In eukaryotic cells, the nuclear envelope governs the nuclear and cytoplasmic trafficking pathways and provides an important feature to control the specificity and spatio-temporal signaling events. Translocation proteins are generally recognized by the nuclear transport receptors (NTRs) based on the nuclear localization signals (NLSs) or nuclear export signals (NESs) on the cargo proteins (Krebs et al., 2010). Importin α, an adaptor of NTRs, consists of tandem armadillo repeats and an auto-inhibitory importin β-binding (IBB) domain, which can bind to the NLS of a cargo and importin β (IMβ), respectively (Cook et al., 2007; Kobe, 1999). Then, the ternary importin α/β−cargo complex passes through the nuclear pore complexes (NPCs) through transient interactions between importin β and Phe/Gly-repeat nucleoporins (Nups) of the NPCs that create a selective permeability barrier (Schmidt and Görlich, 2016). Conversely, chromosome maintenance protein1 (CRM1/XPO1, or exportin 1), a nuclear export receptor, binds to the cargo proteins with NESs and to RanGTP inside the nucleus, traverses the NPCs, and releases cargo into the cytoplasm (Haasen et al., 1999; Monecke et al., 2014; Ossareh-Nazari et al., 2001). Such dynamic changes of nucleocytoplasmic proteins have been implicated in the regulation of plant developmental and environmental responses (Yang et al., 2017).
Genetic screens for suppressors of the autoimmune mutant, snc1 (suppressor of npr1-1, constitutive1), have led to the identification of mos (modifiers of snc1) mutants (Zhang and Li, 2005). Among the MOS genes, MOS3 and MOS7 encode Nup96 and Nup88 of the NPCs, respectively. MOS7 is required for basal resistance, effector-triggered immunity (ETI), and resistance against biotrophic, hemibiotrophic, and necrotrophic pathogens (Cheng et al., 2009; Genenncher et al., 2016). MOS7 is verified to promote the nuclear accumulation of the immunoregulatory proteins such as EDS1 (enhanced disease susceptibility 1), NPR1 (nonexpresser of pathogenesis-related gene 1), and MPK3 (mitogen-activated protein kinase 3). It has been demonstrated that the balance of EDS1 in the cytosol and nucleus is required for efficient basal immune and toll/interleukin1 receptor domain-containing nucleotide-binding leucine-rich repeat (TNL) R protein-triggered resistance (García et al., 2010). Sufficient abundance of MPK3 protein in nucleus is important for full immunity to Botrytis cinerea in Arabidopsis (Genenncher et al., 2016). Moreover, CPR5 (constitutive expresser of PR gene 5) is a novel transmembrane nucleoporin, which associates with NPC core scaffold to allow massive nuclear influx of diverse stress-related signaling cargos after activation of immunoreceptors (Gu et al., 2016). The data clearly indicate an important role of nucleocytoplasmic trafficking, especially nuclear import protein, in plant innate immunity.
Nuclear cargo proteins usually consist of importin α-binding NLSs, which are monopartite or bipartite (separated by 10–12 residues at the linker region) sequence motifs enriched in basic amino acids of three to five residuals (Chang et al., 2012). In Arabidopsis, MOS6, which encodes importin α3 from the nine Arabidopsis importin α isoforms, is required for TNL R gene snc1-mediated resistance against a virulent oomycete pathogen (Palma et al., 2005) and demonstrated to be the main NTR of SNC1 (Lüdke et al., 2021), importing proteins involved in defense signaling in the nucleus. Moreover, an importin β-like protein that is sensitive to abscisic acid (ABA) and drought, SAD2, mediates nuclear import of transcription repressor MYB4 and is required for UV-B response in Arabidopsis (Zhao et al., 2007). As a consequence of the absence of MYB4 in the nucleus, the sad2 mutant shows constitutive expression of cinnamate 4-hydroxylase (C4H) gene, leading to accumulation of UV-absorbing pigments and an enhanced tolerance to UV-B radiation (Zhao et al., 2007). Similarly, nuclear translocation of Tartary buckwheat FtMYB16 is mediated by importin α1 to repress rutin biosynthesis (Li et al., 2019). Furthermore, plant pathogens may take advantage of the plant protein transport system to deliver virulent effector proteins into the host cytoplasm or periplasmic space to interfere with and manipulate host functions. Rice importin α1a and Arabidopsis importin αs interact with the bipartite NLSs of Agrobacterium tumefaciens virulence proteins VirD2 and VirE2, respectively (Bhattacharjee et al., 2008; Chang et al., 2014). Recently, rice importin α1a and importin α1b have shown to be necessary for nuclear import transcription activator-like effectors (TALEs), which are the secreted virulence proteins of Xanthomonas oryzae pv. oryzae (Xoo) and X. oryzae pv. oryzicola (Xoc), the causal agents of bacterial leaf blight and leaf streak, respectively (Hui et al., 2019). Nuclear localization of TALEs increases host susceptibility and modulates host gene expression (Hui et al., 2019; Szurek et al., 2001). The results suggest that nuclear transportation of proteins play a significant role in response to biotic and abiotic challenges.
OsWRKY62 and OsWRKY76, belonging IIa subgroup of WRKY transcription factors (TFs), function negatively in disease resistance against M. oryzae and Xoo (Liu et al., 2016; Peng et al., 2008; Wu et al., 2005; Yokotani et al., 2013). However, OsWRKY62 may act as a positive regulator of defense when it forms heterocomplex with a strong transactivator OsWRKY45 (Fukushima et al., 2016). Stable and transient expression of OsWRKY62.1-GFP shows cytosolic localization in unknown structures or as aggregates (Liu et al., 2016). Interestingly, OsWRKY62.1 is localized in the nuclei when it interacts with OsWRKY76.1 or XA21, a rice pattern recognition receptor for Xoo (Liu et al., 2016; Park and Ronald, 2012), implying a conditioned nuclear import of OsWRKY62.1 for its regulatory function. In a yeast two-hybrid (Y2H) cDNA library screening, we found that OsWRKY76.1 interacted with rice importin α1a (OsIMα1a). Further analysis indicated that OsWRKY62.1 also interacted with OsIMα1a and its close homolog OsIMα1b.