Measuring Plant Protein Secretion

Research output: Contribution to journalArticle (Academic Journal)

Abstract

Fluorescent protein fusions are widely used for visualizing the subcellular localization and mobility of intercellular proteins. There is now a variety of colors, expression vectors, and photoactivated molecules to choose from, each with their own strengths and limitations. In this chapter, the methodologies for expressing and quantifying protein secretion with fluorescent protein fusion constructs using two separate protocols-one in which the retention of a transiently expressed fluorescent marker is measured in seedling roots to quantify a block in secretion, and one in which the secretion of a fluorescent marker into the space of the apoplast is measured to quantify secretion in plant leaves-are described. In the first protocol, seedling roots are transiently transformed with multicistronic constructs; and in the second protocol, markers can be stably expressed and controlled under an inducible promoter in mature plants. Both methods provide tools for quantifying protein secretion and visualizing defects in secretion pathways in Arabidopsis.

Original languageEnglish
Pages (from-to)199-207
Number of pages9
JournalMethods in Molecular Biology
Volume1662
DOIs
Publication statusPublished - 2017

Keywords

  • Agrobacterium tumefaciens/genetics
  • Arabidopsis/drug effects
  • Arabidopsis Proteins/genetics
  • Dexamethasone/pharmacology
  • Gene Expression
  • Genes, Reporter
  • Green Fluorescent Proteins/genetics
  • Luminescent Proteins/genetics
  • Oligopeptides/genetics
  • Optical Imaging/methods
  • Plant Leaves/drug effects
  • Plant Roots/drug effects
  • Plants, Genetically Modified
  • Plasmids/chemistry
  • Promoter Regions, Genetic
  • Protein Sorting Signals/genetics
  • Protein Transport
  • Qa-SNARE Proteins/genetics
  • Recombinant Fusion Proteins/genetics
  • Seedlings/drug effects
  • Transformation, Genetic
  • Transport Vesicles/metabolism
  • Vacuoles/metabolism

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