Osmosensor-mediated control of Ca2+ spiking in pollen germination.

Higher plants survive terrestrial water deficiency and fluctuation by arresting cellular activities (dehydration) and resuscitating processes (rehydration). However, how plants monitor water availability during rehydration is unknown. Although increases in hypo-osmolarity-induced cytosolic Ca2+ concentration (HOSCA) have long been postulated to be the mechanism for sensing hypo-osmolarity in rehydration1,2, the molecular basis remains unknown. Because osmolarity triggers membrane tension and the osmosensing specificity of osmosensing channels can only be determined in vivo3-5, these channels have been classified as a subtype of mechanosensors. Here we identify bona fide cell surface hypo-osmosensors in Arabidopsis and find that pollen Ca2+ spiking is controlled directly by water through these hypo-osmosensors-that is, Ca2+ spiking is the second messenger for water status. We developed a functional expression screen in Escherichia coli for hypo-osmosensitive channels and identified OSCA2.1, a member of the hyperosmolarity-gated calcium-permeable channel (OSCA) family of proteins6. We screened single and high-order OSCA mutants, and observed that the osca2.1/osca2.2 double-knockout mutant was impaired in pollen germination and HOSCA. OSCA2.1 and OSCA2.2 function as hypo-osmosensitive Ca2+-permeable channels in planta and in HEK293 cells. Decreasing osmolarity of the medium enhanced pollen Ca2+ oscillations, which were mediated by OSCA2.1 and OSCA2.2 and required for germination. OSCA2.1 and OSCA2.2 convert extracellular water status into Ca2+ spiking in pollen and may serve as essential hypo-osmosensors for tracking rehydration in plants.

OSCA1 is an osmotic specific sensor: a method to distinguish Ca2+ -mediated osmotic and ionic perception.

Genetic mutants defective in stimulus-induced Ca2+ increases have been gradually isolated, allowing the identification of cell-surface sensors/receptors, such as the osmosensor OSCA1. However, determining the Ca2+ -signaling specificity to various stimuli in these mutants remains a challenge. For instance, less is known about the exact selectivity between osmotic and ionic stresses in the osca1 mutant. Here, we have developed a method to distinguish the osmotic and ionic effects by analyzing Ca2+ increases, and demonstrated that osca1 is impaired primarily in Ca2+ increases induced by the osmotic but not ionic stress. We recorded Ca2+ increases induced by sorbitol (osmotic effect, OE) and NaCl/CaCl2 (OE + ionic effect, IE) in Arabidopsis wild-type and osca1 seedlings. We assumed the NaCl/CaCl2 total effect (TE) = OE + IE, then developed procedures for Ca2+ imaging, image analysis and mathematic fitting/modeling, and found osca1 defects mainly in OE. The osmotic specificity of osca1 suggests that osmotic and ionic perceptions are independent. The precise estimation of these two stress effects is applicable not only to new Ca2+ -signaling mutants with distinct stimulus specificity but also the complex Ca2+ signaling crosstalk among multiple concurrent stresses that occur naturally, and will enable us to specifically fine tune multiple signal pathways to improve crop yields.

[Studies on chemical constituents from Phyllostachys pubescens].

OBJECTIVE: To study the chemical constituents from Phyllostachys pubescens. METHODS: A extract of P. pubescens was prepared by decocting with boiling water, chromatographed on macroporus resin column and eluted by ethanol. The eluant was separated on silica gel column by eluting with various proportions of chloroform-methanol-water. UV, MS, 1H NMR and 13C NMR were used to elucidate these chemical constituents structure. RESULT: Two compounds were isolated as vitexin (I) and orientin (II). CONCLUSION: Two compounds were isolated from this genus for the first time.

[Simultaneous determination of atractylenolide III, atractylenolide I and atractylon in Artactylodis macrocephala using microbore liquid chromatography].

A microbore liquid chromatography (microbore LC) has been developed for the simultaneous determination of atractylenolide III, atractylenolide I and atractylon in Artactylodis macrocephala. In the method, a Microsil C18 column (150 mm x 1.0 mm) was used with the mobile phase containing methanol-acetonitrile-water for gradient elution, the flow rate was 50 microL/min, and the detection wavelength was set at 220 nm. Under the optimized separation conditions, every component was separated thoroughly. The relationships between the concentrations and the peak areas of the three components were all linear. The recoveries were 96.86% for atractylenolide III, 97.13% for atractylenolide I and 98.06% for atractylon, the relative standard deviations (RSD) were 1.63%, 1.31% and 0.39%, respectively. The method is simple, convenient, and can be used for the quality control of Artactylodis macrocephala.