Calcium/calmodulin-dependent protein kinase that operates in the calcium-triggered CaMKK-CaMK1 signaling cascade and, upon calcium influx, activates CREB-dependent gene transcription, regulates calcium-mediated granulocyte function and respiratory burst and promotes basal dendritic growth of hippocampal neurons. In neutrophil cells, required for cytokine-induced proliferative responses and activation of the respiratory burst. Activates the transcription factor CREB1 in hippocampal neuron nuclei. May play a role in apoptosis of erythroleukemia cells. In vitro, phosphorylates transcription factor CREM isoform Beta.
Activation of granulocyte effector functions, such as induction of the respiratory burst and migration, are regulated by a variety of relatively ill-defined signaling pathways. Recently, we identified a novel Ca2+/calmodulin-dependent kinase I-like kinase, CKLiK, which exhibits restricted mRNA expression to human granulocytes. Using a novel antibody generated against the C-terminus of CKLiK, CKLiK was detected in CD34+-derived neutrophils and eosinophils, as well as in mature peripheral blood granulocytes. Activation of human granulocytes by N-formyl-methionyl-leucyl-phenylalanine (fMLP) and platelet-activating factor (PAF), but not the phorbol ester PMA (phorbol 12-myristate-13-acetate), resulted in induction of CKLiK activity, in parallel with a rise of intracellular Ca2+ [Ca2+]i. To study the functionality of CKLiK in human granulocytes, a cell-permeable CKLiK peptide inhibitor (CKLiK297-321) was generated which was able to inhibit kinase activity in a dose-dependent manner. The effect of this peptide was studied on specific granulocyte effector functions such as phagocytosis, respiratory burst, migration, and adhesion. Phagocytosis of Aspergillus fumigatus particles was reduced in the presence of CKLiK297-321 and fMLP-induced reactive oxygen species (ROS) production was potently inhibited by CKLiK297-321 in a dose-dependent manner. Furthermore, fMLP-induced neutrophil migration on albumin-coated surfaces was perturbed, as well as beta2-integrin-mediated adhesion. These findings suggest a critical role for CKLiK in modulating chemoattractant-induced functional responses in human granulocytes.
Among multifunctional Ca(2+)/calmodulin-dependent protein kinases (CaMKs), CaMKI has been shown to comprise a family of four structurally related isoforms (alpha, beta, gamma, and delta) encoded by separate genes with abundant expression in mature brain. In this study, we first examined the developmental gene expression of the four isoforms of CaMKI in mouse brain with special attention to the hippocampal formation by in situ hybridization analysis. The four isoforms of CaMKI were found to exhibit distinct spatiotemporal expression during neuronal development. We also examined the functional involvement of CaMKI in the dendritic formation of cultured hippocampal neurons. The overexpression of kinase-dead mutants of CaMKI reduced the average dendritic length of the transfected neurons without any significant effects on the number of primary dendrites and the branching index. Our present findings provide the detailed anatomical information on the developmental expression of the four isoforms of CaMKI in mouse brain, which represents the possible functional involvement of CaMKI in the basal dendritic growth of hippocampal neurons.
Multifunctional Ca2+/calmodulin-dependent protein kinases (CaMKs) including CaMKI, II and IV, are thought to regulate a variety of neuronal functions. Unlike CaMKII, which is regulated by autophosphorylation, CaMKI as well as CaMKIV are activated by CaMKK. In this study, we examined the cellular and subcellular localization of CaMKIdelta, a recently identified fourth isoform of CaMKI, in the mature brain. In situ hybridization analysis demonstrated wide expression of CaMKIdelta mRNA in the adult mouse brain with prominent expression in the hippocampal pyramidal cells. FLAG-tagged CaMKIdelta was localized at the cytoplasm and neurites without nuclear immunoreactivity in approximately 80% of the transfected primary hippocampal neurons. The stimulation with either KCl depolarization or glutamate triggered the nuclear localization of FLAG-tagged CaMKIdelta by two-fold with a peak at 1 min. In contrast, the catalytically inactive mutants of CaMKIdelta remained cytoplasmic without nuclear translocation during KCl depolarization, indicating the requirement of its activation for the nuclear translocation. Furthermore, we showed that immunoprecipitated CaMKIdelta could phosphorylate cAMP response element binding protein (CREB)alphain vitro and that the over-expression of CaMKIdelta enhanced GAL4-CREB-luciferase activity in PC12 cells stimulated by KCl depolarization. Our present study provides the first evidence for the possible involvement of CaMKIdelta in nuclear functions through its nuclear translocation in response to stimuli that trigger intracellular Ca2+ influx.
