Metabolic syndrome is a pre-diabetic state characterized by several biochemical and physiological alterations,including insulin resistance,visceral fat accumulation,and dyslipidemias,which increase the risk for develo...Metabolic syndrome is a pre-diabetic state characterized by several biochemical and physiological alterations,including insulin resistance,visceral fat accumulation,and dyslipidemias,which increase the risk for developing cardiovascular disease.Metabolic syndrome is associated with augmented sympathetic tone,which could account for the etiology of pre-diabetic cardiomyopathy.This review summarizes the current knowledge of the pathophysiological consequences of enhanced and sustainedβ-adrenergic response in pre-diabetes,focusing on cardiac dysfunction reported in diet-induced experimental models of pre-diabetic cardiomyopathy.The research reviewed indicates that both protein kinase A and Ca^(2+)/calmodulin-dependent protein kinase Ⅱ play important roles in functional responses mediated byβ1-adrenoceptors;therefore,alterations in the expression or function of these kinases can be deleterious.This review also outlines recent information on the role of protein kinase A and Ca^(2+)/calmodulin-dependent protein kinase Ⅱ in abnormal Ca^(2+)handling by cardiomyocytes from diet-induced models of pre-diabetic cardiomyopathy.展开更多
Many of the effects of Ca^2＋ signaling are mediated through the Ca^2＋/calmodulin complex and its acceptors, the Ca^2＋/calmodulin-dependent protein kinases, including PSKHI. Studies of the proteins involved in the c...Many of the effects of Ca^2＋ signaling are mediated through the Ca^2＋/calmodulin complex and its acceptors, the Ca^2＋/calmodulin-dependent protein kinases, including PSKHI. Studies of the proteins involved in the calcium metabolism in oysters will help elucidate the pearl formation mechanism. This paper describes a full-length PSKH1 cDNA isolated from pearl oyster Pinctada fucata. Oyster PSKH1 shares 65% homology with human PSKH1 and 48% similarity with rat CaM kinase I in the amino acid sequence, and contains a calmodulin-binding domain. The results of semi-quantitative reverse transcription-polymerase chain reaction and in situ hybridization revealed that oyster PSKH1 mRNA is highly expressed in the outer epithelial cells of the mantle pallial and in the gill epithelial cells. These studies provide important information describing the complex Ca^2＋ signaling mechanism in oyster calcium metabolism.展开更多
In rice, the Ca^(2+)/calmodulin-dependent protein kinase OsDMI3 is an important positive regulator of abscisic acid (ABA) signaling. In ABA signaling, H_(2)O_(2) is required for ABA-induced activation of OsDMI3, which...In rice, the Ca^(2+)/calmodulin-dependent protein kinase OsDMI3 is an important positive regulator of abscisic acid (ABA) signaling. In ABA signaling, H_(2)O_(2) is required for ABA-induced activation of OsDMI3, which in turn increase H_(2)O_(2) production. However, how OsDMI3 regulates H_(2)O_(2) production in ABA signaling remains unknown. Here we show that OsRbohB is the main NADPH oxidase involved in ABA-induced H_(2)O_(2) production and ABA-mediated physiological responses. OsDMI3 directly interacts with and phosphorylates OsRbohB at Ser-191, which is OsDMI3-mediated site-specific phosphorylation in ABA signaling. Further analyses revealed that OsDMI3-mediated OsRbohB Ser-191 phosphorylation positively regulates the activity of NADPH oxidase and the production of H_(2)O_(2) in ABA signaling, thereby enhancing the sensitivity of seed germination and root growth to ABA and plant tolerance to water stress and oxidative stress. Moreover, we discovered that the OsDMI3-mediated OsRbohB phosphorylation and H_(2)O_(2) production is dependent on the sucrose non-fermenting 1-related protein kinases SAPK8/9/10, which phosphorylate OsRbohB at Ser-140 in ABA signaling. Taken together, these results not only reveal an important regulatory mechanism that directly activates Rboh for ABA-induced H_(2)O_(2) production but also uncover the importance of this regulatory mechanism in ABA signaling.展开更多
Transcription factors can be used to engineer plants for enhanced productivity.However,the mechanism(s)by which the C2H2-type zinc fi nger transcription factor enhances pathogen resistance in cells is not fully unders...Transcription factors can be used to engineer plants for enhanced productivity.However,the mechanism(s)by which the C2H2-type zinc fi nger transcription factor enhances pathogen resistance in cells is not fully understood.Here,Agrobacterium tumefaciens carrying the gene for Arabidopsis thaliana cysteine2/histidine2-type transcription factor 6(ZAT6)was used to engineer rice(Oryza sativa L.),cotton(Gossypium hirsutum L.),and slash pine(Pinus elliottii Engelm.)to generate transgenic cell lines.Transgenic cells were then inoculated with the pathogenic bacterium Pseudomonas syringae.Compared to the control,cell viability of transgenic cells increased 39–47%and growth rate increased 9–15%by 7 days after inoculation in rice,cotton and slash pine.Acid phosphatase activity and alkaline phosphatase activity and transcript levels of Ca 2+-dependent protein kinase genes OsCPK1,OsCPK2,OsCPK6,and OsCPK8 and mitogen-activated protein kinase genes OsMAPK1,OsMAPK2,OsMAPK3,and OsMAPK8 increased signifi cantly in transgenic rice cells by 3 day after inoculation,and extracellular pH had decreased by 10–14%by 96 min after inoculation in transgenic rice,cotton and slash pine cells.These results suggest that ZAT6 enhances P.syringae resistance in plant cells by modulating transcription of CPK and MAPK and oxidase activity.展开更多
