Effect of Chlorsulfuron on Morphogenetic and Disordered Cell Division in Cultures of Passiflora edulis

We examined the effects of a sulfonylurea herbicide, chlorsulfuron, which is known as a potent inhibitor of plant cell division, on morphogenetic cell division and disorganized cell division using the culture system of multiple shoot primordia and callus of Passiflora edulis. The multiple shoot primordia tissue treated with chlorsulfuron failed to achieve shoot morphogenesis, and a large part of the tissue was necrotized during the posttreatment culture, even when it was washed and transferred to chlorsulfuron-free medium. The inhibition of Passiflora shoot morphogenesis by chlorsulfuron was not reversed by the simultaneous addition of branched amino acids, which are known to reverse the inhibitory effect of chlorsulfuron. In contrast, the same treatment of chlorsulfuron on the callus did not kill the cells, although the growth resumption was retarded by a prolonged lag period. The addition of branched amino acids enhanced the recovery growth of the chlorsulfuron-treated callus. These results suggest that the inhibition of disorganized cell division (callus growth) by chlorsulfuron is reversible, whereas morphogenetic cell division (shoot morphogenesis), which is under complex regulation, is inhibited irreversibly by chlorsulfuron. Qualitative differences between morphogenetic cell division and disordered simple proliferative cell division are discussed. Received November 17, 1997; accepted June 4, 1998     

KATP channels in mesenchymal stromal stem cells: strong up-regulation of Kir6.2 subunits upon osteogenic differentiation

The promising use of mesenchymal stromal cells (MSC) in regenerative technologies accounts for necessity of detailed study of their physiology. Proliferation and differentiation of multipotent cells often involve changes in their metabolic state. In the present study, we analyzed the expression of ATP-sensitive potassium (K_ATP) channels in MSC and upon in vitro differentiation. K_ATP channels are present in many cells and regulate a variety of cellular functions by coupling cell metabolism with membrane potential. Kir6.1, Kir6.2 and SUR2A were expressed in undifferentiated MSC, whereas SUR2B and SUR1 were not detected on cDNA and protein level. Upon adipogenic differentiation Kir6.1 and SUR2A showed a significant reduction of the amount of mRNA by 84% and 95%, respectively, whereas Kir6.2 expression was unchanged. Osteogenic differentiation strongly up-regulated Kir6.2 mRNA (28-fold) whereas Kir6.1 and SUR2A showed no significant change in expression. Quantitative Western blot analysis and immunofluorescence staining confirmed the elevated expression of Kir6.2 upon osteogenic differentiation. Taken together, expression changes of K_ATP channels may contribute to in vitro differentiation of MSC and represent changes in the metabolic state of the developing tissue.     

磺酰脲类除草剂残留的微生物降解研究进展

磺酰脲类除草剂是一类高效、低毒和高选择性的除草剂, 此类除草剂能有效地防除阔叶杂草, 其中有些品种对禾本科杂草也有抑制作用。由于该类除草剂易残留药害及容易对地表水造成污染, 因而其在环境中的持久性和环境安全性备受人们关注。本文综述了磺酰脲类除草剂的应用概况及其作用机理、降解磺酰脲类除草剂的常见微生物种类及影响微生物降解效率的因素, 最后展望了微生物修复技术与抗除草剂的转基因作物是解决除草剂残留药害的最佳途径。     

Pollen genotype selection for a simply inherited qualitative factor determining resistance to chlorsulfuron in maize

Pollen genotype selection for genes expressed in both the haploid and diploid phases of the plant life cycle can lead to correlated responses detectable in the sporophyte. A pollen selection was conducted in two genetic backgrounds of maize (Zea mays L.) for chlorsulfuron resistance, conferred by the XA17 allele. Plants of two backcross (BC) lines segregating 1 (heterozygote, resistant) : 1 (homozygote, susceptible) for chlorsulfuron resistance were used as pollen donor. Selection treatment was applied during microspore development, and tassels were cut about 10 days before anthesis and placed into a liquid medium with or without 40 mg l^–1 of chlorsulfuron. Pollen was used to fertilize an unrelated susceptible genotype (tester). The resulting testcrosses (TC) were evaluated in the greenhouse by spraying seedlings with chlorsulfuron at 23 g ha^–1. Non-selected TC progenies derived from heterozygous BC plants showed a proportion of resistant and susceptible plants close to the expected 1:1 ratio, while non-selected TC progenies derived from homozygous BC plants showed susceptible plants only. Selected TC progenies derived from heterozygous BC plants showed a frequency of resistant plants ranging from 89% to 100%. BC plants homozygous for the susceptible allele, subjected to selection treatment, gave poor seed set, and no resistant plants were found in their TC progenies. Resistant TC plants obtained through pollen genotype selection were selfed, and the proportion of resistant plants was close to 3:1 in all selfed families, in accordance with the hypothesis that all of them inherited the XA17 allele through selection. In this study, pollen genotype selection was extremely effective, and its effect persisted in the second sporophytic generation. Received: 19 November 1999 / Accepted: 30 April 2000     

