Photosynthetic Electron Transport in Isolated Maize Bundle Sheath Cells

Abstract

Fragments of bundle sheath strands, free of mesophyll cells and showing a chlorophyll a/b ratio of 6.0 to 6.6 were prepared from Zea mays by a mechanical method. They were unable to photoreduce ferricyanide but were able to photoreduce the membrane-permeant 2,5-dimethylquinone at a rate of 250 to 420 microequivalents per hour per mg chlorophyll (ueq/hr.mg Chi) at 21 C. In the presence of the catalase inhibitor KCN We have attempted to reevaluate the in situ photochemical activities of chloroplasts in isolated maize BS fragments by carefully optimizing the reaction conditions. We used 2,5-dimethylquinone (DMQ) as the PSII electron acceptor, which is a membranepermeant oxidant and is by far the more stable and therefore more reliable as a PSII acceptor than is unsubstituted p-benzoquinone (24). MV, a standard PSI electron acceptor for chloroplast studies, tumed out to be useful also for isolated BS cells. This was not surprising, since MV is a widely used contact herbicide ("paraquat") and is known to penetrate the leaf tissue readily (see, e.g. 3).The in situ photochemical activities of maize BS chloroplasts are quite comparable to the highest activities reported for isolated BS chloroplasts (9). Perhaps more importantly, this paper also describes experiments in which we have successfully detected an electron transport-dependent H+ uptake by isolated BS fragments. Preparation of BS Fragments. BS fragments were isolated from secondary leaves taken from 2-to 4-week-old plants. All isolation procedures were performed at 0 to 4 C using ice-chilled media. Ten to 12 g (fresh weight) of leaves were cut transversely into Ito 2-cm segments. Two to 3 g of these segments were combined with 5 ml of the isolation medium that contained 0.5 M sucrose, 80 mM Tricine-NaOH buffer (pH 7.8), 5 mM MgCl2, 10 mm KCI, 0.2% BSA, and 0.1% PVP (PVP-40, Sigma). The leaf segments were then thoroughly macerated (4-5 min) using a pair of razor blades attached to an electric kitchen knife. This step removed most of the mesophyll cells and the epidermis from the tissue. The fragments were transferred to a beaker together with the medium and kept in suspension using a magnetic stirrer. The procedure was repeated until the entire 10 to 12 g of the original material was processed. The suspension was then filtered though two layers of Miracloth, and the tissue remaining on the cloth was briefly rinsed with distilled H20. The washed fragments were resuspended in 150 ml of the isolation medium and sheared in a Waring Blendor for 9 to 15 sec in 3-sec bursts at low speed. MATERIALS

Differential biogenesis of photosystem‐II in mesophyll and bundle‐sheath cells of ‘malic’ enzyme NADP+‐type C4 plants

“…While the thylakoid membranes of mesophyll chloroplasts are typically composed of appressed and non-appressed regions, those of bundle-sheath chloroplasts, at least in the monocotyledous species, are almost free of grana [3]. In addition, bundle-sheath chloroplasts of Sorghum bicolor and Zeu muys have been reported to be devoid or, at least, largely depleted in photosystem-I1 activity [4, 51. However, quite substantial amounts of oxygen-evolving capacity in bundle-sheath cells of the latter species have been observed by others [6,71. Conflicting results were also obtained when gel electrophoretic and immunological techniques were employed to solve this question [8 -101.…”

The Operation of Two Decarboxylases, Transamination, and Partitioning of C4 Metabolic Processes between Mesophyll and Bundle Sheath Cells Allows Light Capture To Be Balanced for the Maize C4 Pathway

“…In fact, although LEF activity in BS is often neglected (because of negligible expression of the oxygen-evolving complex [Majeran and van Wijk, 2009;Friso et al, 2010] and nonappreciable oxygenevolving activity [Meierhoff and Westhoff, 1993]), evidence that appreciable linear flow can be supported by stromal reductants such as glutathione and ascorbate has been presented (Walker and Izawa, 1979;Ivanov et al, 2001Ivanov et al, , 2005Ivanov et al, , 2007. These reductants are likely to be produced from NADPH, supplied by MAL imported from M (Kanai and Edwards, 1999;Laisk and Edwards, 2000).…”

Study of Energy Storage Processes in Bundle Sheath Cells of Zea mays

“…In our study of BS cells of maize we found no energy storage in chloroplasts through the electron transport chain when MV was present as a strong oxidant (27). In earlier reports it was noticed that the treatment of BS cells with R5P, HCO3 , and NADP stimulates photosynthetic carbon metabolism (10) and therefore increases the demand for NADPH and ATP.…”