Histone methylation readers MRG1/MRG2 interact with the transcription factor TCP14 to positively modulate cytokinin sensitivity in Arabidopsis

Cytokinins were initially identified as a class of plant hormones that stimulate cell division in cultured plant cells (Werner and Schmulling, 2009; Kieber and Schaller, 2014). Since then, an increasing number of studies proved that cytokinins play significant roles in diverse processes related to plant growth and development, including shoot and root development, vascular differentiation, and leaf senescence. Additionally, cytokinins have also been revealed to regulate plant responses to both biotic and abiotic stresses (Werner and Schmulling, 2009; Nishiyama et al., 2011; Kieber and Schaller, 2014; Schaller et al., 2014).

The cytokinin signaling pathway in Arabidopsis is composed of three key components: cytokinin receptors (Arabidopsis histidine kinases; AHKs), phosphotransfer proteins (Arabidopsis histidine phosphotransfer proteins; AHPs), and response regulators (Arabidopsis response regulators; ARRs) (Hwang et al., 2002; Nishimura et al., 2004; To et al., 2004; Hutchison et al., 2006; Kieber and Schaller, 2018). The activation of genes transcription downstream of cytokinin signal perception is mediated by multiple histidine-aspartate phosphorelay reactions (Hwang et al., 2002). The AHKs, including AHK2, AHK3, and AHK4, redundantly perceive cytokinin signals and undergo autophosphorylation (Nishimura et al., 2004), after which the phosphoryl groups are transferred to cytosolic AHPs. Arabidopsis contains six AHP proteins. More specifically, AHP1-AHP5 are conserved proteins with histidine phosphotransfer activities, whereas AHP6 is considered to be a pseudo-phosphotransfer protein lacking the conserved histidine phosphorylation site (Hutchison et al., 2006; Mahonen et al., 2006). The ahp2 ahp3 ahp5 triple mutant exhibits decreased cytokinin sensitivity in root and hypocotyl inhibition assays, and AHP2-overexpressing plants are hypersensitive to cytokinin, suggesting that AHP2, AHP3, and AHP5 function as key redundant positive regulators in the cytokinin signal transduction pathway (Hutchison et al., 2006; Jeon and Kim, 2013; Liu et al., 2017). Phosphorylated AHPs are translocated into the nucleus, where the phosphoryl groups are transferred to type-B ARRs. Activated type-B ARRs control the expression of target genes, including type-A ARR genes, which in turn suppress the cytokinin response (To et al., 2004; Mason et al., 2005; Zubo et al., 2017; Kieber and Schaller, 2018; Xie et al., 2018). Type-A ARRs can also be phosphorylated by AHPs and act as negative regulators in cytokinin signaling (To et al., 2007).

Along with the information obtained from in-depth investigations of the cytokinin signaling pathway, studies have revealed that modifications of histone tails and other epigenetic elements are involved in regulating the cytokinin transcriptional network (Werner and Schmulling, 2009; Li et al., 2013; Potter et al., 2018). Moreover, histone H3 lysine 4 methylation and lysine 27 methylation affect cytokinin-mediated plant growth (Chen et al., 2013; Li et al., 2013). In this study, we revealed that the MORF-RELATED GENE (MRG) family proteins MRG1/2, which are readers of trimethylated histone H3 at lysine 4 and lysine 36 (H3K4me3/H3K36me3), contribute to the regulation of cytokinin signaling. The MRGs containing the chromo-domain in the N-terminal region are conserved in diverse eukaryotes, including yeast, humans, and plants (Bertram and Pereira-Smith, 2001; Liu et al., 2016). Arabidopsis MRG1/2 and rice MRG702 control the flowering time or hypocotyl elongation in response to light by recognizing H3K4me3/H3K36me3 and further modulating the transcription of related genes (Bu et al., 2014; Jin et al., 2015; Peng et al., 2018). Furthermore, MRGs interact with the histone H4-specific acetyltransferases HISTONE ACETYLTRANSFERASE OF THE MYST FAMILY 1 (HAM1) and HAM2 or histone deacetylase HISTONE DEACETYLASE 2C (HD2C) to enhance the histone-4 lysine-5 acetylation (H4K5ac) or to catalyze histone deacetylation (Xu et al., 2014; Guo et al., 2020). Because of the genome-wide distribution of H3K4me3 and H3K36me3 and the potential roles of histone acetylation associated with gene transcriptional regulation, further analyses of the important roles of MRGs in various developmental processes are warranted.

Recent studies proved that MRG1/2 function cooperatively with histone modifiers as well as various transcription factors, such as CONSTANS (CO) and PHYTOCHROME-INTERACTING FACTOR7 (PIF7), to precisely facilitate the expression of downstream genes and regulate biological functions (Bu et al., 2014; Peng et al., 2018). Previous research indicated that several TEOSINTE BRANCHED, CYCLOIDEA, AND PCF (TCP) proteins, which belong to a group of plant-specific transcription factors with a conserved non-canonical basic helix-loop-helix (bHLH) motif known as the TCP domain, also help regulate cytokinin-related developmental processes (Werner and Schmulling, 2009; Martin-Trillo and Cubas, 2010; Steiner et al., 2012a; Steiner et al., 2016). For example, TCP4 in Arabidopsis can diminish leaf responses to cytokinin by inducing the expression of ARR16, which encodes an inhibitor in cytokinin responses (Efroni et al., 2013). In contrast, TCP14 and TCP15 can enhance leaf responses to cytokinin. The Arabidopsis and tomato tcp14 tcp15 mutants are reportedly hyposensitive to exogenously applied cytokinin, and the overexpression of TCP14 or TCP15 increases cytokinin sensitivity (Steiner et al., 2012b; Lucero et al., 2015; Steiner et al., 2016). As described above, TCP proteins are proved to act as transcription factors to regulate cytokinin responses. Recently, it has been reported that the N-terminal of CYCLOIDEA and TEOSINTE BRANCHED 1-like TCP proteins from cucumber (Cucumis sativus L) contains an intrinsically disordered region expected to have conserved histone acetyltransferase (HAT) activity (Yang et al., 2020) and Marchantia TCP protein could densely bind topologically associated domains of genome (Karaaslan et al., 2020), suggesting that TCPs function not only as transcription factors that control the expression of target genes but may also serve as chromatin-remodeling factors that modulate chromatin states. Therefore, it is particularly important to study the relationship between TCPs and other epigenetic regulatory factors (e.g., MRG1/2) in terms of their effects on the cytokinin response and plant development.

In this study, we discovered that MRG1/2 and AtTCP14 positively regulate the cytokinin signaling pathway. Both mrg1 mrg2 and tcp14-2 mutants had pleiotropic cytokinin-hyposensitive phenotypes during the synthetic cytokinin N6-benzylaminopurine (6-BA)-induced inhibition of root elongation and seedling growth as well as the induction of callus growth. We demonstrated that MRG2 can physically interact with the transcription factor AtTCP14 in vitro and in vivo, suggesting they might function as a complex. The complex was recruited to the AHP2 genomic region to enhance the H4K5ac epigenetic modification, which subsequently promoted the transcription of AHP2 and its downstream signaling. The data presented herein lay the foundation for future mechanistic studies on the epigenetic control of plant hormone crosstalk.

留言 (0)

沒有登入
gif