Bioimaging techniques for subcellular localization of plant hemoglobins and measurement of hemoglobin-dependent nitric oxide scavenging in planta.
Plant hemoglobins are ubiquitous in all plant families. They are expressed at low levels in specific tissues. Several studies have established that plant hemoglobins are scavengers of nitric oxide (NO) and that varying the endogenous level of hemoglobin in plant cells negatively modulates bioactivity of NO generated under hypoxic conditions or during cellular signaling. Earlier methods for determination of hemoglobin-dependent scavenging in planta were based on measuring activity in whole plants or organs. Plant hemoglobins do not contain specific organelle localization signals; however, earlier reports on plant hemoglobin have demonstrated either cytosolic or nuclear localization, depending on the method or cell type investigated. We have developed two bioimaging techniques: one for visualization of hemoglobin-catalyzed scavenging of NO in specific cells and another for visualization of subcellular localization of green fluorescent protein-tagged plant hemoglobins in transformed Arabidopsis thaliana plants.
Hebelstrup
Kim H
KH
Department of Plant Science, University of Manitoba, Winnipeg, Manitoba, Canada.
Østergaard-Jensen
Erik
E
Hill
Robert D
RD
D017360:Arabidopsis; D029681:Arabidopsis Proteins / Q000235:genetics / Q000378:metabolism; D016166:Free Radical Scavengers / Q000032:analysis* / Q000378:metabolism; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006454:Hemoglobins / Q000032:analysis / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D018613:Microscopy, Confocal / Q000379:methods; D008856:Microscopy, Fluorescence / Q000379:methods; D009569:Nitric Oxide / Q000378:metabolism*; D010940:Plant Proteins / Q000032:analysis / Q000378:metabolism; D010944:Plants / Q000378:metabolism*; D015202:Protein Engineering / Q000379:methods; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D014018:Tissue Distribution
C470347:AHB1 protein, Arabidopsis; C109042:AHB2 protein, Arabidopsis; D029681:Arabidopsis Proteins; D016166:Free Radical Scavengers; D006454:Hemoglobins; D010940:Plant Proteins; D011993:Recombinant Fusion Proteins; D049452:Green Fluorescent Proteins; D009569:Nitric Oxide