Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:2.7.11.1 (protein kinase)
 81,284 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

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.
 ...
PMID:Identification and characterization of novel components of a Ca2+/calmodulin-dependent protein kinase cascade in HeLa cells. 1293 86

Elevated intracellular Ca(2+) triggers numerous signaling pathways including protein kinases such as the calmodulin-dependent kinases (CaMKs) and the extracellular signal-regulated kinases (ERKs). In the present study we examined Ca(2+)-dependent "cross-talk" between these two protein kinase families. Using a combination of pharmacological inhibitors and dominant-negative kinases (dnKinase), we identified a requirement for CaMKK acting through CaMKI in the stimulation of ERKs upon depolarization of the neuroblastoma cell line, NG108. Depolarization stimulated prolonged ERK and JNK activation that was blocked by the CaMKK inhibitor, STO-609; this inhibition of ERK activation by STO-609 was rescued by expression of a STO-609-insensitive mutant of CaMKK. However, activation of ERK by epidermal growth factor or carbachol were not suppressed by inhibition of CaMKK, indicating specificity for this "cross-talk." To identify the downstream target of CaMKK that mediated ERK activation upon depolarization, dnKinases were expressed. The dnCaMKI completely suppressed ERK2 activation whereas dnAKT/PKB or nuclear-targeted dnCaMKIV, other substrates for CaMKK, were not inhibitory. ERK activation upon depolarization or transfection with constitutively active (ca) CaMKI was blocked by dnRas. Additionally, depolarization of NG108 cells promoted neurite outgrowth, and this effect was blocked by inhibition of either CaMKK (STO-609) or ERK (UO126). Co-transfection with caCaMKK plus caCaMKI also stimulated neurite outgrowth that was blocked by inhibition of ERK (UO126). These data are the first to suggest that ERK activation and neurite outgrowth in response to depolarization are mediated by CaMKK activation of CaMKI.
 ...
PMID:Calcium activation of ERK mediated by calmodulin kinase I. 1515 Feb 58

The AMP-activated protein kinase (AMPK) is an important regulator of cellular metabolism in response to metabolic stress and to other regulatory signals. AMPK activity is absolutely dependent upon phosphorylation of AMPKalphaThr-172 in its activation loop by one or more AMPK kinases (AMPKKs). The tumor suppressor kinase, LKB1, is a major AMPKK present in a variety of tissues and cells, but several lines of evidence point to the existence of other AMPKKs. We have employed three cell lines deficient in LKB1 to study AMPK regulation and phosphorylation, HeLa, A549, and murine embryo fibroblasts derived from LKB(-/-) mice. In HeLa and A549 cells, mannitol, 2-deoxyglucose, and ionomycin, but not 5-aminoimidazole-4-carboxamide-1-beta-d-ribofuranoside (AICAR), treatment activates AMPK by alphaThr-172 phosphorylation. These responses, as well as the downstream effects of AMPK on the phosphorylation of acetyl-CoA carboxylase, are largely inhibited by the Ca(2+)/ calmodulin-dependent protein kinase kinase (CaMKK) inhibitor, STO-609. AMPKK activity in HeLa cell lysates measured in vitro is totally inhibited by STO-609 with an IC50 comparable with that of the known CaMKK isoforms, CaMKKalpha and CaMKKbeta. Furthermore, 2-deoxyglucose- and ionomycin-stimulated AMPK activity, alphaThr-172 phosphorylation, and acetyl-CoA carboxylase phosphorylation are substantially reduced in HeLa cells transfected with small interfering RNAs specific for CaMKKalpha and CaMKKbeta. Lastly, the activation of AMPK in response to ionomycin and 2-deoxyglucose is not impaired in LKB1(-/-) murine embryo fibroblasts. These data indicate that the CaMKKs function in intact cells as AMPKKs, predicting wider roles for these kinases in regulating AMPK activity in vivo.
 ...
PMID:The Ca2+/calmodulin-dependent protein kinase kinases are AMP-activated protein kinase kinases. 1598 64

