Presence of Both Photosystems in Guard Cells of Vicia faba L

Abstract

A new procedure is reported for high-yield isolation of guard cell protoplasts from Viia faba L. Delayed Light emission and P700 content plus absorption and fluorescence emission spectra of these protoplast extracts are reported. It is concluded that both photosystems are present. The presence of photosystem II 12Step 2. The leaf pieces were infitrated under reduced pressure with enzymic digestion medium consisting of 150 ml 0.3 M mannitol, 10 mm sodium ascorbate, 10 mm CaCl2, 4% (w/v) Cellulysin (pH 5.5). Incubation was for 1 h at 30 C in a 1-liter flask in a shaker bath (60 5-cm excursions/min).Step 3. One hundred fifty ml 0.7 M mannitol was added and incubation was continued for 1 h.Step 4. Digestion was interrupted by pouring the material over a 295-,um screen (Nitex) and washing with 0.7 M mannitol containing 10 mm sodium ascorbate. Epidermal strips (coming from both surfaces) and vein nets retained on the screen were transferred to 0.7 M mannitol containing 10 mM sodium ascorbate. Then the epidermal strips were manually separated from the vein nets and transferred to a separate container. The strips had few adhering mesophyll cells, but epidermal cells were intact.In some experiments, mesophyll cells which passed through the screen were carried through steps similar to the remainder of the isolation procedure and used as controls.Step 5. Digestion was continued for I h as in Step 2 except (a) mannitol was increased to 0.7 M, (b) volume was reduced to 100 ml, and (c) shaker-bath speed was reduced to 30 excursions/min.Step 6. The strips were collected and washed as in Step 4. If microscopic examination showed contaminating cells, Steps 5 and 6 were repeated.Step 7. Digestion was continued until guard cell protoplasts were released (about 3 h). These protoplasts were isolated by passage through a cascade of screens (nominally 295, 166, and 20 pm) and collected by centrifugation (lSOOg, 5 min). The protoplasts were washed three times with 0.7 M mannitol containing 10 mM sodium ascorbate and examined for purity.Comments. Isolation of guard-cell protoplasts has been reported (25,32, 43

Cellular Distribution of Calmodulin and Calmodulin-Binding Proteins in Vicia faba L.

“…Guard cell total protein levels (222.5 pg/cell) were similar to a previously published calculation (156 pg/cell), based on the dry weight of guard cell pairs (23). On a protein basis, guard cell protoplasts contained the highest levels of CaM activity (6.36 X 10-2 A.U./g of total protein), 2-fold more than mesophyll cell protoplasts (2.79 X 10-2 A.U./g of total protein).…”

The guard cell chloroplast: a perspective for the twenty-first century

“…Studies of fluorescence transients from gccs provide the earliest evidence for electron transport and photosynthetic carbon fixation in the gcc (Zeiger et al , 1980;Outlaw et al , 1981;Melis & Zeiger, 1982). A study on fluorescence transients from gccs of guard cells in epidermal peels detached from leaves growing in a greenhouse offers a striking demonstration of guard cell plasticity.…”

“…However, the gcc has a normal pigment composition, and functional photosystem I (PSI) and PSII (Zeiger et al , 1980;Outlaw et al , 1981;Shimazaki et al , 1982). Electron transport, oxygen evolution and photophosphorylation have been clearly demonstrated ( Fitzsimons & Weyers, 1983;Shimazaki & Zeiger, 1985).…”

Berger C. Mayne (1920–2011): a friend and his contributions to photosynthesis research

“…In collaboration with William Outlaw and others, Berger Mayne used measurements of delayed and prompt fluorescence and P700 content to demonstrate that both photosystems are present there (Outlaw et al 1981). (Also see Ogawa et al 1982 for a fluorescence study on guard cells.)…”

“…He requested the Editor Martin Gibbs (1922Gibbs ( -2006 to hold up their paper and publish it back to back with Eduardo's (which was submitted after theirs), which he did. They were published in the January 1981 issue (Outlaw et al 1981;Zeiger et al 1981). Somehow, the offprints of Zeiger's were misdated to 1980, so one might read that Berger and Outlaw confirmed Zeiger's findings.…”