-wide profiling demonstrated that VQ21 and VQ22 regulate numerous defense-responsive genes in the SA- and JA-mediated signaling pathway, respectively (Andreasson et al., 2005; Hu et al., 2013a). This notion is further supported by the nuclear localization of some VQ proteins fused with GFP. Interestingly, VQ proteins in group I (MVQ1–MVQ6) possess repression activity, while those from groups II, VI, and X all have activation activity, and members in group VIII do not have transcriptional activity (Fig. 2), suggesting a similar regulatory mode within these individual groups.

In summary, the findings that VQ proteins (1) are mostly targeted to the nucleus, (2) interact with transcription factors and regulate their activities, (3) largely exhibit transcriptional activity, and (4) do not appear to bind to DNA suggest that they likely act as transcription regulators that modulate downstream gene expression through interacting with transcription factors and/or other coregulators. Those VQ proteins lacking transcriptional activity possibly function as cofactors (e.g. to facilitate protein-protein interaction or protein stability).

REGULATION OF VQ PROTEINS

VQ proteins are regulated at several levels. The transcription of VQ genes is modulated by multiple endogenous and environmental signals, consistent with their diverse roles in stress responses and during plant growth and development. For instance, B. cinerea infection and SA treatment rapidly and strongly induce VQ23 and VQ16 transcript levels (Narusaka et al., 2008; Xie et al., 2010; Lai et al., 2011). VQ22 transcript is significantly increased upon JA treatment and quickly accumulates after mechanical wounding (Hu et al., 2013a). Moreover, VQ15 is induced in seedlings exposed to various abiotic stresses, including dehydration, low temperature, or high salinity, while VQ9 expression is increased by NaCl stress treatment (Perruc et al., 2004; Hu et al., 2013b). Furthermore, light represses VQ29 in a phytochrome-dependent manner (Li et al., 2014). A tomato (Solanum lycopersicum) VQ homolog, Avirulence9 Rapidly Elicited169, is rapidly induced by the race-specific elicitor Avirulence9 (Durrant et al., 2000). In addition, expression profiles revealed that many of the Arabidopsis VQ genes were responsive to pathogen infection and SA treatment (Cheng et al., 2012). Similarly, rice VQ genes are also regulated by biotic and abiotic stresses. The transcript levels of a set of rice VQ genes are affected by infection with compatible and incompatible bacterial strains of Xanthomonas oryzae pv oryzae, treatment with abscisic acid, or exposure to drought (Kim et al., 2013), indicating that rice VQ proteins are likely important coregulators in response to environmental stimuli.

Several studies suggest that VQ proteins are also under the control of posttranslational regulation. For example, VQ21 is a substrate of MPK4 and directly interacts with MPK4 in a coimmunoprecipitation assay (Andreasson et al., 2005; Petersen et al., 2010). Activation of MPK4 triggers the phosphorylation of VQ21 at multiple Ser residues, including Ser-72, Ser-108, and Ser-120, as identified by mass spectrometry. VQ21 phosphorylation causes VQ21 and WRKY33 to deassociate from MPK4 (Andreasson et al., 2005; Caspersen et al., 2007). Furthermore, several VQ proteins, including VQ4, VQ13, VQ33, VQ19, VQ11, VQ31, and VQ9, are phosphorylated by MPK3 and/or MPK6, and 22-amino acids flagellin peptide (flg22) elicitor treatment of protoplasts causes a pronounced mobility shift of these proteins, consistent with the higher numbers of predicted mitogen-activated protein kinase phosphorylation sites in these proteins (Pecher et al., 2014). Detailed mutagenesis and phosphorylation analyses revealed that MPK6 targets and phosphorylates 12 Ser/Thr phosphorylation sites of VQ4 (Pecher et al., 2014).

