PubMed · 7027289
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B H Sullivan. 1981. Gastrointestinal bleeding.. https://pubmed.ncbi.nlm.nih.gov/7027289/
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Rhamnogalacturonan II (RG-II) is a structurally complex pectic polysaccharide present in the walls of growing plant cells. We now report that RG-II, released by endopolygalacturonase treatment of the walls of suspension-cultured sycamore cells and etiolated pea stems, exists mainly as a dimer that is cross-linked by a borate ester. The borate ester is completely hydrolyzed at room temperature within 30 min at pH 1, partially hydrolyzed between pH 2 and 4, and stable above pH 4. The dimer is formed in vitro between pH 2.4 and 6. 2 by treating monomeric RG-II (0.5 mM) with boric acid (1.2 mM); the dimer formed after 24 h at pH 3.4 and 5.0 accounts for approximately 30 and approximately 5%, respectively, of the RG-II. In contrast, the dimer accounts for approximately 80 and approximately 54% of the RG-II when the monomer is treated for 24 h at pH 3.4 and 5.0, respectively, with boric acid and 0.5 m Sr2+, Pb2+, or Ba2+. The amount of dimer formed at pH 3.4 or 5.0 is not increased by addition of 0.5 mM Ca2+, Cd2+, Cu2+, Mg2+, Ni2+, and Zn2+. Steric considerations appear to regulate dimer formation since those divalent cations that enhance dimer formation have an ionic radius >1.1 A. Our data suggest that the borate ester is located on C-2 and C-3 of two of the four 3'-linked apiosyl residues of dimeric RG-II. Our results, taken together with the results of two previous studies (Kobayashi, M., Matoh, T., and Azuma, J.-I. (1996) Plant Physiol. 110, 1017-1020; Ishii, T., and Matsunaga, T. (1996) Carbohydr. Res. 284, 1-9) provide substantial evidence that this plant cell wall pectic polysaccharide is covalently cross-linked.
1. A cloned inwardly rectifying K+ channel, IRK2, was expressed in a human cell line, human embryonic kidney (HEK) 293T. Its electrophysiological properties were examined using the patch clamp technique in the whole-cell, cell-attached and inside-out patch configurations. 2. The cells transfected with IRK2 cDNA exhibited a K+ current which showed classical properties of inwardly rectifying K+ channels at both whole-cell and single-channel levels. 3. In the inside-out patch configuration, intracellular Mg2+ (Mg2+i blocked the outward currents in a voltage-dependent and virtually time-independent manner. Mg2+i (1-100 microM) caused a decrease in the unitary current amplitude of the IRK2 channel by inducing subconducting levels. 4. In the absence of Mg2+i, intracellular spermine blocked the outwardly flowing IRK2 currents in a voltage- and time-dependent manner. Spermine (1-100 nM) did not affect the unitary channel current amplitude but reduced the channel open probability. The spermine block showed a slower time and steeper voltage dependence than the Mg2+i++ block. 5. When both these blockers were present, Mg2+i apparently attenuated the inhibitory effect of spermine on the outwardly flowing IRK2 currents. This interaction was voltage and time dependent, and could be well explained by a model in which Mg2+i and spermine competitively bind to the channel with their individual first-order kinetics. This competition would induce time-dependent transits of the channel between the Mg2+i -and spermine-blocked states via a single open state, thereby preserving a certain size of persistent outward currents at depolarized potentials.
The mannose-specific lectin, concanavalin A (ConA), activates Ca2+ entry in human neutrophils by an as yet poorly defined mechanism. The question of whether the sugar specificity of lectins influences signal transduction is unresolved too. Therefore, we studied the effects of ConA in comparison to those of the beta-galactoside-specific lectin, mistletoe lectin I (MLI), on cation entry and exocytosis in human neutrophils. ConA- and MLI-activated influx of Ca2+, Mn2+, Ba2+, Sr2+, and Na+. Lectin-induced cation influxes were inhibited by 1-(beta-[3-(4-methoxyphenyl)propoxy]-4-methoxy-phenethyl) -1H-imidazole hydrochloride (SK&F 96365) and Gd3+. There were differences in the effectiveness of lectins to activate cation entry and of SK&F 96365, Gd3+, and modulators of protein phosphorylation to block entry. MLI but not ConA inhibited thapsigargin-induced Ca2+ entry. Under whole-cell voltage-clamp conditions, MLI activated an inward current that was substantially reduced by removal of extracellular Na+. ConA and MLI synergistically activated Ca2+ entry and lysozyme release. SK&F 96365 and removal of extracellular Ca2+ and Na+ partially inhibited exocytosis. Our data show the following: (1) ConA and MLI activate monovalent and divalent cation entry in human neutrophils by a SK&F 96365- and Gd3+-sensitive pathway, presumably nonselective cation channels. (2) Ca2+ and Na+ entry are involved in the activation of exocytosis by lectins. (3) The differential and/or synergistic effects of ConA and MLI on cation entry and exocytosis may be attributable to mannose- and beta-galactoside-specific activation of signal transduction pathways, i.e., activation of multiple and differentially regulated subtypes of nonselective cation channels.