Spodoptera frugiperda is a highly destructive pest that has become a global problem due to its robust reproductive and migratory capabilities.Transient receptor potential(TRP)channels,which constitute a vast ion chann...Spodoptera frugiperda is a highly destructive pest that has become a global problem due to its robust reproductive and migratory capabilities.Transient receptor potential(TRP)channels,which constitute a vast ion channel family,play pivotal roles in sensing the external environment and maintaining internal homeostasis in insects.TRP channels have been widely investigated for their critical roles in regulating various insect behaviors in recent years.In this study,we identified 15 TRP gene loci encoding 26 transcripts in the genome of S.frugiperda and analyzed their expression profiles at different developmental stages.The results revealed that S.frugiperda possesses four TRPC genes,six TRPA genes,one TRPM gene,two TRPV genes,one TRPN gene,and one TRPML gene,while a canonical TRPP is absent.Moreover,the SfruTRPA1 was functionally characterized using the Xenopus oocyte expression system.The results showed that SfruTRPA1 is activated by temperature increases from 20 to 45℃,and there is no significant desensitization after repeated stimuli within the same temperature range.Additionally,SfruTRPA1 is activated by certain natural chemicals,including allyl isothiocyanate(AITC)and cinnamaldehyde(CA).These findings provide valuable insights to the TRP genes in S.frugiperda.展开更多
OBJECTIVE Two-pore domain potassium channel subtype TREK-1 was widely proved to be activated by inhalational anesthet⁃ics such as chloroform,diethyl ether,halothane,and isoflurane.But little is known about whether TRE...OBJECTIVE Two-pore domain potassium channel subtype TREK-1 was widely proved to be activated by inhalational anesthet⁃ics such as chloroform,diethyl ether,halothane,and isoflurane.But little is known about whether TREK-1 was also a potentially important target of intravenous anesthetics.Etomidate is a popularly used intravenous anesthetic with good safety in clinic.The action of etomidate on TREK-1 was seldom reported.METHODS AND RESULTS By using patch-clamp whole-cell recording tech⁃niques,we found for the first time that etomidate could bidirectionally regulate the TREK-1 potassi⁃um channel in CHO/TREK-1 cells.TREK-1 current amplitudes were observed after the administra⁃tion of etomidate at concentrations ranging from 3 to 100μmol·L-1.Etomidate activated TREK-1 current at concentrations of 3,10,and 15μmol·L-1 with maximum activation at 10μmol·L-1.Interest⁃ingly,at higher concentrations from 20 to 100μmol·L-1,etomidate inhibited TREK-1 current in a concentration-dependent way.According to the concentration-response curve,the fitted criti⁃cal concentration of etomidate between TREK-1 activation and inhibition was 20.7μmol·L-1,which close to the result that etomidate had no obvious effect on TREK-1 at 20μmol·L-1.In addition,etomidate 10μmol·L-1 induced a significant mem⁃brane potential hyperpolarization while etomidate 30μmol·L-1 showed obvious membrane potential depolarization.Furthermore,the bidirectional regulation still existed when the extracellular pH of CHO/TREK-1 cells was decreased.CONCLUSION TREK-1 is activated by etomi⁃date at clinically relevant concentrations but inhib⁃ited by supraclinical concentrations of etomidate,which is different to other volatile anesthetics.TREK-1 might be a potential target for anesthetic such as etomidate and the complicated bidirec⁃tional regulation mechanism of etomidate needed to be fully studied in the future.展开更多
This study was aimed to examine the effect of TREK-1 silencing on the function of astrocytes. Three 21-nucleotide small interfering RNA (siRNA) duplexes (siT1, siT2, siT3) targeting TREK-1 were constructed. Cy3-labele...This study was aimed to examine the effect of TREK-1 silencing on the function of astrocytes. Three 21-nucleotide small interfering RNA (siRNA) duplexes (siT1, siT2, siT3) targeting TREK-1 were constructed. Cy3-labeled dsRNA oligmers were used to determine the transfection efficiency in cultured astrocytes. TREK-1-specific siRNA duplexes (siT1, siT2, siT3) at the optimal concentration were transfected into cultured astrocytes, and the most efficient siRNA was identified by the method of immunocytochemical staining and Western blotting. The proliferation of astrocytes tranfected with TREK-1-targeting siRNA under hypoxia condition was measured by fluorescence-activated cell sorting (FACS). The results showed that TREK-1 was expressed in cultured astrocytes. The dsRNA oligmers targeting TREK-1 could be transfected efficiently in cultured astrocytes and down-regulate the expression of TREK-1 in astrocytes. Moreover, the down-regulation of TREK-1 in astrocytes contributed to the proliferation of astrocytes under hypoxia condition as determined by cell cycle analysis. It was concluded that siRNA is a powerful technique that can be used to knockdown the expression of TREK-1 in astrocytes, which helps further investigate the function of TREK-1 channel in astrocytes under physicological and pathological condition.展开更多