Manganese(Mn)is an essential micronutrient for all living organisms.However,excess Mn supply that can occur in acid or waterlogged soils has toxic effects on plant physiology and development.Although a variety of Mn t...Manganese(Mn)is an essential micronutrient for all living organisms.However,excess Mn supply that can occur in acid or waterlogged soils has toxic effects on plant physiology and development.Although a variety of Mn transporter families have been characterized,we have only a rudimentary understanding of how these transporters are regulated to uphold and adjust Mn homeostasis in plants.Here,we demonstrate that two calcineurin-B-like proteins,CBL2/3,and their interacting kinases,CIPK3/9/26,are key regulators of plant Mn homeostasis.Arabidopsis mutants lacking CBL2 and 3 or their interacting protein kinases CIPK3/9/26 exhibit remarkably high Mn tolerance.Intriguingly,CIPK3/9/26 interact with and phosphorylate the tonoplast-localized Mn and iron(Fe)transporter MTP8 primarily at Ser35,which is conserved among MTP8 proteins from various species.Mn transport complementation assays in yeast combined with multiple physiological assays indicate that CBL-CIPK-mediated phosphorylation of MTP8 negatively regulates its transport activity from the cytoplasm to the vacuole.Moreover,we show that sequential phosphorylation of MTP8,initially at Ser31/32 by the calcium-dependent protein kinase CPK5 and subsequently at Ser35 by CIPK26,provides an activation/deactivation fine-tuning mechanism for differential regulation of Mn transport.Collectively,our findings define a two-tiered calcium-controlled mechanism for dynamic regulation of Mn homeostasis under conditions of fluctuating Mn supply.展开更多
This brief article highlights the results of Fu et al.(Proc Natl Acad Sci USA 119:e2204574119,2022),who recently found that manganese(Mn)deficiency triggers long-lasting multicellular Ca^(2+) oscillations in the elong...This brief article highlights the results of Fu et al.(Proc Natl Acad Sci USA 119:e2204574119,2022),who recently found that manganese(Mn)deficiency triggers long-lasting multicellular Ca^(2+) oscillations in the elongation zone(EZ)of Arabidopsis roots and revealed a Ca^(2+)-CPK21/23-NRAMP1 axis as an important mechanism for plant tolerance and adaptation to low Mn.展开更多
AIM： To examine the existence of Nitric oxide/ cGMP sensitive store-operated Ca^2＋ entry in mouse fibroblast NIH/3T3 cells and its influence on matrix metalloproteinase （MMP） production and adhesion ability of fib...AIM： To examine the existence of Nitric oxide/ cGMP sensitive store-operated Ca^2＋ entry in mouse fibroblast NIH/3T3 cells and its influence on matrix metalloproteinase （MMP） production and adhesion ability of fibroblasts. METHODS： NIH/3T3 cells were cultured. Confocal laser scanning microscopy was used to examine the existence of thapsigargin-induced store-operated Ca^2＋ entry in fibroblasts. Gelatin zymography and semiquantitative reverse transcriptase-polymerase chain reaction （RTPCR） were employed to detect the involvement of [Ca^2＋]i and NO/cGMP in MMP secretion. The involvement of NO/ cGMP-sensitive Ca^2＋ entry in adhesion was determined using matrigel-coated culture plates. RESULTS： 8-bromo-cGMP inhibited the thapsigargin-induced Ca^2＋ entry in 3T3 cells. The cGMP-induced inhibition was abolished by an inhibitor of protein kinase G, KT5823 （1μmol/L）. A similar effect on the Ca^2＋ entry was observed in 3T3 cells in response to a NO donor, （±）-S-nitroso-N-acetylpenicillamine （SNAP）. The inhibitory effect of SNAP on the thapsigargin-induced Ca^2＋ entry was also observed, indicating NO/cGMP-regulated Ca^2＋ entry in 3T3 cells. Results of gelatin zymography assay showed that addition of extracellular Ca^2＋ concentration induced MMP release and activation in a dose-dependent manner. RT-PCR also showed that cGMP and SNAP reduced the production of MMP mRNA in 3T3 cells. Experiments investigating adhesion potentials demonstrated that cGMP and SNAP could upgrade 3T3 cell attachment rate to the matrigel-coated culture plates.CONCLUSION： NO/cGMP sensitive store-operated Ca^2＋ entry occurs in fibroblasts, and attenuates their adhesion potentials through its influence on MMP secretion.展开更多