Human granulocytes are characterized by a variety of specific effector functions involved in host defense. Several widely expressed protein kinases have been implicated in the regulation of these effector functions. A polymerase chain reaction-based strategy was used to identify novel granulocyte-specific kinases. A novel protein kinase complementary DNA with an open reading frame of 357 amino acids was identified with homology to calcium-calmodulin-dependent kinase I (CaMKI). This has been termed CaMKI-like kinase (CKLiK). Analysis of CKLiK messenger RNA (mRNA) expression in hematopoietic cells demonstrated an almost exclusive expression in human polymorphonuclear leukocytes (PMN). Up-regulation of CKLiK mRNA occurs during neutrophilic differentiation of CD34(+) stem cells. CKLiK kinase activity was dependent on Ca(++) and calmodulin as analyzed by in vitro phosphorylation of cyclic adenosine monophosphate responsive element modulator (CREM). Furthermore, CKLiK- transfected cells treated with ionomycin demonstrated an induction of CRE- binding protein (CREB) transcriptional activity compared to control cells. Additionally, CaMK-kinasealpha enhanced CKLiK activity. In vivo activation of CKLiK was shown by addition of interleukin (IL)-8 to a myeloid cell line stably expressing CKLiK. Furthermore inducible activation of CKLiK was sufficient to induce extracellular signal-related kinase (ERK) mitogen-activated protein (MAP) kinase activity. These data identify a novel Ca(++)/calmodulin-dependent PMN- specific kinase that may play a role in Ca(++)-mediated regulation of human granulocyte functions.
Interacting selectively and non-covalently with calmodulin, a calcium-binding protein with many roles, both in the calcium-bound and calcium-free states.
Catalysis of the reactions: ATP + a protein serine = ADP + protein serine phosphate; and ATP + a protein threonine = ADP + protein threonine phosphate. These reactions require the presence of calcium-bound calmodulin.
The immediate defensive reaction (by vertebrate tissue) to infection or injury caused by chemical or physical agents. The process is characterized by local vasodilation, extravasation of plasma into intercellular spaces and accumulation of white blood cells and macrophages.
Multifunctional Ca2+/calmodulin-dependent protein kinases (CaMKs) including CaMKI, II and IV, are thought to regulate a variety of neuronal functions. Unlike CaMKII, which is regulated by autophosphorylation, CaMKI as well as CaMKIV are activated by CaMKK. In this study, we examined the cellular and subcellular localization of CaMKIdelta, a recently identified fourth isoform of CaMKI, in the mature brain. In situ hybridization analysis demonstrated wide expression of CaMKIdelta mRNA in the adult mouse brain with prominent expression in the hippocampal pyramidal cells. FLAG-tagged CaMKIdelta was localized at the cytoplasm and neurites without nuclear immunoreactivity in approximately 80% of the transfected primary hippocampal neurons. The stimulation with either KCl depolarization or glutamate triggered the nuclear localization of FLAG-tagged CaMKIdelta by two-fold with a peak at 1 min. In contrast, the catalytically inactive mutants of CaMKIdelta remained cytoplasmic without nuclear translocation during KCl depolarization, indicating the requirement of its activation for the nuclear translocation. Furthermore, we showed that immunoprecipitated CaMKIdelta could phosphorylate cAMP response element binding protein (CREB)alphain vitro and that the over-expression of CaMKIdelta enhanced GAL4-CREB-luciferase activity in PC12 cells stimulated by KCl depolarization. Our present study provides the first evidence for the possible involvement of CaMKIdelta in nuclear functions through its nuclear translocation in response to stimuli that trigger intracellular Ca2+ influx.
