Isolation of carrot basic leucine zipper transcription factor using yeast one-hybrid screening

Abscisic acid (ABA) is involved in various physiological and developmental processes, including stress responses and seed maturation. Many ABA-regulated genes associated with these processes have been identified and analyzed. Previously, we identified 2 important elements in the promoter of the carrotDcECP31 gene: motif X (CACACGTGGG), which is like an ABA-responsive element (ABRE), and motif Y (CACACGTATC). Together, these are sufficient for embryo-specific ABA-inducible promoter activity. We also showed that motif X functions is an enhancerlike element and that motif Y participates in ABA responsiveness. In this study, we isolated the nuclear protein that interacts with motif Y of theDcECP31 promoter. We performed yeast one-hybrid screening using integrated motif Y as bait and isolated clones. Sequence analysis revealed that clone 22 included the carboxyl-terminal half of bZIP, which contains the basic and leucine zipper domains and binds to G-boxes containing the sequence ACGT. This result supports the hypothesis that carrot C-ABI3, a homologue of theArabidopsis ABI3 protein, functions as a coactivator that interacts with the G-box via protein-protein contacts and suggests that the complex controls the expression of theDcECP31 gene.     

Developmental role and auxin responsiveness of Class III homeodomain leucine zipper gene family members in rice

Members of the Class III homeodomain leucine zipper (Class III HD-Zip) gene family are central regulators of crucial aspects of plant development. To better understand the roles of five Class III HD-Zip genes in rice (Oryza sativa) development, we investigated their expression patterns, ectopic expression phenotypes, and auxin responsiveness. Four genes, OSHB1 to OSHB4, were expressed in a localized domain of the shoot apical meristem (SAM), the adaxial cells of leaf primordia, the leaf margins, and the xylem tissue of vascular bundles. In contrast, expression of OSHB5 was observed only in phloem tissue. Plants ectopically expressing microRNA166-resistant versions of the OSHB3 gene exhibited severe defects, including the ectopic production of leaf margins, shoots, and radialized leaves. The treatment of seedlings with auxin quickly induced ectopic OSHB3 expression in the entire region of the SAM, but not in other tissues. Furthermore, this ectopic expression of OSHB3 was correlated with leaf initiation defects. Our findings suggest that rice Class III HD-Zip genes have conserved functions with their homologs in Arabidopsis (Arabidopsis thaliana), but have also acquired specific developmental roles in grasses or monocots. In addition, some Class III HD-Zip genes may regulate the leaf initiation process in the SAM in an auxin-dependent manner.     

Phylogenetic and expression analysis of the basic helix-loop-helix transcription factor gene family: genomic approach to cellular differentiation

A phylogenetic analysis of seven different species (human, mouse, rat, worm, fly, yeast, and plant) utilizing all (541) basic helix-loop-helix (bHLH) genes identified, including expressed sequence tags (EST), was performed. A super-tree involving six clades and a structural categorization involving the entire coding sequence was established. A nomenclature was developed based on clade distribution to discuss the functional and ancestral relationships of all the genes. The position/location of specific genes on the phylogenetic tree in relation to known bHLH factors allows for predictions of the potential functions of uncharacterized bHLH factors, including EST's. A genomic analysis using microarrays for four different mouse cell types (i.e. Sertoli, Schwann, thymic, and muscle) was performed and considered all known bHLH family members on the microarray for comparison. Cell-specific groups of bHLH genes helped clarify those bHLH genes potentially involved in cell specific differentiation. This phylogenetic and genomic analysis of the bHLH gene family has revealed unique aspects of the evolution and functional relationships of the different genes in the bHLH gene family.