Expression analysis of RING zinc finger genes from Triticum aestivum and identification of TaRZF70 that contains four RING-H2 domains and differentially responds to water deficit between leaf and root
ABSTRACT RING zinc finger proteins are known for their role predominantly in targeted protein degradation and participate in gene regulation through interaction with other regulatory proteins. In this study seven RING zinc finger genes from Triticum aestivum (bread wheat) were analysed for expression profiles in various organs (leaf, root, stem, spike, endosperm and embryo) and during leaf development and aging as well as in their responses to water deficit. Expression levels of six of these seven genes varied markedly among the six organs examined. All seven genes changed their expression levels in the leaf from the growing to senescing stage. Four genes were responsive to water deficit. A RING-H2 zinc finger gene, TaRZF70 showed differential response to water deprivation, namely up-regulation in the leaf and down-regulation in the root. This differential response was also observed in abscisic acid (ABA)-treated plants. Sequence analysis revealed that TaRZF70 contained four RING-H2 domains, the largest number of RING-H2 domains in any RING-H2 zinc finger proteins reported to date. These results indicate that these RING zinc finger genes are involved in diverse physiological processes in wheat, including response to drought.
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ABSTRACT: Intense interest in the RING domain has arisen because of its widespread occurrence and involvement in human disease. Several intriguing characteristics evident from the study of this cysteine-rich, zinc-binding domain have made it difficult to establish a single defining biochemical function for RINGs. These proteins are found throughout the cell and mediate diverse cellular processes, e.g. oncogenesis, apoptosis, development and viral infection. Recent developments indicate that RING-mediated protein interactions are critical for transcriptional repression and for ubiquitination. These data are in addition to previously established functions for RINGs in RNA processing, cell-cycle control and peroxisomal biogenesis, to name a few. At first glance, there appears to be little to link such disparate actions. Collectively, these results suggest that RINGs function in formation and architecture of large protein complexes that contribute to diverse cellular processes. Here, new developments, in the context of previous results, are discussed in an attempt to establish a unifying theory for RING function.Journal of Molecular Biology 03/2000; 295(5):1103-12. · 3.91 Impact Factor
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ABSTRACT: The KRAB domain is a 75 amino acid residue transcriptional repression module commonly found in eukaryotic zinc-finger proteins. KRAB-mediated gene silencing requires binding to the corepressor KAP-1. The KRAB:KAP-1 interaction requires the RING-B box-coiled coil (RBCC) domain of KAP-1, which is a widely distributed motif, hypothesized to be a protein-protein interface. Little is known about RBCC-mediated ligand binding and the role of the individual sub-domains in recognition and specificity. We have addressed these issues by reconstituting and characterizing the KRAB:KAP-1-RBCC interaction using purified components. Our results show that KRAB binding to KAP-1 is direct and specific, as the related RBCC domains from TIF1alpha and MID1 do not bind the KRAB domain. A combination of gel filtration, analytical ultracentrifugation, chemical cross-linking, non-denaturing gel electrophoresis, and site-directed mutagenesis techniques has revealed that the KAP-1-RBCC must oligomerize likely as a homo-trimer in order to bind the KRAB domain. The RING finger, B2 box, and coiled-coil region are required for oligomerization of KAP-1-RBCC and KRAB binding, as mutations in these domains concomitantly abolished these functions. KRAB domain binding stabilized the homo-oligomeric state of the KAP-1-RBCC as detected by chemical cross-linking and velocity sedimentation studies. Mutant KAP-1-RBCC molecules hetero-oligomerize with the wild-type KAP-1, but these complexes were inactive for KRAB binding, suggesting a potential dominant negative activity. Substitution of the coiled-coil region with heterologous dimerization, trimerization, or tetramerization domains failed to recapitulate KRAB domain binding. Chimeric KAP-1-RBCC proteins containing either the RING, RING-B box, or coiled-coil regions from MID1 also failed to bind the KRAB domain. The KAP-1-RBCC mediates a highly specific, direct interaction with the KRAB domain, and it appears to function as an integrated, possibly cooperative structural unit wherein each sub-domain contributes to oligomerization and/or ligand recognition. These observations provide the first principles for RBCC domain-mediated protein-protein interaction and have implications for identifying new ligands for RBCC domain proteins.Journal of Molecular Biology 03/2000; 295(5):1139-62. · 3.91 Impact Factor
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ABSTRACT: We describe the identification and structural characterization of a novel family of Arabidopsis genes related to ATL2 which encode a variant of the RING zinc finger domain, known as RING-H2. Analysis of genes selected by us and of sequences from Arabidopsis stored in databases permitted the prediction of several RING-H2 proteins that contain highly homologous RING domains. The ATL gene family is represented by fifteen sequences that contain, in addition to the RING, a transmembrane domain which is located in most of them towards the N-terminal end. Transgenic Arabidopsis seedlings carrying the ATL2 promoter fused to the GUS reporter gene revealed that the expression of ATL2 is rapidly induced after exposure to chitin or inactivated crude cellulase preparations. Rapid induction of transcript accumulation of another member of the ATL family was also observed under the same conditions. These results suggest that some ATLs may be involved in the early stages of the defense response triggered in plants in response to pathogen attack.Plant Molecular Biology 08/1999; 40(4):579-90. · 3.52 Impact Factor