Potassium transporters play crucial roles in K^＋ uptake and translocation in plants. However, so far little is known about the regulatory mechanism of potassium transporters. Here, we show that a Shaker-like potassiu...Potassium transporters play crucial roles in K^＋ uptake and translocation in plants. However, so far little is known about the regulatory mechanism of potassium transporters. Here, we show that a Shaker-like potassium channel AtKC1, encoded by the AtLKT1 gene cloned from the Arabidopsis thaliana low-K^＋ （LK）-tolerant mutant Atlktl, significantly regulates AKTl-mediated K^＋ uptake under LK conditions. Under LK conditions, the Atkcl mutants maintained their root growth, whereas wild-type plants stopped their root growth. Lesion of AtKC1 significantly enhanced the tolerance of the Atkcl mutants to LK stress and markedly increased K^＋ uptake and K^＋ accumulation in the Atkclmutant roots under LK conditions. Electrophysiological results showed that AtKC1 inhibited the AKT1-mediated inward K^＋ currents and negatively shifted the voltage dependence of AKT1 channels. These results demonstrate that the ‘silent＇ K^＋ channel α-subunit AtKC1 negatively regulates the AKTl-mediated K^＋ uptake in Arabidopsis roots and consequently alters the ratio of root-to-shoot under LK stress conditions.展开更多
AIM: To investigate the mechanisms of chloride intracellular channel 1 (CLIC1) in the metastasis of colon cancer under hypoxia-reoxygenation (H-R) conditions.
AIM:To investigate the effect of chloride intracellular channel 1(CLIC1) on the cell proliferation,apoptosis,migration and invasion of gastric cancer cells.METHODS:CLIC1 expression was evaluated in human gastric cance...AIM:To investigate the effect of chloride intracellular channel 1(CLIC1) on the cell proliferation,apoptosis,migration and invasion of gastric cancer cells.METHODS:CLIC1 expression was evaluated in human gastric cancer cell lines SGC-7901 and MGC-803 by real time polymerase chain reaction(RT-PCR).Four segments of small interference RNA(siRNA) targeting CLIC1 mRNA and a no-sense control segment were designed by bioinformatics technology.CLIC1 siRNA was selected using Lipofectamine 2000 and transfected transiently into human gastric cancer SGC-7901 and MGC-803 cells.The transfected efficiency was observed under fluorescence microscope.After transfection,mRNA expression of CLIC1 was detected with RT-PCR and Western blotting was used to detect the protein expression.Proliferation was examined by methyl thiazolyl tetrazolium and apoptosis was detected with flow cytometry.Polycarbonate membrane transwell chamber and Matrigel were used for the detection of the changes of invasion and migration of the two cell lines.RESULTS:In gastric cancer cell lines SGC-7901 and MGC-803,CLIC1 was obviously expressed and CLIC1 siRNA could effectively suppress the expression of CLIC1 protein and mRNA.Proliferation of cells transfected with CLIC1 siRNA3 was enhanced notably,and the highest proliferation rate was 23.3%(P = 0.002) in SGC-7901 and 35.55%(P = 0.001) in MGC-803 cells at 48 h.The G2/M phase proportion increased,while G0/G1 and S phase proportions decreased.The apoptotic rate of the CLIC1 siRNA3 group obviously decreased in both SGC-7901 cells(62.24%,P = 0.000) and MGC-803 cells(52.67%,P = 0.004).Down-regulation of CLIC1 led to the inhibition of invasion and migration by 54.31%(P = 0.000) and 33.62%(P = 0.001) in SGC-7901 and 40.74%(P = 0.000) and 29.26%(P = 0.002) in MGC-803.However,there was no significant difference between the mock group cells and the negative control group cells.展开更多
基金funded by the Shenzhen Science and Technology Program,China(KQTD20180411143628272)the Special Funds for Science Technology Innovation and Industrial Development of Shenzhen Dapeng New District,China(pt202101-02)the National Key R&D Program of China(2022YFE0116500).