AIM:To explore the effect and mechanism of gastrin and its antagonists prog lumide and somatostatin on colorectal carcinoma and their clinical significance.METHODS:A model of transplanted human colonic carcinoma was e...AIM:To explore the effect and mechanism of gastrin and its antagonists prog lumide and somatostatin on colorectal carcinoma and their clinical significance.METHODS:A model of transplanted human colonic carcinoma was established from SW480 cell line in gymnomouse body.The volume and weight of transplanted carcinoma was observed under the effect of pentagatrin (PG), proglumide (PGL) and octapeptide somotostatin (SMS201-995, SMS). The cAMP content of carcinoma cell was determined by radioimmunoassay and the DNA, protein content and cell cycle were determined by flow-cytometry. The amount of viable cells was determined by MTT colorimetric analysis,IP(3) content was determined by radioimmunoassay, Ca(2+) concentration in cell by fluorometry and PKC activity by isotopic enzymolysis. The expression of gastrin, c-myc, c-fos and rasP21 in 48 cases of colorectal carcinoma tissue was detected by the immuno-cytochemistry SP method. Argyrophilia nucleolar organizer regions was determined with argyrophilia stain.RESULTS:The volume,weight, cAMP, DNA and protein content in carcinoma cell, cell amount and proliferation index of S and G(2)M phase in PG group were all significantly higher than those of control group. When PG was at the concentration of 25mg/L, the amount of viable cells, IP(3) content and Ca(2+) concentration in cell and membrane PKC activity in PG group were significantly higher than those in control group; when PGL was at a concentration of 32mg/L, they dropped to the lowest level in PG (25mg/L)+PGL group, but without significant difference from the control group. The positive expression rate of gastrin, c-myc, c-fos and rasP21 in carcinoma tissue was 39.6%, 54.2%, 47.9% and 54.2% respectively and significantly higher than that in mucosa 3cm and 6cm adjacent to carcinoma tissue and normal colorectal mucosa. The positive expression rate of gastrin of highly differentiated adenocarcinoma group was significantly higher than that of poorly differentiated and mucinous adenocar-cinoma groups. The AgNORs count of carcinoma tissue was significantly higher than that in mucosa 3cm and 6cm adjacent to carcinoma tissue and normal colorectal mucosa; and the positive expression of c-myc and c-fos and the AgNORs count in gastrin-positive group was significantly higher than those in gastrin negative group.CONCLUSION:Pentagastrin has a promoting effect on the growth of transplanted human colonic carcinoma from SW480 cell line. PGL has no obvious effect on the growth of human colonic carcinoma SW480 cell line, but could inhibit the growth promoting effect of PG on transplanted carcinoma. Somatostatin can not only inhibit the growth of transplanted human colonic carcinoma from SW480 cell line directly but also depress the growth-promoting effect of gastrin on the transplanted carcinoma. Some colorectal carcinoma cells can produce and secrete gastrin through autocrine, highly differentiated adenocarcinoma express the highest level gastrin.Endogenous gastrin can stimulate the cell division and proliferation of carcinoma cell and promote the growth of colorectal carcinoma regulating the expression of oncogene c-myc, c-fos. Our study has provided experimental basis for the adjuvant treatment using gastrin antagonist such as PGL, somatostatin of patients with colorectal carcinoma.展开更多
Membrane depolarization induces the release of the serine proteinase tissue-type plasminogen activator（t PA） from the presynaptic terminal of cerebral cortical neurons.Once in the synaptic cleft this t PA promotes t...Membrane depolarization induces the release of the serine proteinase tissue-type plasminogen activator（t PA） from the presynaptic terminal of cerebral cortical neurons.Once in the synaptic cleft this t PA promotes the exocytosis and subsequent endocytic retrieval of glutamate-containing synaptic vesicles,and regulates the postsynaptic response to the presynaptic release of glutamate.Indeed,t PA has a bidirectional effect on the composition of the postsynaptic density（PSD） that does not require plasmin generation or the presynaptic release of glutamate,but varies according to the baseline level of neuronal activity.Hence,in inactive neurons t PA induces phosphorylation and accumulation in the PSD of the Ca^（2＋）/calmodulin-dependent protein kinase IIα（pCa MKIIα）,followed by pCa MKIIα-induced phosphorylation and synaptic recruitment of Glu R1-containing α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid（AMPA） receptors.In contrast,in active neurons with increased levels of pCa MKIIα in the PSD t PA induces pCa MKIIα and p Glu R1 dephosphorylation and their subsequent removal from the PSD.These effects require active synaptic N-methyl-D-aspartate（NMDA） receptors and cyclin-dependent kinase 5（Cdk5）-induced phosphorylation of the protein phosphatase 1（PP1） at T320.These data indicate that t PA is a homeostatic regulator of the postsynaptic response of cerebral cortical neurons to the presynaptic release of glutamate via bidirectional regulation of the pCa MKIIα/PP1 switch in the PSD.展开更多
