GeBP and GeBP-Like Proteins Are Noncanonical Leucine-Zipper Transcription Factors That Regulate Cytokinin Response in Arabidopsis  

Abstract

Understanding the role of transcription factors (TFs) is essential in reconstructing developmental regulatory networks. The plant-specific GeBP TF family of Arabidopsis thaliana (Arabidopsis) comprises 21 members, all of unknown function. A subset of four members, the founding member GeBP and GeBP-like proteins (GPL) 1, 2, and 3, shares a conserved C-terminal domain. Here we report that GeBP/GPL genes represent a newly defined class of leucine-zipper (Leu-zipper) TFs and that they play a redundant role in cytokinin hormone pathway regulation. Specifically, we demonstrate using yeast, in vitro, and split-yellow fluorescent protein in planta assays that GeBP/GPL proteins form homo-and heterodimers through a noncanonical Leu-zipper motif located in the C-terminal domain. A triple loss-of-function mutant of the three most closely related genes gebp gpl1 gpl2 shows a reduced sensitivity to exogenous cytokinins in a subset of cytokinin responses such as senescence and growth, whereas root inhibition is not affected. We find that transcript levels of type-A cytokinin response genes, which are involved in the negative feedback regulation of cytokinin signaling, are higher in the triple mutant. Using a GPL version that acts as a constitutive transcriptional activator, we show that the regulation of Arabidopsis response regulators (ARRs) is mediated by at least one additional, as yet unknown, repressor acting genetically downstream in the GeBP/GPL pathway. Our results indicate that GeBP/GPL genes encode a new class of unconventional Leu-zipper TF proteins and suggest that their role in the cytokinin pathway is to antagonize the negative feedback regulation on ARR genes to trigger the cytokinin response.

STOREKEEPER RELATED1/G-Element Binding Protein (STKR1) Interacts with Protein Kinase SnRK1

“…2A). This indicates that the STKR1 N terminus is required for its interaction with the SnRK1 a-subunits.Previous studies have shown that STKR1/GeBP and its close homologs GPL1, GPL2, and GPL3 belong to a plant-specific family of transcription factors that form homodimers and heterodimers through their noncanonical Leu zipper motif located at the C terminus of the proteins (Chevalier et al, 2008). A direct interaction assay confirmed that STKR1 forms homodimers and heteromerizes with the GPL proteins via its C-terminal part (Fig.…”

“…2B), indicating that dimerization and SnRK1 interaction are mediated by different domains of the protein. Because STKR1/GeBP and GPL proteins have been proposed to act redundantly (Chevalier et al, 2008), we tested the ability of GPL1 and GPL3 to interact with SnRK1 in yeast. The data revealed that these proteins did not bind to the SnRK1 a-subunits, suggesting a specific functional relationship of STKR1 with SnRK1 (Fig.…”

Increased vegetative development and sturdiness of storekeeper-transgenic tobacco

“…Therefore, we deduced that a low threshold value existed for the transgenic gene product to realize the heterochronic and morphological responses observed in 35S::stk01-and 35S::stk03-transgenic tobacco. We and others [7] observed that STK-like proteins were able form homo-and heterodimers and therefore it appeared likely that also interactions occurred between transgenic Arabidopsis and resident tobacco STK-like proteins. To date, ten STK-like sequences have been identified in tobacco and deposited within the TOBFAC database.…”

“…More important with respect to the data presented, the overexpression of GEBP in Arabidopsis did not result in a phenotype deviating from the wild type [6]. Recently, GEBP and three STK-like proteins (At2g25650, At2g36340 and At5g14270, also termed GEBP-like proteins: GPL1, 2 and 3) were shown to form homoand heterodimers via unconventional leucine zipper motifs and to participate in the regulation of cytokinin response in Arabidopsis thaliana [7]. We have started characterizing stk-like genes after screening Arabidopsis expression libraries in the Yeast OneHybrid system using as a bait 700 bp of the 5´-silencer region within intron 2 of the Arabidopsis floral homeotic gene agamous (ag; [8] [GEBP]) when the plant-specific GAGA-factor BPC3 (At1g68120) was co-expressed in yeast (J.F.…”

“…In the future we will expand target gene screenings of our STK proteins to identify further factors involved in the development of the sturdy S-plants. Putative targets may not only include genes identified by our Yeast Two-Hybrid screenings but also along the pathway of cytokinin signalling since a triple mutant in stk-like genes of Arabidopsis was affected in a subset of cytokinin responses [7].…”