The Arabidopsis tandem zinc finger protein AtTZF1 affects ABA- and GA-mediated growth, stress and gene expression responses

Tandem zinc finger (TZF) proteins are characterized by two zinc-binding CCCH motifs arranged in tandem. Human TZFs such as tristetraproline (TTP) bind to and trigger the degradation of mRNAs encoding cytokines and various regulators. Although the molecular functions of plant TZFs are unknown, recent genetic studies have revealed roles in hormone-mediated growth and environmental responses, as well as in the regulation of gene expression. Here we show that expression of AtTZF1 (AtCTH/AtC3H23) mRNA is repressed by a hexokinase-dependent sugar signaling pathway. However, AtTZF1 acts as a positive regulator of ABA/sugar responses and a negative regulator of GA responses, at least in part by modulating gene expression. RNAi of AtTZF1-3 caused early germination and slightly stress-sensitive phenotypes, whereas plants over-expressing AtTZF1 were compact, late flowering and stress-tolerant. The developmental phenotypes of plants over-expressing AtTZF1 were only partially rescued by exogenous application of GA, implying a reduction in the GA response or defects in other mechanisms. Likewise, the enhanced cold and drought tolerance of plants over-expressing AtTZF1 were not associated with increased ABA accumulation, suggesting that it is mainly ABA responses that are affected. Consistent with this notion, microarray analysis showed that over-expression of AtTZF1 mimics the effects of ABA or GA deficiency on gene expression. Notably, a gene network centered on a GA-inducible and ABA/sugar-repressible putative peptide hormone encoded by GASA6 was severely repressed by AtTZF1 over-expression. Hence AtTZF1 may serve as a regulator connecting sugar, ABA, GA and peptide hormone responses.     

The Arabidopsis thaliana tandem zinc finger 1 (AtTZF1) protein in RNA binding and decay

The Arabidopsis thaliana tandem zinc finger 1 (AtTZF1) protein is characterized by two tandem‐arrayed CCCH‐type zinc fingers. We have previously found that AtTZF1 affects hormone‐mediated growth, stress and gene expression responses. While much has been learned at the genetic and physiological level, the molecular mechanisms underlying the effects of AtTZF1 on gene expression remain obscure. A human TZF protein, hTTP, is known to bind and trigger the degradation of mRNAs containing AU‐rich elements (AREs) at the 3′ untranslated regions. However, while the TZF motif of hTTP is characterized by C_X8C_X5C_X3H‐_X18‐C_X8C_X5C_X3H, AtTZF1 contains an atypical motif of C_X7C_X5C_X3H‐_X16‐C_X5C_X4C_X3H. Moreover, the TZF motif of AtTZF1 is preceded by an arginine‐rich (RR) region that is unique to plants. Using fluorescence anisotropy and electrophoretic mobility shift binding assays, we have demonstrated that AtTZF1 binds to RNA molecules with specificity and the interaction is dependent on the presence of zinc. Compared with hTTP, in which TZF is solely responsible for RNA binding, both TZF and RR regions of AtTZF1 are required to achieve high‐affinity RNA binding. Moreover, zinc finger integrity is vital for RNA binding. Using a plant protoplast transient expression analysis we have further revealed that AtTZF1 can trigger the decay of ARE‐containing mRNAs in vivo. Taken together, our results support the notion that AtTZF1 is involved in RNA turnover.     

A CCCH-type zinc finger nucleic acid-binding protein quantitatively confers resistance against rice bacterial blight disease

Bacterial blight is a devastating disease of rice (Oryza sativa) caused by Xanthomonas oryzae pv oryzae (Xoo). Zinc finger proteins harboring the motif with three conserved cysteine residues and one histidine residue (CCCH) belong to a large family. Although at least 67 CCCH-type zinc finger protein genes have been identified in the rice genome, their functions are poorly understood. Here, we report that one of the rice CCCH-type zinc finger proteins, C3H12, containing five typical CX(8)-CX(5)-CX(3)-H zinc finger motifs, is involved in the rice-Xoo interaction. Activation of C3H12 partially enhanced resistance to Xoo, accompanied by the accumulation of jasmonic acid (JA) and induced expression of JA signaling genes in rice. In contrast, knockout or suppression of C3H12 resulted in partially increased susceptibility to Xoo, accompanied by decreased levels of JA and expression of JA signaling genes in rice. C3H12 colocalized with a minor disease resistance quantitative trait locus to Xoo, and the enhanced resistance of randomly chosen plants in the quantitative trait locus mapping population correlated with an increased expression level of C3H12. The C3H12 protein localized in the nucleus and possessed nucleic acid-binding activity in vitro. These results suggest that C3H12, as a nucleic acid-binding protein, positively and quantitatively regulates rice resistance to Xoo and that its function is likely associated with the JA-dependent pathway.     