The AMP-activated protein kinase (AMPK) is a critical regulator of energy balance at both the cellular and whole-body levels. Two upstream kinases have been reported to activate AMPK in cell-free assays, i.e., the tumor suppressor LKB1 and calmodulin-dependent protein kinase kinase. However, evidence that this is physiologically relevant currently only exists for LKB1. We now report that there is a significant basal activity and phosphorylation of AMPK in LKB1-deficient cells that can be stimulated by Ca2+ ionophores, and studies using the CaMKK inhibitor STO-609 and isoform-specific siRNAs show that CaMKKbeta is required for this effect. CaMKKbeta also activates AMPK much more rapidly than CaMKKalpha in cell-free assays. K(+)-induced depolarization in rat cerebrocortical slices, which increases intracellular Ca2+ without disturbing cellular adenine nucleotide levels, activates AMPK, and this is blocked by STO-609. Our results suggest a potential Ca(2+)-dependent neuroprotective pathway involving phosphorylation and activation of AMPK by CaMKKbeta.
 ...
PMID:Calmodulin-dependent protein kinase kinase-beta is an alternative upstream kinase for AMP-activated protein kinase. 1605 95

The Ca(2+)/calmodulin (CaM) competitive inhibitor KN-93 has previously been used to evaluate 5'-AMP-activated protein kinase (AMPK)-independent Ca(2+)-signaling to contraction-stimulated glucose uptake in muscle during intense electrical stimulation ex vivo. With the use of low-intensity tetanic contraction of mouse soleus and extensor digitorum longus (EDL) muscles ex vivo, this study demonstrates that KN-93 can potently inhibit AMPK phosphorylation and activity after 2 min but not 10 min of contraction while strongly inhibiting contraction-stimulated 2-deoxyglucose uptake at both the 2- and 10-min time points. These data suggest inhibition of Ca(2+)/CaM-dependent signaling events upstream of AMPK, the most likely candidate being the novel AMPK kinase CaM-dependent protein kinase kinase (CaMKK). CaMKK protein expression was detected in mouse skeletal muscle. Similar to KN-93, the CaMKK inhibitor STO-609 strongly reduced AMPK phosphorylation and activity at 2 min and less potently at 10 min. Pretreatment with STO-609 inhibited contraction-stimulated glucose uptake at 2 min in soleus, but not EDL, and in both muscles after 10 min. Neither KN-93 nor STO-609 inhibited 5-aminoimidazole-4-carboxamide-1-beta-4-ribofuranoside-stimulated glucose uptake, AMPK phosphorylation, or recombinant LKB1 activity, suggestive of an LKB1-independent effect. Finally, neither KN-93 nor STO-609 had effects on the reductions in glucose uptake seen in mice overexpressing a kinase-dead AMPK construct, indicating that the effects of KN-93 and STO-609 on glucose uptake require inhibition of AMPK activity. We propose that CaMKKs act in mouse skeletal muscle regulating AMPK phosphorylation and glucose uptake at the onset of mild tetanic contraction and that an intensity- and/or time-dependent switch occurs in the relative importance of AMPKKs during contraction.
 ...
PMID:Possible CaMKK-dependent regulation of AMPK phosphorylation and glucose uptake at the onset of mild tetanic skeletal muscle contraction. 1721 73