Intramuscular fat （IMF） content in chickens significantly contributes to meat quality. The main objective of this study was to assess the expression of calcineudn （CAN） and Ca^2＋/calmodutin-dependent protein kina...Intramuscular fat （IMF） content in chickens significantly contributes to meat quality. The main objective of this study was to assess the expression of calcineudn （CAN） and Ca^2＋/calmodutin-dependent protein kinase （CAME） in lipogene- sis in chicken muscle. The chickens were slaughtered and sampled at the ages of 4, 8, and 16 weeks, respectively. IMF content and the expression of CaN subunits and CaMK isoforms were measured in thigh muscle tissue. The results showed that the IMF contents were higher in chickens at the age of 16 weeks compared with those in chickens at the ages of 4 and 8 weeks （P〈0.05）. The expression levels of fatty acid synthase （FAS） and fatty acid translocase CD36 （FAT/CD36） mRNA in 16-week-old chickens were all significantly up-regulated compared with those in 4-week-old chickens （P〈0.05）. The mRNA levels of CaNB and CaMK IV in 16-week-old chickens were significantly lower than those in 4-week-old chickens （P〈0.05）. But the CaMK II mRNA levels in 16-week-old chickens were significantly higher than those in 4-week-old chickens （P〈0.05）. To investigate the roles of CaMK and CaN in adipogenesis, SV cells were incubated in standard adipogenesis medium for 24 h and treated with specific inhibitor of CaMK and CaN. The ex- pressions of CCAAT/enhancer binding protein β（C/EBPJ3）, sterol regulatory element- binding protein 1 （SREBP1） and peroxisome proliferation-activated receptor ）, （PPARy） were dramatically enhanced by CsA and CaN inhibitor （P〈0.05）. KN93, a CaMK Ⅱ inhibitor, dramatically repressed the expression of those lipogenic genes （P〈0.05）. All the results above indicated that CaN and CaMK had different effects on adipogenesis in the muscle of chickens.展开更多
Background Gabapentin has been widely and successfully used in the clinic for many neuropathic pain syndromes since last decade, however its analgesic mechanisms are still elusive. Our study was to investigate whether...Background Gabapentin has been widely and successfully used in the clinic for many neuropathic pain syndromes since last decade, however its analgesic mechanisms are still elusive. Our study was to investigate whether Ca^2＋/calmodulin-dependent protein kinase Ⅱ （CaMKⅡ） contributes to the analgesic effect of gabapentin on a chronic constriction injury （CCI） model. Methods Gabapentin （2%, 100 mg/kg） or saline （0.5 ml/100 g） was injected intraperitoneally 15 minutes prior to surgery and then every 12 hours from postoperative day 0-4 to all rats in control, sham and CCI groups. The analgesic effect of gabapentin was assessed by measuring mechanical allodynia and thermal hyperalgesia of rats. Expression and activation of CaMKⅡ were quantified by reverse-transcriptional polymerase chain reaction and Western blotting. Results The analgesic effect of gabapentin on mechanical allodynia and thermal hyperalgesia was significant in the CCI model, with maximal reduction reached on postoperative day 8. Gabapentin decreased the expression of the total CaMKⅡ and phosphorylated CaMKⅡ in CCI rats. Conclusion The analgesic effect of gabapentin on CCI rats may be related to the decreased expression and phosphorylation of CaMKⅡ in the spinal cord.展开更多
G_(q)-coupled receptors regulate numerous physiological processes by activating enzymes and inducing intracellular Ca^(2+)signals.There is a strong need for an optogenetic tool that enables powerful experimental contr...G_(q)-coupled receptors regulate numerous physiological processes by activating enzymes and inducing intracellular Ca^(2+)signals.There is a strong need for an optogenetic tool that enables powerful experimental control over G_(q) signaling.Here,we present chicken opsin 5(cOpn5)as the long sought-after,single-component optogenetic tool that mediates ultra-sensitive optical control of intracellular G_(q) signaling with high temporal and spatial resolution.Expressing cOpn5 in HEK 293T cells and primary mouse astrocytes enables blue light-triggered,G_(q)-dependent Ca^(2+) release from intracellular stores and protein kinase C activation.Strong Ca^(2+) transients were evoked by brief light pulses of merely 10 ms duration and at 3 orders lower light intensity of that for common optogenetic tools.Photostimulation of cOpn5-expressing cells at the subcellular and single-cell levels generated fast intracellular Ca^(2+)transition,thus demonstrating the high spatial precision of cOpn5 optogenetics.The cOpn5-mediated optogenetics could also be applied to activate neurons and control animal behavior in a circuit-dependent manner.cOpn5 optogenetics may find broad applications in studying the mechanisms and functional relevance of G_(q) signaling in both non-excitable cells and excitable cells in all major organ systems.展开更多
