Out of the blue: Phototropins of the leaf vascular bundle sheath mediate the regulation of leaf hydraulic conductance by blue light

Abstract

The Arabidopsis (Arabidopsis thaliana) leaf veins bundle sheath cells (BSCs) – a selective barrier to water and solutes entering the mesophyll – increase the leaf radial hydraulic conductance (Kleaf) by acidifying the xylem sap by their plasma membrane H+-ATPase, AHA2. Based on this and on the BSCs’ expression of phototropins PHOT1 and PHOT2, and the known blue-light (BL)-induced Kleaf increase, we hypothesized that, resembling the guard cells, BL perception by the BSCs’ phots activates its H+-ATPase, which, consequently, upregulates Kleaf. Indeed, under BL, the Kleaf of the knockout mutant lines phot1-5, phot2-1, phot1-5 phot2-1, and aha2-4 was lower than that of the wild type (WT). BSC-only-directed complementation of phot1-5 or aha2-4 by PHOT1 or AHA2, respectively, restored the BL-induced Kleaf increase. BSC-specific silencing of PHOT1 or PHOT2 prevented such Kleaf increase. A xylem-fed kinase inhibitor (tyrphostin 9) replicated this also in WT plants. White light – ineffective in the phot1-5 mutant – acidified the xylem sap (relative to darkness) in WT and in the PHOT1-complemented phot1-5. These results, supported by BL increase of BSC protoplasts’ water permeability and cytosolic pH and their hyperpolarization by BL, identify the BSCs as a second phot-controlled water conductance element in leaves, in series with stomatal conductance. Through both, BL regulates the leaf water balance.

Leaf hydraulic maze; Differential effect of ABA on bundle-sheath, palisade, and spongy mesophyll hydraulic conductance

“…6). It is interesting to note that similarities between the regulation mechanisms of guard cells and BSC have also been reported in the context of blue light signal-transduction pathways, which initiate stomatal opening while increasing bundle sheath Pf (Iino et al, 1985;Zait et al, 2017;Torne et al, 2021;Grunwald, 2022). Our findings also reinforce the importance of the BSC as a leaf hydraulic checkpoint (Ache et al, 2010;Griffiths et al, 2013;Caringella et al, 2015).…”

A tale of two pumps: Blue light and ABA alter Arabidopsis leaf hydraulics via bundle sheath cells’ H+-pumps and channels

“…7). Based on our earlier demonstration of the AHA2's prominent role in the leaf xylem sap acidification (Grunwald et al, 2021(Grunwald et al, , 2022, we now identify AHA2 as the BSCs-specific target of the ABA-inducible signaling cascade. This resembles the Arabidopsis seedlings hypocotyl (Hayashi et al, 2014) 2D-2F).…”

“…Together, these results demonstrated that the BSCs’ external pH (pH EXT ) affected K leaf by regulating the individual BSCs’ P f (Grunwald et al, 2021). Our finding that pH EXT mediated the promoting effect of BL on the BSCs P f and through this – on K leaf (Grunwald et al, 2022), invited a question whether pH EXT mediated also the above-mentioned inhibitory effects of ABA on both the BSCs’ P f and K leaf (Shatil-Cohen et al, 2011). Furthermore, since aquaporins are reportedly inhibited by low cytosolic pH (Tournaire-Roux et al, 2003) we wondered whether the inhibitory effects of ABA on K leaf involved also the acidification of the BSCs cytosol.…”

“…We monitored the pH and membrane potential (EM) resulting from the H + -pumping using fluorescence imaging of pH-and EM-reporting ratiometric fluorescent probes: Wigoda et al, 2017;Grunwald et al, 2022), also preloaded into the isolated protoplasts.…”

“…Whether achieved by pH buffers (ibid. ), or by illumination with blue light, which stimulated the BSCs’ H + -ATPase, AHA2, via phot receptors (Grunwald et al, 2022), xylem sap acidification increased the K leaf , while xylem sap alkalinization (by a pH buffer or by an absence of blue light) decreased it. Similarly, the P f of isolated BSCs’ protoplasts was higher in an acidic solution (pH 6) than in an alkaline solution (pH 7.5; (Grunwald et al, 2021)).…”