On potential interactions between non-selective cation channel TRPM4 and sulfonylurea receptor SUR1

The sulfonylurea receptor SUR1 associates with Kir6.2 or Kir6.1 to form K(ATP) channels, which link metabolism to excitability in multiple cell types. The strong physical coupling of SUR1 with Kir6 subunits appears exclusive, but recent studies argue that SUR1 also modulates TRPM4, a member of the transient receptor potential family of non-selective cation channels. It has been reported that, following stroke, brain, or spinal cord injury, SUR1 is increased in neurovascular cells at the site of injury. This is accompanied by up-regulation of a non-selective cation conductance with TRPM4-like properties and apparently sensitive to sulfonylureas, leading to the postulation that post-traumatic non-selective cation currents are determined by TRPM4/SUR1 channels. To investigate the mechanistic hypothesis for the coupling between TRPM4 and SUR1, we performed electrophysiological and FRET studies in COSm6 cells expressing TRPM4 channels with or without SUR1. TRPM4-mediated currents were Ca(2+)-activated, voltage-dependent, underwent desensitization, and were inhibited by ATP but were insensitive to glibenclamide and tolbutamide. These properties were not affected by cotransfection with SUR1. When the same SUR1 was cotransfected with Kir6.2, functional K(ATP) channels were formed. In cells cotransfected with Kir6.2, SUR1, and TRPM4, we measured K(ATP)-mediated K(+) currents and Ca(2+)-activated, sulfonylurea-insensitive Na(+) currents in the same patch, further showing that SUR1 controls K(ATP) channel activity but not TRPM4 channels. FRET signal between fluorophore-tagged TRPM4 subunits was similar to that between Kir6.2 and SUR1, whereas there was no detectable FRET efficiency between TRPM4 and SUR1. Our data suggest that functional or structural association of TRPM4 and SUR1 is unlikely.     

Role of Derlin-1 protein in proteostasis regulation of ATP-sensitive potassium channels

ATP-sensitive potassium (K(ATP)) channels composed of sulfonylurea receptor 1 (SUR1) and Kir6.2 regulate insulin secretion by linking glucose metabolism with membrane potential. The number of K(ATP) channels in the plasma membrane affects the sensitivity of β-cells to glucose. Aberrant surface channel expression leads to insulin secretion disease. Previously, we have shown that K(ATP) channel proteins undergo endoplasmic reticulum (ER)-associated degradation (ERAD) via the ubiquitin-proteasome pathway, and inhibition of proteasome function results in an increase in channel surface expression. Here, we investigated whether Derlin-1, a protein involved in retrotranslocation of misfolded or misassembled proteins across the ER membrane for degradation by cytosolic proteasomes, plays a role in ERAD and, in turn, biogenesis efficiency of K(ATP) channels. We show that both SUR1 and Kir6.2 form a complex with Derlin-1 and an associated AAA-ATPase, p97. Overexpression of Derlin-1 led to a decrease in the biogenesis efficiency and surface expression of K(ATP) channels. Conversely, knockdown of Derlin-1 by RNA interference resulted in increased processing of SUR1 and a corresponding increase in surface expression of K(ATP) channels. Importantly, knockdown of Derlin-1 increased the abundance of disease-causing misfolded SUR1 or Kir6.2 proteins and even partially rescued surface expression in a mutant channel. We conclude that Derlin-1, by being involved in ERAD of SUR1 and Kir6.2, has a role in modulating the biogenesis efficiency and surface expression of K(ATP) channels. The results suggest that physiological or pathological changes in Derlin-1 expression levels may affect glucose-stimulated insulin secretion by altering surface expression of K(ATP) channels.     