文摘Metabolic syndrome is a pre-diabetic state characterized by several biochemical and physiological alterations,including insulin resistance,visceral fat accumulation,and dyslipidemias,which increase the risk for developing cardiovascular disease.Metabolic syndrome is associated with augmented sympathetic tone,which could account for the etiology of pre-diabetic cardiomyopathy.This review summarizes the current knowledge of the pathophysiological consequences of enhanced and sustainedβ-adrenergic response in pre-diabetes,focusing on cardiac dysfunction reported in diet-induced experimental models of pre-diabetic cardiomyopathy.The research reviewed indicates that both protein kinase A and Ca^(2+)/calmodulin-dependent protein kinase Ⅱ play important roles in functional responses mediated byβ1-adrenoceptors;therefore,alterations in the expression or function of these kinases can be deleterious.This review also outlines recent information on the role of protein kinase A and Ca^(2+)/calmodulin-dependent protein kinase Ⅱ in abnormal Ca^(2+)handling by cardiomyocytes from diet-induced models of pre-diabetic cardiomyopathy.
基金Supported by the National High-Tech Research and Development (863) Program of China (No. 2003AA603430) and the National Natural Science Foundation of China (No. 30371092)
文摘Many of the effects of Ca^2＋ signaling are mediated through the Ca^2＋/calmodulin complex and its acceptors, the Ca^2＋/calmodulin-dependent protein kinases, including PSKHI. Studies of the proteins involved in the calcium metabolism in oysters will help elucidate the pearl formation mechanism. This paper describes a full-length PSKH1 cDNA isolated from pearl oyster Pinctada fucata. Oyster PSKH1 shares 65% homology with human PSKH1 and 48% similarity with rat CaM kinase I in the amino acid sequence, and contains a calmodulin-binding domain. The results of semi-quantitative reverse transcription-polymerase chain reaction and in situ hybridization revealed that oyster PSKH1 mRNA is highly expressed in the outer epithelial cells of the mantle pallial and in the gill epithelial cells. These studies provide important information describing the complex Ca^2＋ signaling mechanism in oyster calcium metabolism.
基金supported by the National Natural Science Foundation of China(31971824 and 32170316).
文摘In rice, the Ca^(2+)/calmodulin-dependent protein kinase OsDMI3 is an important positive regulator of abscisic acid (ABA) signaling. In ABA signaling, H_(2)O_(2) is required for ABA-induced activation of OsDMI3, which in turn increase H_(2)O_(2) production. However, how OsDMI3 regulates H_(2)O_(2) production in ABA signaling remains unknown. Here we show that OsRbohB is the main NADPH oxidase involved in ABA-induced H_(2)O_(2) production and ABA-mediated physiological responses. OsDMI3 directly interacts with and phosphorylates OsRbohB at Ser-191, which is OsDMI3-mediated site-specific phosphorylation in ABA signaling. Further analyses revealed that OsDMI3-mediated OsRbohB Ser-191 phosphorylation positively regulates the activity of NADPH oxidase and the production of H_(2)O_(2) in ABA signaling, thereby enhancing the sensitivity of seed germination and root growth to ABA and plant tolerance to water stress and oxidative stress. Moreover, we discovered that the OsDMI3-mediated OsRbohB phosphorylation and H_(2)O_(2) production is dependent on the sucrose non-fermenting 1-related protein kinases SAPK8/9/10, which phosphorylate OsRbohB at Ser-140 in ABA signaling. Taken together, these results not only reveal an important regulatory mechanism that directly activates Rboh for ABA-induced H_(2)O_(2) production but also uncover the importance of this regulatory mechanism in ABA signaling.
文摘Transcription factors can be used to engineer plants for enhanced productivity.However,the mechanism(s)by which the C2H2-type zinc fi nger transcription factor enhances pathogen resistance in cells is not fully understood.Here,Agrobacterium tumefaciens carrying the gene for Arabidopsis thaliana cysteine2/histidine2-type transcription factor 6(ZAT6)was used to engineer rice(Oryza sativa L.),cotton(Gossypium hirsutum L.),and slash pine(Pinus elliottii Engelm.)to generate transgenic cell lines.Transgenic cells were then inoculated with the pathogenic bacterium Pseudomonas syringae.Compared to the control,cell viability of transgenic cells increased 39–47%and growth rate increased 9–15%by 7 days after inoculation in rice,cotton and slash pine.Acid phosphatase activity and alkaline phosphatase activity and transcript levels of Ca 2+-dependent protein kinase genes OsCPK1,OsCPK2,OsCPK6,and OsCPK8 and mitogen-activated protein kinase genes OsMAPK1,OsMAPK2,OsMAPK3,and OsMAPK8 increased signifi cantly in transgenic rice cells by 3 day after inoculation,and extracellular pH had decreased by 10–14%by 96 min after inoculation in transgenic rice,cotton and slash pine cells.These results suggest that ZAT6 enhances P.syringae resistance in plant cells by modulating transcription of CPK and MAPK and oxidase activity.