Any process that increases the rate, frequency or extent of neuron projection development. Neuron projection development is the process whose specific outcome is the progression of a neuron projection over time, from its formation to the mature structure. A neuron projection is any process extending from a neural cell, such as axons or dendrites (collectively called neurites).
Any process that increases the frequency, rate, or extent of neutrophil chemotaxis. Neutrophil chemotaxis is the directed movement of a neutrophil cell, the most numerous polymorphonuclear leukocyte found in the blood, in response to an external stimulus, usually an infection or wounding.
Activation of granulocyte effector functions, such as induction of the respiratory burst and migration, are regulated by a variety of relatively ill-defined signaling pathways. Recently, we identified a novel Ca2+/calmodulin-dependent kinase I-like kinase, CKLiK, which exhibits restricted mRNA expression to human granulocytes. Using a novel antibody generated against the C-terminus of CKLiK, CKLiK was detected in CD34+-derived neutrophils and eosinophils, as well as in mature peripheral blood granulocytes. Activation of human granulocytes by N-formyl-methionyl-leucyl-phenylalanine (fMLP) and platelet-activating factor (PAF), but not the phorbol ester PMA (phorbol 12-myristate-13-acetate), resulted in induction of CKLiK activity, in parallel with a rise of intracellular Ca2+ [Ca2+]i. To study the functionality of CKLiK in human granulocytes, a cell-permeable CKLiK peptide inhibitor (CKLiK297-321) was generated which was able to inhibit kinase activity in a dose-dependent manner. The effect of this peptide was studied on specific granulocyte effector functions such as phagocytosis, respiratory burst, migration, and adhesion. Phagocytosis of Aspergillus fumigatus particles was reduced in the presence of CKLiK297-321 and fMLP-induced reactive oxygen species (ROS) production was potently inhibited by CKLiK297-321 in a dose-dependent manner. Furthermore, fMLP-induced neutrophil migration on albumin-coated surfaces was perturbed, as well as beta2-integrin-mediated adhesion. These findings suggest a critical role for CKLiK in modulating chemoattractant-induced functional responses in human granulocytes.
Any process that increases the rate frequency or extent of a phase of elevated metabolic activity, during which oxygen consumption increases; this leads to the production, by an NADH dependent system, of hydrogen peroxide (H2O2), superoxide anions and hydroxyl radicals.
Evidence
1:
Inferred from Mutant PhenotypeUniProtKB
Activation of granulocyte effector functions, such as induction of the respiratory burst and migration, are regulated by a variety of relatively ill-defined signaling pathways. Recently, we identified a novel Ca2+/calmodulin-dependent kinase I-like kinase, CKLiK, which exhibits restricted mRNA expression to human granulocytes. Using a novel antibody generated against the C-terminus of CKLiK, CKLiK was detected in CD34+-derived neutrophils and eosinophils, as well as in mature peripheral blood granulocytes. Activation of human granulocytes by N-formyl-methionyl-leucyl-phenylalanine (fMLP) and platelet-activating factor (PAF), but not the phorbol ester PMA (phorbol 12-myristate-13-acetate), resulted in induction of CKLiK activity, in parallel with a rise of intracellular Ca2+ [Ca2+]i. To study the functionality of CKLiK in human granulocytes, a cell-permeable CKLiK peptide inhibitor (CKLiK297-321) was generated which was able to inhibit kinase activity in a dose-dependent manner. The effect of this peptide was studied on specific granulocyte effector functions such as phagocytosis, respiratory burst, migration, and adhesion. Phagocytosis of Aspergillus fumigatus particles was reduced in the presence of CKLiK297-321 and fMLP-induced reactive oxygen species (ROS) production was potently inhibited by CKLiK297-321 in a dose-dependent manner. Furthermore, fMLP-induced neutrophil migration on albumin-coated surfaces was perturbed, as well as beta2-integrin-mediated adhesion. These findings suggest a critical role for CKLiK in modulating chemoattractant-induced functional responses in human granulocytes.