Studies in nonmuscle cells have demonstrated that Ca(2+)/calmodulin-dependent protein kinase kinases (CaMKKs) are upstream regulators of AMP-activated protein kinase (AMPK) and Akt. In skeletal muscle, activation of AMPK and Akt has been implicated in the regulation of glucose uptake. The objective of this study was to determine whether CaMKKalpha regulates skeletal muscle glucose uptake, and whether it is dependent on AMPK and/or Akt activation. Expression vectors containing constitutively active CaMKKalpha (caCaMKKalpha) or empty vector were transfected into mouse muscles by in vivo electroporation. After 2 weeks, caCaMKKalpha was robustly expressed and increased CaMKI (Thr(177/180)) phosphorylation, a known CaMKK substrate. In muscles from wild-type mice, caCaMKKalpha increased in vivo [(3)H]-2-deoxyglucose uptake 2.5-fold and AMPKalpha1 and -alpha2 activities 2.5-fold. However, in muscles from AMPKalpha2 inactive mice (AMPKalpha2i), caCaMKKalpha did not increase AMPKalpha1 or -alpha2 activities, but it did increase glucose uptake 2.5-fold, demonstrating that caCaMKKalpha stimulates glucose uptake independent of AMPK. Akt (Thr(308)) phosphorylation was not altered by CaMKKalpha, and caCaMKKalpha plus insulin stimulation did not increase the insulin-induced phosphorylation of Akt (Thr(308)). These results suggest that caCaMKKalpha stimulates glucose uptake via insulin-independent signaling mechanisms. To assess the role of CaMKK in contraction-stimulated glucose uptake, isolated muscles were treated with or without the CaMKK inhibitor STO-609 and then electrically stimulated to contract. Contraction increased glucose uptake 3.5-fold in muscles from both wild-type and AMPKalpha2i mice, but STO-609 significantly decreased glucose uptake (approximately 24%) only in AMPKalpha2i mice. Collectively, these results implicate CaMKKalpha in the regulation of skeletal muscle glucose uptake independent of AMPK and Akt activation.
 ...
PMID:Ca2+/calmodulin-dependent protein kinase kinase-alpha regulates skeletal muscle glucose uptake independent of AMP-activated protein kinase and Akt activation. 1728 69

Ca(2+)/calmodulin-dependent protein kinase kinases (CaMKKs) are upstream protein kinases that phosphorylate and activate CaMKI and CaMKIV, both of which are involved in a variety of neuronal functions. Here, we first demonstrated that the two isoforms of CaMKK were differentially expressed during neural development by in situ hybridization. We also demonstrated that both dominant negative and pharmacological interference with CaMKK inhibitor, STO-609 resulted in a significant decrease in the number of primary dendrites of cultured hippocampal neurons. Our present findings provide the detailed anatomical information on the developmental expression of CaMKKs and the functional involvement of CaMKK in the formation of primary dendrites.
 ...
PMID:Distinct developmental expression of two isoforms of Ca2+/calmodulin-dependent protein kinase kinases and their involvement in hippocampal dendritic formation. 1766 91

Calcium (Ca(2+))/calmodulin-dependent protein kinase kinase (CaMKK) is a novel member of Ca(2+)/calmodulin-dependent protein kinase (CaMK) family, whose physiological roles in regulating meiotic cell cycle needs to be determined. We showed by Western blot that CaMKK was expressed in pig oocytes at various maturation stages. Confocal microscopy was employed to observe CaMKK distribution. In oocytes at the germinal vesicle (GV) or prometaphase I (pro-MI) stage, CaMKK was distributed in the nucleus, around the condensed chromatin and the cortex of the cell. At metaphase I (MI) stage, CaMKK was concentrated in the cortex of the cell. After transition to anaphase I or telophase I stage, CaMKK was detected around the separating chromosomes and in the cortex of the cell. At metaphase II (MII) stage, CaMKK was localized to the cortex of the cell, with a thicker area near the first polar body (PB1). Treatment of pig cumulus-enclosed oocytes with STO-609, a membrane-permeable CaMKK inhibitor, resulted in the delay/inhibition of the meiotic resumption and the inhibition of first polar body emission. The correlation between CaMKK and microfilaments during meiotic maturation of pig oocytes was then studied. CaMKK and microfilaments were colocalized from MI to MII during porcine oocyte maturation. After oocytes were treated with STO-609, microfilaments were depolymerized, while in oocytes exposed to cytochalasin B (CB), a microfilament polymerization inhibitor, CaMKK became diffused evenly throughout the cell. These data suggest that CaMKK is involved in regulating the meiotic cell cycle probably by interacting with microfilaments in pig oocytes.
 ...
PMID:Involvement of calcium/calmodulin-dependent protein kinase kinase in meiotic maturation of pig oocytes. 1836 50