Phosphorylation often leads to the targeted degradation of a protein by the ubiquitin-proteasome pathway (Henriques et al., 2009). Pecher et al. (2014) observed that the protein abundance of Arabidopsis VQ4, VQ13, VQ6, VQ14, and VQ9 was reduced in protoplasts after flg22 treatment. They further found that the rapid flg22-induced VQ4 reduction was abrogated in mutants in which all 12 phosphorylation sites were mutated, indicating that phosphorylation of VQ4 promotes its destabilization. Furthermore, a recent study showed that methyl jasmonate treatment induced VQ22 degradation, which was attenuated by the 26S proteasome inhibitor MG132. Wounding also triggered the degradation of VQ22 in the wild type but not in coronatine insensitive1 (coi1) mutant plants. These results demonstrate that jasmonate or wounding triggers the degradation of VQ22 via the 26S proteasome pathway in a COI1-dependent manner (Hu et al., 2013a). The degradation of VQ22 upon insect attack or pathogen infection likely releases its inhibitory role on positive effectors of plant defense responses (such as WRKY transcription factors). Since COI1 does not interact with VQ22, the E3 ligase mediating VQ22 degradation remains to be identified. We anticipate that some other VQ proteins might also be subjected to posttranslational regulation.

MOLECULAR FUNCTION OF THE VQ MOTIF

The conserved sequence of the VQ motif indicates that it possesses essential roles. Site-directed mutagenesis of the VQ motif helped to dissect the function and mode of action of VQ proteins. For example, the VQ motif was found to be important for VQ14 function. Transgenes harboring mutations in the VQ motif (IVQQ to EDLE) of VQ14 failed to rescue the vq14/iku1 seed phenotype. However, mutations in other conserved regions did not affect the function of VQ14 (Wang et al., 2010).

Several lines of evidence suggest that the VQ motif mediates protein localization and the interaction between VQ proteins and other regulators. First, mutation in the VQ motif altered the nuclear localization of VQ9 (Hu et al., 2013b). The region of VQ21 containing the VQ motif is also needed for its nuclear localization (Petersen et al., 2010). However, this is not the case for VQ23, as substitutions in the VQ motif did not change the subcellular targeting of VQ23 (Lai et al., 2011). This might be because VQ23 localizes to both the nucleus and chloroplast, due to its chloroplast-targeting signal peptide and nuclear localization signal outside the VQ motif.

Second, the VQ motif in VQ9 and VQ23 is essential for the interaction with their partners. Mutations in the VQ motif (VVQK to EDLE) of VQ9 abolished the interaction with WRKY8, whereas substitutions in VQ23 (VQ to AA) impaired the VQ23-WRKY33 interaction (Lai et al., 2011; Hu et al., 2013b). Although truncation of the N-terminal portions containing the VQ motif abolished the VQ14-MINI3 and VQ21-WRKY33 interactions (Petersen et al., 2010; Wang et al., 2010), the role of the VQ motif in mediating these interactions remains to be determined. Most intriguingly, the VQ motif in MVQ proteins is essential for their interaction with the WRKY transcription factors, but it is not necessary for their interaction with or phosphorylation by MPK3/MPK6 (Pecher et al., 2014). However, two amino acid replacements in VQ29 (V70D and Q71L) did not affect its interaction with PIF1 (Li et al., 2014). Therefore, the role of the VQ motif in modulating protein-protein interactions varies.

Third, a single amino acid substitution in VQ29 (V70A or V70D) abolished its repressive activity, indicating that the VQ motif is required for mediating the transcriptional activity of VQ29. Interestingly, double substitutions in the VQ motif (V70D and Q71L) led to a significant induction of the LUCIFERASE reporter gene driven by the 35S minimal promoter fused with a GAL4-binding sequence (Li et al., 2014). Similarly, the VQ motif variant VQ4DL had a significant induction of the stress promoter-driven LUCIFERASE reporter expression (Pecher et al., 2014). These studies suggest that the transcriptional activity could be switched from repression to activation or the reverse through modifying the VQ motif, and they further imply that recruitment of transcription factors/regulators might control this regulation. In addition, VQ2 and VQ14 were predicted to undergo alternative splicing that results in proteins without the VQ motif (Fig. 2). This posttranscriptional regulation, therefore, offers a possible regulatory role through VQ domain exclusion.

CONCLUSION AND PERSPECTIVES

Although much progress has been made in understanding the role of VQ proteins in Arabidopsis, our knowledge of the functions and mechanisms of action of this family remains limited. Based on the available information, we propose that VQ proteins constitute a group of transcription regulators that interact with transcription factors to modulate downstream gene expression. Furthermore, these proteins may interact with other proteins or protein complexes to form regulatory modules, such as WRKY-VQ-MPK, and fine-tune gene transcription. Consequently, members of the VQ protein family either positively or negatively regulate multiple responses, including plant immunity, abiotic stresses, and growth and development, and the expression of their underlying genes is altered in response to either internal or external signals.

