Plant HKT proteins comprise a family of cation transporters together with prokaryotic KtrB, TrkH, and KdpA transporter subunits and fungal Trk proteins. These transporters contain four loop domains in one polypeptide with a proposed distant homology to K ؉ channel selectivity filters. Functional expression in yeast and Xenopus oocytes revealed that wheat HKT1 mediates Na ؉ -coupled K ؉ transport. Arabidopsis AtHKT1, however, transports only Na ؉ in eukaryotic expression systems. To understand the molecular basis of this difference we constructed a series of AtHKT1͞HKT1 chimeras and introduced point mutations to AtHKT1 and wheat HKT1 at positions predicted to be critical for K ؉ selectivity. A single-point mutation, Ser-68 to glycine, was sufficient to restore K ؉ permeability to AtHKT1. The reverse mutation in HKT1, Gly-91 to serine, abrogated K ؉ permeability. This glycine in P-loop A of AtHKT1 and HKT1 can be modeled as the first glycine of the K ؉ channel selectivity filter GYG motif. The importance of such filter glycines for K ؉ selectivity was confirmed by interconversion of Ser-88 and Gly-88 in the rice paralogues OsHKT1 and OsHKT2. Surprisingly, all HKT homologues known from dicots have a serine at the filter position in P-loop A, suggesting that these proteins function mainly as Na ؉ transporters in plants and that Na ؉ ͞K ؉ symport in HKT proteins is associated with a glycine in the filter residue. These data provide experimental evidence that the glycine residues in selectivity filters of HKT proteins are structurally related to those of K ؉ channels.

In response to environmental variation, angiosperm trees bend their stems by forming tension wood, which consists of a cellulose-rich G (gelatinous)-layer in the walls of fiber cells and generates abnormal tensile stress in the secondary xylem. We produced transgenic poplar plants overexpressing several endoglycanases to reduce each specific polysaccharide in the cell wall, as the secondary xylem consists of primary and secondary wall layers. When placed horizontally, the basal regions of stems of transgenic poplars overexpressing xyloglucanase alone could not bend upward due to low strain in the tension side of the xylem. In the wild-type plants, xyloglucan was found in the inner surface of G-layers during multiple layering. In situ xyloglucan endotransglucosylase (XET) activity showed that the incorporation of whole xyloglucan, potentially for wall tightening, began at the inner surface layers S1 and S2 and was retained throughout G-layer development, while the incorporation of xyloglucan heptasaccharide (XXXG) for wall loosening occurred in the primary wall of the expanding zone. We propose that the xyloglucan network is reinforced by XET to form a further connection between wall-bound and secreted xyloglucans in order to withstand the tensile stress created within the cellulose G-layer microfibrils.

The differences between cell wall formation at night, when the tangential strain used as an index of the volumetric changes in differentiating cells is high, and in the day, when the tangential strain is low, were investigated in Cryptomeria japonica D. Don. Samples containing differentiating xylem were collected at 0500 hours and 1400 hours. The innermost surface of developing secondary walls in differentiating tracheids was observed by field emission scanning electron microscopy. In the specimens collected at 0500 hours, an amorphous material was observed covering the cellulose microfibrils. The cell wall surface was immunogold-labeled with an anti-glucomannan antiserum. After chlorite treatment, the amorphous material disappeared, and immunogold labeling was rarely observed. In the specimens collected at 1400 hours, cellulose microfibrils were clearly evident, and amorphous material and immunogold labeling were rarely observed. We thus confirmed that much amorphous material containing glucomannans is observed at night, when differentiating tracheids are turgid due to the increase in their volume, while the amorphous material was rarely observed during the day when cellulose microfibrils are clearly observed.

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