文摘Spodoptera frugiperda is a highly destructive pest that has become a global problem due to its robust reproductive and migratory capabilities.Transient receptor potential(TRP)channels,which constitute a vast ion channel family,play pivotal roles in sensing the external environment and maintaining internal homeostasis in insects.TRP channels have been widely investigated for their critical roles in regulating various insect behaviors in recent years.In this study,we identified 15 TRP gene loci encoding 26 transcripts in the genome of S.frugiperda and analyzed their expression profiles at different developmental stages.The results revealed that S.frugiperda possesses four TRPC genes,six TRPA genes,one TRPM gene,two TRPV genes,one TRPN gene,and one TRPML gene,while a canonical TRPP is absent.Moreover,the SfruTRPA1 was functionally characterized using the Xenopus oocyte expression system.The results showed that SfruTRPA1 is activated by temperature increases from 20 to 45℃,and there is no significant desensitization after repeated stimuli within the same temperature range.Additionally,SfruTRPA1 is activated by certain natural chemicals,including allyl isothiocyanate(AITC)and cinnamaldehyde(CA).These findings provide valuable insights to the TRP genes in S.frugiperda.
文摘OBJECTIVE Two-pore domain potassium channel subtype TREK-1 was widely proved to be activated by inhalational anesthet⁃ics such as chloroform,diethyl ether,halothane,and isoflurane.But little is known about whether TREK-1 was also a potentially important target of intravenous anesthetics.Etomidate is a popularly used intravenous anesthetic with good safety in clinic.The action of etomidate on TREK-1 was seldom reported.METHODS AND RESULTS By using patch-clamp whole-cell recording tech⁃niques,we found for the first time that etomidate could bidirectionally regulate the TREK-1 potassi⁃um channel in CHO/TREK-1 cells.TREK-1 current amplitudes were observed after the administra⁃tion of etomidate at concentrations ranging from 3 to 100μmol·L-1.Etomidate activated TREK-1 current at concentrations of 3,10,and 15μmol·L-1 with maximum activation at 10μmol·L-1.Interest⁃ingly,at higher concentrations from 20 to 100μmol·L-1,etomidate inhibited TREK-1 current in a concentration-dependent way.According to the concentration-response curve,the fitted criti⁃cal concentration of etomidate between TREK-1 activation and inhibition was 20.7μmol·L-1,which close to the result that etomidate had no obvious effect on TREK-1 at 20μmol·L-1.In addition,etomidate 10μmol·L-1 induced a significant mem⁃brane potential hyperpolarization while etomidate 30μmol·L-1 showed obvious membrane potential depolarization.Furthermore,the bidirectional regulation still existed when the extracellular pH of CHO/TREK-1 cells was decreased.CONCLUSION TREK-1 is activated by etomi⁃date at clinically relevant concentrations but inhib⁃ited by supraclinical concentrations of etomidate,which is different to other volatile anesthetics.TREK-1 might be a potential target for anesthetic such as etomidate and the complicated bidirec⁃tional regulation mechanism of etomidate needed to be fully studied in the future.
基金supported by grants from the National Natural Science Foundation of China (No.30971007)the Natural Science Foundation of Hubei Province for Outstanding Young Scholars (No.2010CDA103)the National Basic Research Program(No.2011CB504403)
文摘This study was aimed to examine the effect of TREK-1 silencing on the function of astrocytes. Three 21-nucleotide small interfering RNA (siRNA) duplexes (siT1, siT2, siT3) targeting TREK-1 were constructed. Cy3-labeled dsRNA oligmers were used to determine the transfection efficiency in cultured astrocytes. TREK-1-specific siRNA duplexes (siT1, siT2, siT3) at the optimal concentration were transfected into cultured astrocytes, and the most efficient siRNA was identified by the method of immunocytochemical staining and Western blotting. The proliferation of astrocytes tranfected with TREK-1-targeting siRNA under hypoxia condition was measured by fluorescence-activated cell sorting (FACS). The results showed that TREK-1 was expressed in cultured astrocytes. The dsRNA oligmers targeting TREK-1 could be transfected efficiently in cultured astrocytes and down-regulate the expression of TREK-1 in astrocytes. Moreover, the down-regulation of TREK-1 in astrocytes contributed to the proliferation of astrocytes under hypoxia condition as determined by cell cycle analysis. It was concluded that siRNA is a powerful technique that can be used to knockdown the expression of TREK-1 in astrocytes, which helps further investigate the function of TREK-1 channel in astrocytes under physicological and pathological condition.
