Arabidopsis ZINC FINGER PROTEIN1 Acts Downstream of GL2 to Repress Root Hair Initiation and Elongation by Directly Suppressing bHLH Genes

Abstract

Cys2His2-like fold group (C2H2)-type zinc finger proteins promote root hair growth and development by regulating their target genes. However, little is known about their potential negative roles in root hair initiation and elongation. Here, we show that the C2H2-type zinc finger protein named ZINC FINGER PROTEIN1 (AtZP1), which contains an ERF-associated amphiphilic repression (EAR) motif, negatively regulates Arabidopsis (Arabidopsis thaliana) root hair initiation and elongation. Our results demonstrate that AtZP1 is highly expressed in root hairs and that AtZP1 inhibits transcriptional activity during root hair development. Plants overexpressing AtZP1 lacked root hairs, while loss-of-function mutants had longer and more numerous root hairs than the wild type. Transcriptome analysis indicated that AtZP1 downregulates genes encoding basic helix-loop-helix (bHLH) transcription factors associated with root hair cell differentiation and elongation. Mutation or deletion of the EAR motif substantially reduced the inhibitory activity of AtZP1. Chromatin immunoprecipitation assays, AtZP1:glucocorticoid receptor (GR) induction experiments, electrophoretic mobility shift assays, and yeast one-hybrid assays showed that AtZP1 directly targets the promoters of bHLH transcription factor genes, including the key root hair initiation gene ROOT HAIR DEFECTIVE6 (RHD6) and root hair elongation genes ROOT HAIR DEFECTIVE 6-LIKE 2 (RSL2) and RSL4, and suppresses root hair development. Our findings suggest that AtZP1 functions downstream of GL2 and negatively regulates root hair initiation and elongation, by suppressing RHD6, RSL4, and RSL2 transcription via the GL2/ZP1/ RSL pathway.

C2H2 Zinc Finger Proteins: Master Regulators of Abiotic Stress Responses in Plants

“…The conserved EAR motif in class II ERF TFs plays a key role in their inhibitory activity. For example, the C-terminal inhibitory domain of AtERF4 is DLDLNL; if a mutation occurs in this domain, the protein's inhibitory function is abolished (Ohta et al, 2001;Han et al, 2019a).…”

“…Several abioticstress-related C2H2-type zinc finger proteins, such as AZF1, AZF2, AZF3, and ZAT10/STZ, specifically bind to the A(G/C)T repeat sequences in their target promoters (Takatsuji et al, 1994;Sakamoto et al, 2004;Kodaira et al, 2011). The DNA sequence A [AG/CT]CNAC (Sakai et al, 1995;Kubo et al, 1998;Zhou et al, 2011;Han et al, 2019a), the TGCTANNATTG element (Huang et al, 2009b), and TACAAT motifs (Shi et al, 2014) are also possible binding domains of C2H2 TFs in plants. Under abiotic stress conditions, C2H2 zinc finger proteins directly target downstream stress-related genes to activate or inhibit their expression.…”

“…These regulatory proteins include protein kinases [including mitogen activated protein kinases (MAPK), calcium dependent protein kinases (CDPK), receptor protein kinases, ribosomal protein kinases, and transcriptional regulatory protein kinases] (Tibbles and Woodgett, 1999;Wimalasekera and Scherer, 2018;Divya et al, 2019), protein phosphatases (Singh et al, 2010;Gong et al, 2018), transcription factors (TFs) (Han et al, 2014;Sun et al, 2018), and proteins involved in inorganic phosphate (Pi) turnover (Takahashi et al, 2001;Fabiańska et al, 2019). TFs bind to specific sequences in the promoters of their target genes, thereby regulating gene expression and affecting biological phenotypes (Han et al, 2019a). TFs are key regulatory components of abiotic stress signaling pathways (Schmidt et al, 2012;Castelán-Muñoz et al, 2019).…”

Genome-wide characterization of the C2H2 zinc-finger genes in Cucumis sativus and functional analyses of four CsZFPs in response to stresses

“…For example, JcZFP8 , a C2H2 zinc finger protein gene from Jatropha curcas L., participates the trichome development in transgenic tobacco [ 47 ]. The C2H2-typezinc finger protein named ZINC FINGER PROTEIN 1 ( AtZP1 ) negatively regulates Arabidopsis thaliana root hair initiation and elongation [ 48 ].…”

Autocrine regulation of root hair size by the RALF‐FERONIA‐RSL4 signaling pathway

“…1). Recently, more genetic components that regulate RH growth were reported, including ZINC FINGER PROTEIN1 (AtZP1), which negatively regulates RHD6 , RSL4 , and RSL2 expression and thereby represses RH growth (Han et al , 2020). In a similar manner, GT‐2‐LIKE1 (GTL1) and its homolog DF1 bind to the RSL4 promoter and repress RH growth (Shibata et al , 2018).…”