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Abstract

Genes for histidyl-aspartyl (His-Asp) phosphorelay components (His-containing phosphotransfer proteins, HP, and response regulators, RR) were isolated from Zea mays L. to characterize their function in cytokinin signaling. Six type-A RRs (ZmRR1, ZmRR2, ZmRR4-ZmRR7), 3 type-B RRs (ZmRR8-ZmRR10), and 3 HPs (ZmHP1-ZmHP3) were found in leaves. All type-A RR genes expressed in leaves were up-regulated by exogenous cytokinin. Transient expression of fusion products of the signaling modules with green fluorescent protein in epidermal leaf cells suggested cytosolic and nuclear localizations of ZmHPs, whereas type-B ZmRR8 was restricted to the nucleus. Type-A RRs were localized partly to the cytosol (ZmRR1, ZmRR2, and ZmRR3) and partly to the nucleus (ZmRR4, ZmRR5, and ZmRR6). In the yeast two-hybrid assay, ZmHP1 and ZmHP3 interacted with both cytosolic ZmRR1 and nuclear type-B ZmRRs. In vitro experiments demonstrated that ZmHPs function as a phospho-donor for ZmRRs; turnover rates of the phosphorylated state were tenfold lower in ZmRR8 and ZmRR9 than in ZmRR1 and ZmRR4. These results suggest that the His-Asp phosphorelay signaling pathway might diverge into a cytosolic and a nuclear branch in leaves of maize, and that the biochemical nature of ZmRRs is different in terms of stability of the phosphorylated status.

Atypical response regulators expressed in the maize endosperm transfer cells link canonical two component systems and seed biology

“…Three HPs are currently known in maize [30], located on chromosomes 1, 2 and 4. A homology search did not show any additional loci in the maize genome.…”

“…There is however ample evidence to support their direct upstream interaction with histidine phosphotransfer proteins within the transduction route [30,35]. To study whether ZmTCRRs show coincidence with maize HPs in timing and location, polyclonal antibodies were raised against ZmTCRR-2, ZmHP1 and ZmHP2, the polyclonal anti-ZmTCRR-1 was already available [25].…”

Ligand-binding properties and subcellular localization of maize cytokinin receptors

“…Since the input domain of cytokinin receptors contains putative transmembrane regions, these receptors are thought to be integrated into membranes. However, while the subcellular location of the downstream modules (His-phosphotransfer proteins and response regulators) is established in A. thaliana and maize (Hwang and Sheen, 2001; Imamura et al , 2001; Asakura et al , 2003), the location of the primary receptors is unclear to date. Early concepts presuming that cytokinin receptors are located in the plasma membrane (Ueguchi et al , 2001; Inoue et al , 2001) were based on a bioinformatic approach.…”

Laser Microdissection of Narrow Sheath Mutant Maize Uncovers Novel Gene Expression in the Shoot Apical Meristem

“…ZmHP1 has been shown to function in vitro as a phospho-donor for the maize response regulators (ZmRRs) ZmRR1, ZmRR4, ZmRR8, and ZmRR9, whereas accumulation of ZmHP1 in both the cytoplasm and nucleus is consistent with its predicted function during two-component signaling [19]. In situ hybridization of Zmhp1 reveals transcript accumulation at two lateral foci within the nonmutant SAM (red arrows in Figure 5E and 5G), which mimics the ns expression domain (Figure 2C) [2].…”

“…A total of 11 functional categories are identified (Figure 3), including genes predicted to be involved in two-component signaling, auxin transport and signaling, jasmonate-induction/sugar signaling, intercellular transport, RNA processing, chromatin remodeling, transcription regulation, growth/cell division, ribosome structure, and general metabolism. Of the 66 genes identified in our microarray analyses, just one (Zmhp1) has been characterized biochemically [19], and none have been subjected to genetic analysis in maize. In addition, nine genes of unknown function are differentially expressed in ns mutant SAMs (Table 1).…”