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Plant nonsymbiotic hemoglobins: Hexacoordinated hemoglobins involved in hormonally -regulated cell differentiation

Emily Jane Hume Ross, University of Nebraska - Lincoln


Hemoglobins (Hbs) are heme proteins that are widespread in nature. They are α-helical and characterized by the globin backbone. Functionally, Hbs bind small gaseous ligands and small cellular molecules. Plants contain three types of Hbs: truncated, symbiotic, and nonsymbiotic (ns). The nsHbs are the topic of this dissertation. ^ Recombinant (r) Hb2 from rice (Oryza sativa) was analyzed for its ligand-binding and hexacoordination kinetics. The observed data were used in structural modeling studies, using the known solution structure of rice rHb1. rHb2 and rHb1 exhibit similar gaseous ligand-binding kinetics, however they display marked differences in visible absorption spectra, binding to bulky ligands, and hexacoordination parameters. The structural features responsible for these differences are discussed using computer-based modeling. ^ Immunoblotting analysis indicates that the rice nsHb isoproteins are present during cereal seed germination and in all seedling tissues through the first two weeks following germination. Immunodetection using confocal laser scanning microcopy provides evidence that nsHbs are localized to differentiating cell types, such as root cap cells, aleurone, and xylem. ^ Mapping and sequence analysis of the hemoglobin (hb) gene family was analyzed in rice, which consists of four functional hb gene copies organized in two clusters: hb1/hb3/hb4 and hb2. Sequence comparison with databases revealed that a number of potential promoters exist upstream from the hb genes suggesting that these isogenes are differentially expressed. ^ In silico analysis of the putative rice NSHB1-4 and Arabidopsis NSHB1-2 promoters suggests the presence of hormonally-regulated cis-elements. Rice NSHB2 promoter was analyzed for tissue specific activation using the GUS reporter gene in transgenic Arabidopsis thaliana. Using transient assays, the rice NSHB2 promoter was analyzed for cytokinin activation through deletion and mutational studies, and by co-bombardment with the Arabidopsis ARR1 gene. The implications of these results, specifically hormonal regulation of NSHB gene expression are discussed. ^ Finally, the regulation of these genes, role(s) of these proteins, and potential functions of the nsHbs in the regulation of plant cell metabolism are discussed. ^

Subject Area

Biology, Cell|Chemistry, Biochemistry|Biology, Plant Physiology

Recommended Citation

Ross, Emily Jane Hume, "Plant nonsymbiotic hemoglobins: Hexacoordinated hemoglobins involved in hormonally -regulated cell differentiation" (2002). ETD collection for University of Nebraska - Lincoln. AAI3059968.