基金This research was funded by a grant from the National Natural Science Foundation of China(31770289 to C.W.)Northwest A&F University(Z111021604 to C.W.)+3 种基金the National Natural Science Foundation of China(31900236 to Z.Z.)supported in part by the open funds of China Postdoctoral Science Foundation(2018M643740 to Z.Z.)the Natural Science Basic Research Plan in Shaanxi Province of China(program no.2019JQ-150)the State Key Laboratory of Plant Physiology and Biochemistry(SKLPPBKF2101 to C.W.).
文摘Manganese(Mn)is an essential micronutrient for all living organisms.However,excess Mn supply that can occur in acid or waterlogged soils has toxic effects on plant physiology and development.Although a variety of Mn transporter families have been characterized,we have only a rudimentary understanding of how these transporters are regulated to uphold and adjust Mn homeostasis in plants.Here,we demonstrate that two calcineurin-B-like proteins,CBL2/3,and their interacting kinases,CIPK3/9/26,are key regulators of plant Mn homeostasis.Arabidopsis mutants lacking CBL2 and 3 or their interacting protein kinases CIPK3/9/26 exhibit remarkably high Mn tolerance.Intriguingly,CIPK3/9/26 interact with and phosphorylate the tonoplast-localized Mn and iron(Fe)transporter MTP8 primarily at Ser35,which is conserved among MTP8 proteins from various species.Mn transport complementation assays in yeast combined with multiple physiological assays indicate that CBL-CIPK-mediated phosphorylation of MTP8 negatively regulates its transport activity from the cytoplasm to the vacuole.Moreover,we show that sequential phosphorylation of MTP8,initially at Ser31/32 by the calcium-dependent protein kinase CPK5 and subsequently at Ser35 by CIPK26,provides an activation/deactivation fine-tuning mechanism for differential regulation of Mn transport.Collectively,our findings define a two-tiered calcium-controlled mechanism for dynamic regulation of Mn homeostasis under conditions of fluctuating Mn supply.
基金supported by grants from National Natural Science Foundation of China(Grant No.31900216)National Key Laboratory of Plant Molecular Genetics.
文摘This brief article highlights the results of Fu et al.(Proc Natl Acad Sci USA 119:e2204574119,2022),who recently found that manganese(Mn)deficiency triggers long-lasting multicellular Ca^(2+) oscillations in the elongation zone(EZ)of Arabidopsis roots and revealed a Ca^(2+)-CPK21/23-NRAMP1 axis as an important mechanism for plant tolerance and adaptation to low Mn.
基金Supported by the Major State Basic Research Development Program (973 Program) of China, No.2003CB515507
文摘AIM： To examine the existence of Nitric oxide/ cGMP sensitive store-operated Ca^2＋ entry in mouse fibroblast NIH/3T3 cells and its influence on matrix metalloproteinase （MMP） production and adhesion ability of fibroblasts. METHODS： NIH/3T3 cells were cultured. Confocal laser scanning microscopy was used to examine the existence of thapsigargin-induced store-operated Ca^2＋ entry in fibroblasts. Gelatin zymography and semiquantitative reverse transcriptase-polymerase chain reaction （RTPCR） were employed to detect the involvement of [Ca^2＋]i and NO/cGMP in MMP secretion. The involvement of NO/ cGMP-sensitive Ca^2＋ entry in adhesion was determined using matrigel-coated culture plates. RESULTS： 8-bromo-cGMP inhibited the thapsigargin-induced Ca^2＋ entry in 3T3 cells. The cGMP-induced inhibition was abolished by an inhibitor of protein kinase G, KT5823 （1μmol/L）. A similar effect on the Ca^2＋ entry was observed in 3T3 cells in response to a NO donor, （±）-S-nitroso-N-acetylpenicillamine （SNAP）. The inhibitory effect of SNAP on the thapsigargin-induced Ca^2＋ entry was also observed, indicating NO/cGMP-regulated Ca^2＋ entry in 3T3 cells. Results of gelatin zymography assay showed that addition of extracellular Ca^2＋ concentration induced MMP release and activation in a dose-dependent manner. RT-PCR also showed that cGMP and SNAP reduced the production of MMP mRNA in 3T3 cells. Experiments investigating adhesion potentials demonstrated that cGMP and SNAP could upgrade 3T3 cell attachment rate to the matrigel-coated culture plates.CONCLUSION： NO/cGMP sensitive store-operated Ca^2＋ entry occurs in fibroblasts, and attenuates their adhesion potentials through its influence on MMP secretion.