The physiological relevance and molecular mechanisms of most of the VQ genes remain to be characterized. Some closely related VQ protein genes, such as VQ1 and VQ10 (group VIII), VQ24 and VQ15 (group X), and VQ12 and VQ29 (group IV), may be functionally redundant (Cheng et al., 2012). Generating double or higher order mutants or artificial microRNA that targets multiple genes in one group could help to unravel the roles of these genes. Moreover, previous studies showed that many VQ proteins interact with WRKY transcription factors in yeast cells, indicating that these factors may also be involved in biotic or abiotic responses. These findings highlight the importance of this protein family in regulating plant immunity and stress tolerance that require further investigation. However, it should be noted that only a subset of the VQ proteins interact with WRKYs, as well as MPK3 and MPK6, and that some members of this family mediate plant growth and development, with roles in seed development, seedling growth, and plant architecture. Determining the other roles, if any, of the remaining VQ genes will be of great interest. It was recently shown that the heterologous overexpression of Arabidopsis VQ21 in K. blossfeldiana and P. hybrida altered the growth and development and also the disease tolerance of the transgenic plants (Gargul et al., 2015). Characterizing the function and mode of action of VQ genes in other species will provide insight into the evolutionary role of this plant-specific protein family. Likewise, manipulating the expression of VQ genes might be a strategy to increase crop productivity or adaptation to suboptimal environments.

Glossary

SA

salicylic acid

JA

jasmonic acid

Pst

Pseudomonas syringae pv tobacco

CaM

calmodulin

Footnotes

1

This work was supported by the National Natural Science Foundation of China (grant nos. 31170221 and 31325002 to R.L.) and the Ministry of Agriculture of China (grant no. 2014ZX08009–003 to R.L.).

