Methods to Study Plant Programmed Cell Death
Programmed cell death (PCD) is a critical component of plant development, defense against invading pathogens, and response to environmental stresses. In this chapter, we provide detailed technical methods for studying PCD associated with plant development or induced by abiotic stress. A root hair assay or electrolyte leakage assay are excellent techniques for the quantitative determination of PCD and/or cellular injury induced in response to abiotic stress, whereas the lace plant provides a unique model that facilitates the study of genetically regulated PCD during leaf development.
Key wordsProgrammed cell death Root hair assay Electrolyte leakage Developmental programmed cell death Leaf morphogenesis Abiotic stress
Department of Agriculture, Food & Marine (Ireland) is acknowledged for funding the work on ryegrass stress responses in the McCabe lab (VICCI grant. 14/S/819). The authors extend their thanks to the National Sciences and Engineering Research Council of Canada (NSERC) and the Killam Trusts for providing PhD funding to AND. AHLANG received funding from the Canadian Foundation for Innovation (CFI; Leaders Opportunity Fund) and the NSERC Discovery Grants Program.
Time-lapse video of perforation formation via developmentally regulated programmed cell death (PCD) in a window stage lace plant (Aponogeton madagascariensis) leaf. Differential interference contrast (DIC) and dark field (DF) optics were used to monitor the same areole of a window stage leaf over a period of 48 h. PCD initiates in centralized cells and radiates outward before halting several cell layers before the longitudinal and transverse veins of the leaf (MOV 15,904 kb)
Time-lapse video of a lace plant (Aponogeton madagascariensis) late programmed cell death (LPCD) stage cell. The thin and transparent leaves of the lace plant make it an ideal tissue source for live cell imaging. In this fresh, unstained specimen the nuclei and nucleoli of cells are clearly visible. Chloroplasts can also be seen streaming within the cytoplasm or associated with the nuclei in a ring-like pattern. Aggregates of defunct organelles such as mitochondria and chloroplast can be seen undergoing Brownian motion in the vacuole. Nuclear displacement occurs late in the process, which is followed by collapse of the tonoplast and then plasma membrane. Following PCD, a packaged cell corpse is observed. 50× playback speed (MOV 73,470 kb)
- 1.Kacprzyk J, Daly CT, McCabe PF (2011) The botanical dance of death: programmed cell death in plants. In: Kader JC, Delseny M (eds) Advances in botanical research, vol 60. Academic, UK, pp 169–261Google Scholar
- 12.Gunawardena AHLAN, Navachandrabala C, Kane M, Dengler NG (2006) Lace plant: a novel system for studying developmental programmed cell death. In: da Silva JA T (ed) Floriculture, ornamental and plant biotechnology: advances and tropical issues. Global Science Books, Middlesex, pp 157–162Google Scholar
- 16.Kacprzyk J, McCabe PF (2015) A root hair assay to expedite cell death research. In: Estevez JM (ed) Plant cell expansion, vol 1242, Methods Mol Biol. Humana Press, New York, pp 73–82Google Scholar
- 20.Mishiba KI, Nagashima Y, Suzuki E, Hayashi N, Ogata Y, Shimada Y, Koizumi N (2013) Defects in IRE1 enhance cell death and fail to degrade mRNAs encoding secretory pathway proteins in the Arabidopsis unfolded protein response. Proc Natl Acad Sci U S A 110:5713–5718CrossRefPubMedPubMedCentralGoogle Scholar