文摘AIM:To explore the effect and mechanism of gastrin and its antagonists prog lumide and somatostatin on colorectal carcinoma and their clinical significance.METHODS:A model of transplanted human colonic carcinoma was established from SW480 cell line in gymnomouse body.The volume and weight of transplanted carcinoma was observed under the effect of pentagatrin (PG), proglumide (PGL) and octapeptide somotostatin (SMS201-995, SMS). The cAMP content of carcinoma cell was determined by radioimmunoassay and the DNA, protein content and cell cycle were determined by flow-cytometry. The amount of viable cells was determined by MTT colorimetric analysis,IP(3) content was determined by radioimmunoassay, Ca(2+) concentration in cell by fluorometry and PKC activity by isotopic enzymolysis. The expression of gastrin, c-myc, c-fos and rasP21 in 48 cases of colorectal carcinoma tissue was detected by the immuno-cytochemistry SP method. Argyrophilia nucleolar organizer regions was determined with argyrophilia stain.RESULTS:The volume,weight, cAMP, DNA and protein content in carcinoma cell, cell amount and proliferation index of S and G(2)M phase in PG group were all significantly higher than those of control group. When PG was at the concentration of 25mg/L, the amount of viable cells, IP(3) content and Ca(2+) concentration in cell and membrane PKC activity in PG group were significantly higher than those in control group; when PGL was at a concentration of 32mg/L, they dropped to the lowest level in PG (25mg/L)+PGL group, but without significant difference from the control group. The positive expression rate of gastrin, c-myc, c-fos and rasP21 in carcinoma tissue was 39.6%, 54.2%, 47.9% and 54.2% respectively and significantly higher than that in mucosa 3cm and 6cm adjacent to carcinoma tissue and normal colorectal mucosa. The positive expression rate of gastrin of highly differentiated adenocarcinoma group was significantly higher than that of poorly differentiated and mucinous adenocar-cinoma groups. The AgNORs count of carcinoma tissue was significantly higher than that in mucosa 3cm and 6cm adjacent to carcinoma tissue and normal colorectal mucosa; and the positive expression of c-myc and c-fos and the AgNORs count in gastrin-positive group was significantly higher than those in gastrin negative group.CONCLUSION:Pentagastrin has a promoting effect on the growth of transplanted human colonic carcinoma from SW480 cell line. PGL has no obvious effect on the growth of human colonic carcinoma SW480 cell line, but could inhibit the growth promoting effect of PG on transplanted carcinoma. Somatostatin can not only inhibit the growth of transplanted human colonic carcinoma from SW480 cell line directly but also depress the growth-promoting effect of gastrin on the transplanted carcinoma. Some colorectal carcinoma cells can produce and secrete gastrin through autocrine, highly differentiated adenocarcinoma express the highest level gastrin.Endogenous gastrin can stimulate the cell division and proliferation of carcinoma cell and promote the growth of colorectal carcinoma regulating the expression of oncogene c-myc, c-fos. Our study has provided experimental basis for the adjuvant treatment using gastrin antagonist such as PGL, somatostatin of patients with colorectal carcinoma.
基金supported in part by National Institutes of Health Grants NS-079331(to MY)and NS-091201(to MY)
文摘Membrane depolarization induces the release of the serine proteinase tissue-type plasminogen activator（t PA） from the presynaptic terminal of cerebral cortical neurons.Once in the synaptic cleft this t PA promotes the exocytosis and subsequent endocytic retrieval of glutamate-containing synaptic vesicles,and regulates the postsynaptic response to the presynaptic release of glutamate.Indeed,t PA has a bidirectional effect on the composition of the postsynaptic density（PSD） that does not require plasmin generation or the presynaptic release of glutamate,but varies according to the baseline level of neuronal activity.Hence,in inactive neurons t PA induces phosphorylation and accumulation in the PSD of the Ca^（2＋）/calmodulin-dependent protein kinase IIα（pCa MKIIα）,followed by pCa MKIIα-induced phosphorylation and synaptic recruitment of Glu R1-containing α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid（AMPA） receptors.In contrast,in active neurons with increased levels of pCa MKIIα in the PSD t PA induces pCa MKIIα and p Glu R1 dephosphorylation and their subsequent removal from the PSD.These effects require active synaptic N-methyl-D-aspartate（NMDA） receptors and cyclin-dependent kinase 5（Cdk5）-induced phosphorylation of the protein phosphatase 1（PP1） at T320.These data indicate that t PA is a homeostatic regulator of the postsynaptic response of cerebral cortical neurons to the presynaptic release of glutamate via bidirectional regulation of the pCa MKIIα/PP1 switch in the PSD.