References

  1. Andreasson E, Jenkins T, Brodersen P, Thorgrimsen S, Petersen NHT, Zhu S, Qiu JL, Micheelsen P, Rocher A, Petersen M, et al. (2005) The MAP kinase substrate MKS1 is a regulator of plant defense responses. EMBO J 24: 2579–2589 [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Buscaill P, Rivas S (2014) Transcriptional control of plant defence responses. Curr Opin Plant Biol 20: 35–46 [DOI] [PubMed] [Google Scholar]
  3. Caspersen MB, Qiu JL, Zhang X, Andreasson E, Naested H, Mundy J, Svensson B (2007) Phosphorylation sites of Arabidopsis MAP kinase substrate 1 (MKS1). Biochim Biophys Acta 1774: 1156–1163 [DOI] [PubMed] [Google Scholar]
  4. Cheng Y, Zhou Y, Yang Y, Chi YJ, Zhou J, Chen JY, Wang F, Fan B, Shi K, Zhou YH, et al. (2012) Structural and functional analysis of VQ motif-containing proteins in Arabidopsis as interacting proteins of WRKY transcription factors. Plant Physiol 159: 810–825 [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Chi Y, Yang Y, Zhou Y, Zhou J, Fan B, Yu JQ, Chen Z (2013) Protein-protein interactions in the regulation of WRKY transcription factors. Mol Plant 6: 287–300 [DOI] [PubMed] [Google Scholar]
  6. Crooks GE, Hon G, Chandonia JM, Brenner SE (2004) WebLogo: a sequence logo generator. Genome Res 14: 1188–1190 [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Durrant WE, Rowland O, Piedras P, Hammond-Kosack KE, Jones JD (2000) cDNA-AFLP reveals a striking overlap in race-specific resistance and wound response gene expression profiles. Plant Cell 12: 963–977 [DOI] [PMC free article] [PubMed] [Google Scholar]
  8. Fiil BK, Petersen M (2011) Constitutive expression of MKS1 confers susceptibility to Botrytis cinerea infection independent of PAD3 expression. Plant Signal Behav 6: 1425–1427 [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Garcia D, Saingery V, Chambrier P, Mayer U, Jürgens G, Berger F (2003) Arabidopsis haiku mutants reveal new controls of seed size by endosperm. Plant Physiol 131: 1661–1670 [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Gargul JM, Mibus H, Serek M (2015) Manipulation of MKS1 gene expression affects Kalanchoë blossfeldiana and Petunia hybrida phenotypes. Plant Biotechnol J 13: 51–61 [DOI] [PubMed] [Google Scholar]
  11. Glazebrook J. (2005) Contrasting mechanisms of defense against biotrophic and necrotrophic pathogens. Annu Rev Phytopathol 43: 205–227 [DOI] [PubMed] [Google Scholar]
  12. Henriques R, Jang IC, Chua NH (2009) Regulated proteolysis in light-related signaling pathways. Curr Opin Plant Biol 12: 49–56 [DOI] [PubMed] [Google Scholar]
  13. Hu P, Zhou W, Cheng Z, Fan M, Wang L, Xie D (2013a) JAV1 controls jasmonate-regulated plant defense. Mol Cell 50: 504–515 [DOI] [PubMed] [Google Scholar]
  14. Hu Y, Chen L, Wang H, Zhang L, Wang F, Yu D (2013b) Arabidopsis transcription factor WRKY8 functions antagonistically with its interacting partner VQ9 to modulate salinity stress tolerance. Plant J 74: 730–745 [DOI] [PubMed] [Google Scholar]
  15. Jiao Y, Lau OS, Deng XW (2007) Light-regulated transcriptional networks in higher plants. Nat Rev Genet 8: 217–230 [DOI] [PubMed] [Google Scholar]
  16. Kim DY, Kwon SI, Choi C, Lee H, Ahn I, Park SR, Bae SC, Lee SC, Hwang DJ (2013) Expression analysis of rice VQ genes in response to biotic and abiotic stresses. Gene 529: 208–214 [DOI] [PubMed] [Google Scholar]
  17. Knight H. (2000) Calcium signaling during abiotic stress in plants. Int Rev Cytol 195: 269–324 [DOI] [PubMed] [Google Scholar]
  18. Lai Z, Li Y, Wang F, Cheng Y, Fan B, Yu JQ, Chen Z (2011) Arabidopsis sigma factor binding proteins are activators of the WRKY33 transcription factor in plant defense. Plant Cell 23: 3824–3841 [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Li Y, Jing Y, Li J, Xu G, Lin R (2014) Arabidopsis VQ MOTIF-CONTAINING PROTEIN29 represses seedling deetiolation by interacting with PHYTOCHROME-INTERACTING FACTOR1. Plant Physiol 164: 2068–2080 [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Luo M, Dennis ES, Berger F, Peacock WJ, Chaudhury A (2005) MINISEED3 (MINI3), a WRKY family gene, and HAIKU2 (IKU2), a leucine-rich repeat (LRR) kinase gene, are regulators of seed size in Arabidopsis. Proc Natl Acad Sci USA 102: 17531–17536 [DOI] [PMC free article] [PubMed] [Google Scholar]
