Hypersensitive cell death (HR) of adaxial leaf epidermal cells of barley containing the M1a gene for resistance to powdery mildew germlings, Erysiphe graminis f. sp. hordei, was investigated for temporal relations between cell death, onset of autofluorescence, and accumulation of insoluble silicon (Si). Physiological cell viability or death of barley epidermal cells was determined by concomitant uptake of the vital dye neutral red, and by cell plasmolysis using alpha methyl-d-glucose. Cell death, as judged by lack of neutral red uptake and lack of plasmolysis, first occurred in a low percentage of attacked epidermal cells at 15 h after inoculation and increased to a maximum of 72% at 21 and 24 h. Cells judged dead by physiological criteria always exhibited whole-cell autofluoresence by blue light excitation. Autofluorescence was weak in dying and recently dead cells, but it increased in intensity with time following cell death. It was concluded that autofluorescence accompanies cell death and is perhaps due to release of phenolic compounds from cell vacuoles after cell membranes have lost their semipermeable properties. Insoluble Si accumulation, as determined by energy dispersive X-ray microanalysis using a scanning electron microscope, did not directly correspond to the initial appearance of autofluorescence in HR-dead cells, but occurred gradually after death and in apparent relation to increasing autofluorescence intensity. An hypothesis is stated that phenolics are released by decompartmentalization following cell death and accumulate in the dead cell's wall area causing autofluorescence, and that these phenolics form insoluble complexes with Si moving apoplastically in epidermal wall areas due to passive, transpiration stream transport.
Bibliographical noteFunding Information:
Research was conducted under ,Minnesota Agricuhural Experimental Station project No. 22-70, with support provided by USDA Competitive Grants Office--Grant 85-CRCR-I-1596; and the Science and Technology Agency of Japan.