Smooth muscle contraction is initiated by a rise in intracellular calcium, leading to activation of smooth muscle myosin light chain kinase (MLCK) via calcium/calmodulin (CaM). Activated MLCK then phosphorylates the regulatory myosin light chains, triggering cross-bridge cycling and contraction. Here, we show that MLCK is a substrate of AMP-activated protein kinase (AMPK). The phosphorylation site in chicken MLCK was identified by mass spectrometry to be located in the CaM-binding domain at Ser(815). Phosphorylation by AMPK desensitized MLCK by increasing the concentration of CaM required for half-maximal activation. In primary cultures of rat aortic smooth muscle cells, vasoconstrictors activated AMPK in a calcium-dependent manner via CaM-dependent protein kinase kinase-beta, a known upstream kinase of AMPK. Indeed, vasoconstrictor-induced AMPK activation was abrogated by the STO-609 CaM-dependent protein kinase kinase-beta inhibitor. Myosin light chain phosphorylation was increased under these conditions, suggesting that contraction would be potentiated by ablation of AMPK. Indeed, in aortic rings from mice in which alpha1, the major catalytic subunit isoform in arterial smooth muscle, had been deleted, KCl- or phenylephrine-induced contraction was increased. The findings suggest that AMPK attenuates contraction by phosphorylating and inactivating MLCK. This might contribute to reduced ATP turnover in the tonic phase of smooth muscle contraction.
 ...
PMID:AMP-activated protein kinase phosphorylates and desensitizes smooth muscle myosin light chain kinase. 1842 92

These studies explore the connections between simvastatin, Rac1, and AMP-activated protein kinase (AMPK) pathways in cultured vascular endothelial cells and in arterial preparations isolated from statin-treated mice. In addition to their prominent effects on lipoprotein metabolism, statins can regulate the small GTPase Rac1, and may also affect the phosphorylation of the ubiquitous AMPK. We explored pathways of statin-modulated Rac1 and AMPK activation both in arterial preparations from statin-treated mice as well as in cultured endothelial cells. We treated adult mice with simvastatin daily for 2 weeks and then harvested and analyzed arterial preparations. Simvastatin treatment of mice led to a significant increase in AMPK and LKB1 phosphorylation and to a decrease in protein kinase A activity relative to control animals, associated with a marked increase in Rac1 activation. Exposure of bovine aortic endothelial cells to simvastatin for 24 h strikingly increased GTP-bound Rac1 and led to increased phosphorylation of AMPK as well as the AMPK kinase LKB1. These responses to simvastatin were blocked by mevalonate or geranylgeranyl pyrophosphate but not by farnesyl pyrophosphate. Small interfering RNA (siRNA)-mediated knockdown of AMPK abrogated simvastatin-induced Rac1 activation and LKB1 phosphorylation. Importantly, siRNA-mediated knockdown of the key AMPK kinase, calcium/calmodulin-dependent protein kinase kinase beta, completely blocked simvastatin-induced endothelial cell migration and also abrogated statin-promoted phosphorylation of AMPK and LKB1, as did pharmacological inhibition with the specific calcium/calmodulin-dependent protein kinase beta inhibitor STO-609. Moreover, siRNA-mediated knockdown of Rac1 completely blocked simvastatin-induced LKB1 phosphorylation, but without affecting simvastatin-induced AMPK phosphorylation. These findings establish a key role for simvastatin in activation of a novel Rac1-dependent signaling pathway in the vascular wall.
 ...
PMID:Regulation of Rac1 by simvastatin in endothelial cells: differential roles of AMP-activated protein kinase and calmodulin-dependent kinase kinase-beta. 1933 49


1 2 3 4 5 Next >>