  21. Marchler-Bauer A, Derbyshire MK, Gonzales NR, Lu S, Chitsaz F, Geer LY, Geer RC, He J, Gwadz M, Hurwitz DI, et al. (2015) CDD: NCBI’s conserved domain database. Nucleic Acids Res 43: D222–D226 [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Morikawa K, Shiina T, Murakami S, Toyoshima Y (2002) Novel nuclear-encoded proteins interacting with a plastid sigma factor, Sig1, in Arabidopsis thaliana. FEBS Lett 514: 300–304 [DOI] [PubMed] [Google Scholar]
  23. Narusaka M, Kawai K, Izawa N, Seki M, Shinozaki K, Seo S, Kobayashi M, Shiraishi T, Narusaka Y (2008) Gene coding for SigA-binding protein from Arabidopsis appears to be transcriptionally up-regulated by salicylic acid and NPR1-dependent mechanisms. J Gen Plant Pathol 74: 345–354 [Google Scholar]
  24. Pandey SP, Somssich IE (2009) The role of WRKY transcription factors in plant immunity. Plant Physiol 150: 1648–1655 [DOI] [PMC free article] [PubMed] [Google Scholar]
  25. Pecher P, Eschen-Lippold L, Herklotz S, Kuhle K, Naumann K, Bethke G, Uhrig J, Weyhe M, Scheel D, Lee J (2014) The Arabidopsis thaliana mitogen-activated protein kinases MPK3 and MPK6 target a subclass of ‘VQ-motif’-containing proteins to regulate immune responses. New Phytol 203: 592–606 [DOI] [PubMed] [Google Scholar]
  26. Perruc E, Charpenteau M, Ramirez BC, Jauneau A, Galaud JP, Ranjeva R, Ranty B (2004) A novel calmodulin-binding protein functions as a negative regulator of osmotic stress tolerance in Arabidopsis thaliana seedlings. Plant J 38: 410–420 [DOI] [PubMed] [Google Scholar]
  27. Petersen K, Qiu JL, Lütje J, Fiil BK, Hansen S, Mundy J, Petersen M (2010) Arabidopsis MKS1 is involved in basal immunity and requires an intact N-terminal domain for proper function. PLoS One 5: e14364. [DOI] [PMC free article] [PubMed] [Google Scholar]
  28. Petersen M, Brodersen P, Naested H, Andreasson E, Lindhart U, Johansen B, Nielsen HB, Lacy M, Austin MJ, Parker JE, et al. (2000) Arabidopsis MAP kinase 4 negatively regulates systemic acquired resistance. Cell 103: 1111–1120 [DOI] [PubMed] [Google Scholar]
  29. Qiu JL, Fiil BK, Petersen K, Nielsen HB, Botanga CJ, Thorgrimsen S, Palma K, Suarez-Rodriguez MC, Sandbech-Clausen S, Lichota J, et al. (2008) Arabidopsis MAP kinase 4 regulates gene expression through transcription factor release in the nucleus. EMBO J 27: 2214–2221 [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Qu LJ, Zhu YX (2006) Transcription factor families in Arabidopsis: major progress and outstanding issues for future research. Curr Opin Plant Biol 9: 544–549 [DOI] [PubMed] [Google Scholar]
  31. Rushton PJ, Somssich IE, Ringler P, Shen QJ (2010) WRKY transcription factors. Trends Plant Sci 15: 247–258 [DOI] [PubMed] [Google Scholar]
  32. Shiu SH, Shih MC, Li WH (2005) Transcription factor families have much higher expansion rates in plants than in animals. Plant Physiol 139: 18–26 [DOI] [PMC free article] [PubMed] [Google Scholar]
  33. Wang A, Garcia D, Zhang H, Feng K, Chaudhury A, Berger F, Peacock WJ, Dennis ES, Luo M (2010) The VQ motif protein IKU1 regulates endosperm growth and seed size in Arabidopsis. Plant J 63: 670–679 [DOI] [PubMed] [Google Scholar]
  34. Weyhe M, Eschen-Lippold L, Pecher P, Scheel D, Lee J (2014) Ménage à trois: the complex relationships between mitogen-activated protein kinases, WRKY transcription factors and VQ-motif-containing proteins. Plant Signal Behav 9: e29519. [DOI] [PMC free article] [PubMed] [Google Scholar]
  35. Wray GA, Hahn MW, Abouheif E, Balhoff JP, Pizer M, Rockman MV, Romano LA (2003) The evolution of transcriptional regulation in eukaryotes. Mol Biol Evol 20: 1377–1419 [DOI] [PubMed] [Google Scholar]
  36. Xie YD, Li W, Guo D, Dong J, Zhang Q, Fu Y, Ren D, Peng M, Xia Y (2010) The Arabidopsis gene SIGMA FACTOR-BINDING PROTEIN 1 plays a role in the salicylate- and jasmonate-mediated defence responses. Plant Cell Environ 33: 828–839 [DOI] [PMC free article] [PubMed] [Google Scholar]
  37. Yamaguchi-Shinozaki K, Shinozaki K (2006) Transcriptional regulatory networks in cellular responses and tolerance to dehydration and cold stresses. Annu Rev Plant Biol 57: 781–803 [DOI] [PubMed] [Google Scholar]
  38. Yamasaki K, Kigawa T, Seki M, Shinozaki K, Yokoyama S (2013) DNA-binding domains of plant-specific transcription factors: structure, function, and evolution. Trends Plant Sci 18: 267–276 [DOI] [PubMed] [Google Scholar]

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