基金Acknowledgments We thank Dr Emily Liman （University of Southern California, USA） for providing the pGEMHE vector for the Xenopus oocyte experiments. We also thank Dr Richer Gaber （Northwestern Uni- versity, USA） for providing the yeast mutant strain with K＋ transport deficiency. We are grateful to Dr Rainer Hedrich （University of Wurzburg, Germany） for critical discussion. This work was supported by the National Natural Science Foundation of China （grant no. 30830013 to WHW）, the Beijing Municipal Education Commission （grant no. YB20081001901 to WHW） and the Program of Introducing Talents of Discipline to Universities （grant no. B06003 to WHW）.
文摘Potassium transporters play crucial roles in K^＋ uptake and translocation in plants. However, so far little is known about the regulatory mechanism of potassium transporters. Here, we show that a Shaker-like potassium channel AtKC1, encoded by the AtLKT1 gene cloned from the Arabidopsis thaliana low-K^＋ （LK）-tolerant mutant Atlktl, significantly regulates AKTl-mediated K^＋ uptake under LK conditions. Under LK conditions, the Atkcl mutants maintained their root growth, whereas wild-type plants stopped their root growth. Lesion of AtKC1 significantly enhanced the tolerance of the Atkcl mutants to LK stress and markedly increased K^＋ uptake and K^＋ accumulation in the Atkclmutant roots under LK conditions. Electrophysiological results showed that AtKC1 inhibited the AKT1-mediated inward K^＋ currents and negatively shifted the voltage dependence of AKT1 channels. These results demonstrate that the ‘silent＇ K^＋ channel α-subunit AtKC1 negatively regulates the AKTl-mediated K^＋ uptake in Arabidopsis roots and consequently alters the ratio of root-to-shoot under LK stress conditions.
基金Supported by The "Eleventh Five-year Plan" for Medical Sci-ence Development of PLA,No.06MB243the National Natural Science Foundation of China,No.81101101 and No.51273165+1 种基金the Key Project of Chinese Ministry of Education,No.212149the Projects of Sichuan Province,No.2010SZ0294,No.2011JQ0032 and No.12ZB038
文摘AIM: To investigate the mechanisms of chloride intracellular channel 1 (CLIC1) in the metastasis of colon cancer under hypoxia-reoxygenation (H-R) conditions.
基金Supported by The National Natural Science Foundation of China,No.30560151the Key Research Project of Guangxi Municipal Health Bureau,No.200824+1 种基金the Research Project of Guangxi Educational Department,No.201012MS062 and No. 2011105981002M204the Natural Science Foundation of Guangxi,No.0832113
文摘AIM:To investigate the effect of chloride intracellular channel 1(CLIC1) on the cell proliferation,apoptosis,migration and invasion of gastric cancer cells.METHODS:CLIC1 expression was evaluated in human gastric cancer cell lines SGC-7901 and MGC-803 by real time polymerase chain reaction(RT-PCR).Four segments of small interference RNA(siRNA) targeting CLIC1 mRNA and a no-sense control segment were designed by bioinformatics technology.CLIC1 siRNA was selected using Lipofectamine 2000 and transfected transiently into human gastric cancer SGC-7901 and MGC-803 cells.The transfected efficiency was observed under fluorescence microscope.After transfection,mRNA expression of CLIC1 was detected with RT-PCR and Western blotting was used to detect the protein expression.Proliferation was examined by methyl thiazolyl tetrazolium and apoptosis was detected with flow cytometry.Polycarbonate membrane transwell chamber and Matrigel were used for the detection of the changes of invasion and migration of the two cell lines.RESULTS:In gastric cancer cell lines SGC-7901 and MGC-803,CLIC1 was obviously expressed and CLIC1 siRNA could effectively suppress the expression of CLIC1 protein and mRNA.Proliferation of cells transfected with CLIC1 siRNA3 was enhanced notably,and the highest proliferation rate was 23.3%(P = 0.002) in SGC-7901 and 35.55%(P = 0.001) in MGC-803 cells at 48 h.The G2/M phase proportion increased,while G0/G1 and S phase proportions decreased.The apoptotic rate of the CLIC1 siRNA3 group obviously decreased in both SGC-7901 cells(62.24%,P = 0.000) and MGC-803 cells(52.67%,P = 0.004).Down-regulation of CLIC1 led to the inhibition of invasion and migration by 54.31%(P = 0.000) and 33.62%(P = 0.001) in SGC-7901 and 40.74%(P = 0.000) and 29.26%(P = 0.002) in MGC-803.However,there was no significant difference between the mock group cells and the negative control group cells.