Synergistic effect on insulin secretion from INS-1 cells of a sulfonylurea and a phosphodiesterase 3 inhibitor

The effects on insulin secretion from INS-1 cells of varying concentrations of the sulfonylurea glyburide and the PDE3 inhibitor milrinone, separately and in combination were measured. Over a range of concentrations the effects of the two drugs in combination were more than additive. A response surface model was fit to the data and was found to describe the data well. From this model, it was apparent that a significant synergistic effect upon insulin secretion existed over a wide range of combinations of the two drugs.     

Herbicide-tolerant corn by pollen selection

Summary Maize pollen was exposed to the herbicide Chlorsulfuron (CS), and segregation for tolerance was observed. The resulting plant generation exhibited significantly greater tolerance to CS than other (control) progeny. Since the increase in tolerance occurred after only one generation of pollen exposure, this result demonstrates that pollen selection can be used to develop herbicide-resistant crop species, even when the species are not amenable to cell culture. It also suggests a possible mechanism for the rapid evolution of herbicide tolerance in weeds.     

Recognition of sulfonylurea receptor (ABCC8/9) ligands by the multidrug resistance transporter P-glycoprotein (ABCB1): functional similarities based on common structural features between two multispecific ABC proteins

ATP-sensitive K(+) (K(ATP)) channels are the target of a number of pharmacological agents, blockers like hypoglycemic sulfonylureas and openers like the hypotensive cromakalim and diazoxide. These agents act on the channel regulatory subunit, the sulfonylurea receptor (SUR), which is an ABC protein with homologies to P-glycoprotein (P-gp). P-gp is a multidrug transporter expressed in tumor cells and in some healthy tissues. Because these two ABC proteins both exhibit multispecific recognition properties, we have tested whether SUR ligands could be substrates of P-gp. Interaction with P-gp was assayed by monitoring ATPase activity of P-gp-enriched vesicles. The blockers glibenclamide, tolbutamide, and meglitinide increased ATPase activity, with a rank order of potencies that correlated with their capacity to block K(ATP) channels. P-gp ATPase activity was also increased by the openers SR47063 (a cromakalim analog), P1075 (a pinacidil analog), and diazoxide. Thus, these molecules bind to P-gp (although with lower affinities than for SUR) and are possibly transported by P-gp. Competition experiments among these molecules as well as with typical P-gp substrates revealed a structural similarity between drug binding domains in the two proteins. To rationalize the observed data, we addressed the molecular features of these proteins and compared structural models, computerized by homology from the recently solved structures of murine P-gp and bacterial ABC transporters MsbA and Sav1866. Considering the various residues experimentally assigned to be involved in drug binding, we uncovered several hot spots, which organized spatially in two main binding domains, selective for SR47063 and for glibenclamide, in matching regions of both P-gp and SUR.     

Synthesis,structural characterization and in vivo anti-diabetic evaluation of some new sulfonylurea derivatives in normal and silicate coated nanoparticle forms as anti-hyperglycemic agents

Series of new sulfonylurea derivatives (gliclazide analogues) was synthesized and characterized. Thus, p-tolylsulfonylisocyanate was left to react with different amino derivatives under mild conditions to afford the desired sulfonylurea derivatives 1–5. The molecular structure of the compound N-(2,6-Dichlorophenylcarbamoyl)-4-methylbenzenesulfonamide, 1c has been elucidated by single crystal X-ray diffraction. Anti-diabetic properties of the synthesized compounds relative to anti-diabetic drug (gliclazidem MR60) were carried out, where most of the tested compounds showed significant activity for reducing the blood glucose level. The results revealed that compounds 1c and 5 showed better anti-diabetic activities compared with gliclazide. Activity of the most potent derivatives of sulfonylurea compounds namely 1c and 5 were increased using coated nanostructure tetraethyl orthosilicate (TEOS) as a modified release (MR) agent. The effect of the prepared sulfonylurea compounds against the diabetic condition was investigated using specific selected biomarkers as of liver enzyme activities as transaminases (AST, ALT) and alkaline phosphatase (ALP), lipids profiles; total cholesterol (TC), triacylglycerols (TG) and total lipid (TL). The antioxidants, oxidative stress biomarkers and histological examination were also examined and discussed.