Identification of K+ Channels in the Plasma Membrane of Maize Subsidiary Cells

Abstract

The stomatal complex of Zea mays consists of two guard cells with the pore in between them and two flanking subsidiary cells. Both guard cells and subsidiary cells are important elements for stoma physiology because a wellcoordinated transmembrane shuttle transport of potassium and chloride ions occurs between these cells during stomatal movement. To shed light upon the corresponding transport systems from subsidiary cells, subsidiary cell protoplasts were enzymatically isolated and in turn, analyzed with the patch-clamp technique. Thereby, two K + -selective channel types were identified in the plasma membrane of subsidiary cells. With regard to their voltage-dependent gating behavior, they may act as hyperpolarization-dependent K + uptake and depolarization-activated K + release channels during stomatal movement. Interestingly, the K + channels from subsidiary cells and guard cells similarly responded to membrane voltage as well as to changes in the K + gradient. Further, the inward-and outward-rectifying K + current amplitude decreased upon a rise in the intracellular free Ca 2+ level from 2 nM to the m m m mM-range. The results indicate that the plasma membrane of subsidiary cells and guard cells has to be inversely polarized in order to achieve the anti-parallel direction of K + fluxes between these cell types during stomatal movement.

The K+ Channel KZM1 Mediates Potassium Uptake into the Phloem and Guard Cells of the C4 Grass Zea mays

“…4A, inset, compare with Ref. 53), survive this mechanical treatment. Thus, KZM1 expression in the maize leaf is restricted to guard/subsidiary cells and the phloem-enriched vascular/bundle sheath strands.…”

“…By using the patch clamp technique, KZM1-like, inward-rectifying K ϩ channels have been identified in maize guard cells (60,61) and subsidiary cells (53). Thus KZM1, together with KZM2, 3 seems to carry a major part of the inward K ϩ current in Z. mays guard cells and subsidiary cells during stomatal movement (compare Refs.…”

Differential expression of K+ channels between guard cells and subsidiary cells within the maize stomatal complex

“…Isolation and collection of subsidiary cell and guard cell protoplasts Maize protoplasts were isolated from lower epidermal strips of 8-day-old leaves as previously described (Majore et al 2002) with minor modifications. Briefly, for isolation of subsidiary cell protoplasts, maize epidermal peels were incubated at 30°C for 110 min in an enzyme solution composed of 0.6% (w/v) cellulase Onozuka RS (Serva, Heidelberg, Germany), 0.064% (w/v) pectolyase Y-23 (Seishin Corp., Tokyo, Japan), 0.4% macerozyme (Serva), 0.8% (w/v) bovine serum albumine (BSA; Sigma), 0.4% (w/v) polyvinylpyrrolidone (PVP-40; Sigma), 0.8 mM CaCl 2 , pH 6.5/KOH, p=530 mosmol kg À1 (Dmannitol).…”

“…For instance, during stomatal opening subsidiary cells release K + ions while guard cells take up these osmolytes. Inward-and outward-rectifying K + currents have already been analyzed by patch-clamp studies on isolated guard cell and subsidiary cell protoplasts (Fairley-Grenot and Assmann 1992;Majore et al 2002). So far, the postulated maize orthologs of KAT1 and GORK in guard cells, however, have not been characterized at the molecular level.…”

Anatomy and Morphology

“…The subsidiary cells have been shown to function as a reservoir for K + and Cl − ions that move into guard cells during stomatal opening (Majore et al 2002;Franks & Farquhar 2007). The guard cells are slightly recessed below the external surface of the pore (Color Plate 2.8a-b), which probably establishes a boundary layer to slow transpiration (see Chapter 10).…”