EARLY RESPONSIVE TO DEHYDRATION 15, a negative regulator of abscisic acid responses in Arabidopsis

EARLY RESPONSIVE TO DEHYDRATION 15 (ERD15) is rapidly induced in response to various abiotic and biotic stress stimuli in Arabidopsis (Arabidopsis thaliana). Modulation of ERD15 levels by overexpression or RNAi silencing altered the responsiveness of the transgenic plants to the phytohormone abscisic acid (ABA). Overexpression of ERD15 reduced the ABA sensitivity of Arabidopsis manifested in decreased drought tolerance and in impaired ability of the plants to increase their freezing tolerance in response to this hormone. In contrast, RNAi silencing of ERD15 resulted in plants that were hypersensitive to ABA and showed improved tolerance to both drought and freezing, as well as impaired seed germination in the presence of ABA. The modulation of ERD15 levels not only affected abiotic stress tolerance but also disease resistance: ERD15 overexpression plants showed improved resistance to the bacterial necrotroph Erwinia carotovora subsp. carotovora accompanied with enhanced induction of marker genes for systemic acquired resistance. We propose that ERD15 is a novel mediator of stress-related ABA signaling in Arabidopsis.     

Gene-specific involvement of beta-oxidation in wound-activated responses in Arabidopsis

The coordinated induced expression of beta-oxidation genes is essential to provide the energy supply for germination and postgerminative development. However, very little is known about other functions of beta-oxidation in nonreserve organs. We have identified a gene-specific pattern of induced beta-oxidation gene expression in wounded leaves of Arabidopsis. Mechanical damage triggered the local and systemic induction of only ACX1 among acyl-coenzyme A oxidase (ACX) genes, and KAT2/PED1 among 3-ketoacyl-coenzyme A thiolase (KAT) genes in Arabidopsis. In turn, wounding induced KAT5/PKT2 only systemically. Although most of the beta-oxidation genes were activated by wound-related factors such as dehydration and abscisic acid, jasmonic acid (JA) induced only ACX1 and KAT5. Reduced expression of ACX1 or KAT2 genes, in transgenic plants expressing their corresponding mRNAs in antisense orientation, correlated with defective wound-activated synthesis of JA and with reduced expression of JA-responsive genes. Induced expression of JA-responsive genes by exogenous application of JA was unaffected in those transgenic plants, suggesting that ACX1 and KAT2 play a major role in driving wound-activated responses by participating in the biosynthesis of JA in wounded Arabidopsis plants.     

Comprehensive analysis of CCCH-type zinc finger gene family in citrus (Clementine mandarin) by genome-wide characterization

The CCCH-type zinc finger proteins comprise a large gene family of regulatory proteins and are widely distributed in eukaryotic organisms. The CCCH proteins have been implicated in multiple biological processes and environmental responses in plants. Little information is available, however, about CCCH genes in plants, especially in woody plants such as citrus. The release of the whole-genome sequence of citrus allowed us to perform a genome-wide analysis of CCCH genes and to compare the identified proteins with their orthologs in model plants. In this study, 62 CCCH genes and a total of 132 CCCH motifs were identified, and a comprehensive analysis including the chromosomal locations, phylogenetic relationships, functional annotations, gene structures and conserved motifs was performed. Distribution mapping revealed that 54 of the 62 CCCH genes are unevenly dispersed on the nine citrus chromosomes. Based on phylogenetic analysis and gene structural features, we constructed 5 subfamilies of 62 CCCH members and integrative subfamilies from citrus, Arabidopsis, and rice, respectively. Importantly, large numbers of SNPs and InDels in 26 CCCH genes were identified from Poncirus trifoliata and Fortunella japonica using whole-genome deep re-sequencing. Furthermore, citrus CCCH genes showed distinct temporal and spatial expression patterns in different developmental processes and in response to various stress conditions. Our comprehensive analysis of CleC3Hs is a valuable resource that further elucidates the roles of CCCH family members in plant growth and development. In addition, variants and comparative genomics analyses deepen our understanding of the evolution of the CCCH gene family and will contribute to further genetics and genomics studies of citrus and other plant species.