基金Supported by Natural Science Foundation of Hubei Province of China(2011CDB012)Project of State Key Laboratory of Animal Nutrition(2004DA125184F1012)
文摘Intramuscular fat （IMF） content in chickens significantly contributes to meat quality. The main objective of this study was to assess the expression of calcineudn （CAN） and Ca^2＋/calmodutin-dependent protein kinase （CAME） in lipogene- sis in chicken muscle. The chickens were slaughtered and sampled at the ages of 4, 8, and 16 weeks, respectively. IMF content and the expression of CaN subunits and CaMK isoforms were measured in thigh muscle tissue. The results showed that the IMF contents were higher in chickens at the age of 16 weeks compared with those in chickens at the ages of 4 and 8 weeks （P〈0.05）. The expression levels of fatty acid synthase （FAS） and fatty acid translocase CD36 （FAT/CD36） mRNA in 16-week-old chickens were all significantly up-regulated compared with those in 4-week-old chickens （P〈0.05）. The mRNA levels of CaNB and CaMK IV in 16-week-old chickens were significantly lower than those in 4-week-old chickens （P〈0.05）. But the CaMK II mRNA levels in 16-week-old chickens were significantly higher than those in 4-week-old chickens （P〈0.05）. To investigate the roles of CaMK and CaN in adipogenesis, SV cells were incubated in standard adipogenesis medium for 24 h and treated with specific inhibitor of CaMK and CaN. The ex- pressions of CCAAT/enhancer binding protein β（C/EBPJ3）, sterol regulatory element- binding protein 1 （SREBP1） and peroxisome proliferation-activated receptor ）, （PPARy） were dramatically enhanced by CsA and CaN inhibitor （P〈0.05）. KN93, a CaMK Ⅱ inhibitor, dramatically repressed the expression of those lipogenic genes （P〈0.05）. All the results above indicated that CaN and CaMK had different effects on adipogenesis in the muscle of chickens.
基金This research is supported by a grant from the National Natural Science Foundation of China （No. 30672029）.
文摘Background Gabapentin has been widely and successfully used in the clinic for many neuropathic pain syndromes since last decade, however its analgesic mechanisms are still elusive. Our study was to investigate whether Ca^2＋/calmodulin-dependent protein kinase Ⅱ （CaMKⅡ） contributes to the analgesic effect of gabapentin on a chronic constriction injury （CCI） model. Methods Gabapentin （2%, 100 mg/kg） or saline （0.5 ml/100 g） was injected intraperitoneally 15 minutes prior to surgery and then every 12 hours from postoperative day 0-4 to all rats in control, sham and CCI groups. The analgesic effect of gabapentin was assessed by measuring mechanical allodynia and thermal hyperalgesia of rats. Expression and activation of CaMKⅡ were quantified by reverse-transcriptional polymerase chain reaction and Western blotting. Results The analgesic effect of gabapentin on mechanical allodynia and thermal hyperalgesia was significant in the CCI model, with maximal reduction reached on postoperative day 8. Gabapentin decreased the expression of the total CaMKⅡ and phosphorylated CaMKⅡ in CCI rats. Conclusion The analgesic effect of gabapentin on CCI rats may be related to the decreased expression and phosphorylation of CaMKⅡ in the spinal cord.
基金supported by Ministry of Science and Technology China Brain Initiative Grant(2021ZD0202803)the Research Unit of Medical Neurobiology at Chinese Academy of Medical Sciences(2019RU003)Beijing Municipal Government。
文摘G_(q)-coupled receptors regulate numerous physiological processes by activating enzymes and inducing intracellular Ca^(2+)signals.There is a strong need for an optogenetic tool that enables powerful experimental control over G_(q) signaling.Here,we present chicken opsin 5(cOpn5)as the long sought-after,single-component optogenetic tool that mediates ultra-sensitive optical control of intracellular G_(q) signaling with high temporal and spatial resolution.Expressing cOpn5 in HEK 293T cells and primary mouse astrocytes enables blue light-triggered,G_(q)-dependent Ca^(2+) release from intracellular stores and protein kinase C activation.Strong Ca^(2+) transients were evoked by brief light pulses of merely 10 ms duration and at 3 orders lower light intensity of that for common optogenetic tools.Photostimulation of cOpn5-expressing cells at the subcellular and single-cell levels generated fast intracellular Ca^(2+)transition,thus demonstrating the high spatial precision of cOpn5 optogenetics.The cOpn5-mediated optogenetics could also be applied to activate neurons and control animal behavior in a circuit-dependent manner.cOpn5 optogenetics may find broad applications in studying the mechanisms and functional relevance of G_(q) signaling in both non-excitable cells and excitable cells in all major organ systems.