Among multifunctional Ca(2+)/calmodulin-dependent protein kinases (CaMKs), CaMKI has been shown to comprise a family of four structurally related isoforms (alpha, beta, gamma, and delta) encoded by separate genes with abundant expression in mature brain. In this study, we first examined the developmental gene expression of the four isoforms of CaMKI in mouse brain with special attention to the hippocampal formation by in situ hybridization analysis. The four isoforms of CaMKI were found to exhibit distinct spatiotemporal expression during neuronal development. We also examined the functional involvement of CaMKI in the dendritic formation of cultured hippocampal neurons. The overexpression of kinase-dead mutants of CaMKI reduced the average dendritic length of the transfected neurons without any significant effects on the number of primary dendrites and the branching index. Our present findings provide the detailed anatomical information on the developmental expression of the four isoforms of CaMKI in mouse brain, which represents the possible functional involvement of CaMKI in the basal dendritic growth of hippocampal neurons.
Any process that modulates the rate, frequency or extent of granulocyte chemotaxis. Granulocyte chemotaxis is the movement of a granulocyte in response to an external stimulus.
Evidence
1:
Inferred from Mutant PhenotypeUniProtKB
Activation of granulocyte effector functions, such as induction of the respiratory burst and migration, are regulated by a variety of relatively ill-defined signaling pathways. Recently, we identified a novel Ca2+/calmodulin-dependent kinase I-like kinase, CKLiK, which exhibits restricted mRNA expression to human granulocytes. Using a novel antibody generated against the C-terminus of CKLiK, CKLiK was detected in CD34+-derived neutrophils and eosinophils, as well as in mature peripheral blood granulocytes. Activation of human granulocytes by N-formyl-methionyl-leucyl-phenylalanine (fMLP) and platelet-activating factor (PAF), but not the phorbol ester PMA (phorbol 12-myristate-13-acetate), resulted in induction of CKLiK activity, in parallel with a rise of intracellular Ca2+ [Ca2+]i. To study the functionality of CKLiK in human granulocytes, a cell-permeable CKLiK peptide inhibitor (CKLiK297-321) was generated which was able to inhibit kinase activity in a dose-dependent manner. The effect of this peptide was studied on specific granulocyte effector functions such as phagocytosis, respiratory burst, migration, and adhesion. Phagocytosis of Aspergillus fumigatus particles was reduced in the presence of CKLiK297-321 and fMLP-induced reactive oxygen species (ROS) production was potently inhibited by CKLiK297-321 in a dose-dependent manner. Furthermore, fMLP-induced neutrophil migration on albumin-coated surfaces was perturbed, as well as beta2-integrin-mediated adhesion. These findings suggest a critical role for CKLiK in modulating chemoattractant-induced functional responses in human granulocytes.
In this report, we cloned a novel calmodulin-kinase (CaM-KIdelta) from HeLa cells and characterized its activation mechanism. CaM-KIdelta exhibits Ca(2+)/CaM-dependent activity that is enhanced (approximately 30-fold) in vitro by phosphorylation of its Thr180 by CaM-K kinase (CaM-KK)alpha, consistent with detection of CaM-KIdelta-activating activity in HeLa cells. We also identified a novel CaM-KKbeta isoform (CaM-KKbeta-3) in HeLa cells whose activity was highly Ca(2+)/CaM-independent. Transiently expressed CaM-KIdelta exhibited enhanced protein kinase activity in HeLa cells without ionomycin stimulation. This sustained activation of CaM-KIdelta was completely abolished by Thr180Ala mutation and inhibited by CaM-KK inhibitor, STO-609, indicating a functional CaM-KK/CaM-KIdelta cascade in HeLa cells.
Activated by Ca(2+)/calmodulin. Binding of calmodulin results in conformational change that relieves intrasteric autoinhibition and allows phosphorylation of Thr-180 within the activation loop by CaMKK1 or CaMKK2. Phosphorylation of Thr-180 results in several fold increase in total activity. Unlike CaMK4, may be unable to exhibit autonomous activity after Ca(2+)/calmodulin activation.
Human granulocytes are characterized by a variety of specific effector functions involved in host defense. Several widely expressed protein kinases have been implicated in the regulation of these effector functions. A polymerase chain reaction-based strategy was used to identify novel granulocyte-specific kinases. A novel protein kinase complementary DNA with an open reading frame of 357 amino acids was identified with homology to calcium-calmodulin-dependent kinase I (CaMKI). This has been termed CaMKI-like kinase (CKLiK). Analysis of CKLiK messenger RNA (mRNA) expression in hematopoietic cells demonstrated an almost exclusive expression in human polymorphonuclear leukocytes (PMN). Up-regulation of CKLiK mRNA occurs during neutrophilic differentiation of CD34(+) stem cells. CKLiK kinase activity was dependent on Ca(++) and calmodulin as analyzed by in vitro phosphorylation of cyclic adenosine monophosphate responsive element modulator (CREM). Furthermore, CKLiK- transfected cells treated with ionomycin demonstrated an induction of CRE- binding protein (CREB) transcriptional activity compared to control cells. Additionally, CaMK-kinasealpha enhanced CKLiK activity. In vivo activation of CKLiK was shown by addition of interleukin (IL)-8 to a myeloid cell line stably expressing CKLiK. Furthermore inducible activation of CKLiK was sufficient to induce extracellular signal-related kinase (ERK) mitogen-activated protein (MAP) kinase activity. These data identify a novel Ca(++)/calmodulin-dependent PMN- specific kinase that may play a role in Ca(++)-mediated regulation of human granulocyte functions.
In this report, we cloned a novel calmodulin-kinase (CaM-KIdelta) from HeLa cells and characterized its activation mechanism. CaM-KIdelta exhibits Ca(2+)/CaM-dependent activity that is enhanced (approximately 30-fold) in vitro by phosphorylation of its Thr180 by CaM-K kinase (CaM-KK)alpha, consistent with detection of CaM-KIdelta-activating activity in HeLa cells. We also identified a novel CaM-KKbeta isoform (CaM-KKbeta-3) in HeLa cells whose activity was highly Ca(2+)/CaM-independent. Transiently expressed CaM-KIdelta exhibited enhanced protein kinase activity in HeLa cells without ionomycin stimulation. This sustained activation of CaM-KIdelta was completely abolished by Thr180Ala mutation and inhibited by CaM-KK inhibitor, STO-609, indicating a functional CaM-KK/CaM-KIdelta cascade in HeLa cells.
Protein involved in the localized protective response to tissue damage, microbial infection, or the presence of foreign matter. It is characterized by swelling, redness, heat and pain and involves a complex series of events including vascular changes and accumulation of blood cells, such as neutrophil leucocytes and mononuclear phagocytes, at the site of injury.
Protein which catalyzes the phosphorylation of serine or threonine residues on target proteins by using ATP as phosphate donor. Such phosphorylation may cause changes in the function of the target protein. Protein kinases share a conserved catalytic core common to both serine/ threonine and tyrosine protein kinases.
Enzyme whose activity is modified by the noncovalent binding of an allosteric effector at a site other than the active site. This binding mediates conformational changes, altering its catalytic or binding properties.
A reference proteome is a set of protein sequences derived from a complete proteome which constitutes a defined standard for a particular user community. Reference proteomes are manually defined according to a number of criteria. They cover the proteomes of well- studied model organisms and other proteomes of interest for biomedical and biotechnological research. Reference proteomes have been selected to provide broad coverage of the tree of life, and constitute a representative cross-section of the taxonomic diversity to be found within UniProtKB.
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