Phosphorylation of Calmodulin by Plasma‐Membrane‐Associated Protein Kinase(s)

Benguría, Alberto; Soriano, Montserrat; Joyal, John L.; Sacks, David B.; Villalobo, Antonio

doi:10.1111/j.1432-1033.1995.050_c.x

Cited by 11 publications

(12 citation statements)

References 46 publications

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“…Phosphorylation of CaM tightly bound to purified rat liver plasma membrane fractions by unidentified membrane‐bound protein kinases has been described previously [64,65]. This phosphorylation was inhibited by Ca 2+ , does not require the addition of exogenous polycations and takes place exclusively at serine residues [64,65].…”

Section: Phosphorylation Of Calmodulin By Nonreceptor Protein‐tyrosinmentioning

confidence: 77%

“…This phosphorylation was inhibited by Ca 2+ , does not require the addition of exogenous polycations and takes place exclusively at serine residues [64,65]. In contrast, the phosphorylation of exogenous CaM by these plasma membrane preparations requires the presence of polycations presenting a biphasic behaviour toward the polycation/calmodulin (mol/mol) ratio, and takes place at serine, threonine plus tyrosine residues [65]. The phosphorylation of exogenous CaM was more sensitive to the inhibitory action of Ca 2+ than that of endogenous CaM, presenting a K′ i of 1 µ m by the former and higher than 0.1 m m by the latter [65].…”

Section: Phosphorylation Of Calmodulin By Nonreceptor Protein‐tyrosinmentioning

confidence: 99%

“…In contrast, the phosphorylation of exogenous CaM by these plasma membrane preparations requires the presence of polycations presenting a biphasic behaviour toward the polycation/calmodulin (mol/mol) ratio, and takes place at serine, threonine plus tyrosine residues [65]. The phosphorylation of exogenous CaM was more sensitive to the inhibitory action of Ca 2+ than that of endogenous CaM, presenting a K′ i of 1 µ m by the former and higher than 0.1 m m by the latter [65]. The differential phosphorylation of endogenous and exogenous CaM by protein kinases associated to the plasma membrane is an interesting observation, as underscores that both P‐CaM pools could have different functions in the cell.…”

Section: Phosphorylation Of Calmodulin By Nonreceptor Protein‐tyrosinmentioning

confidence: 99%

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Phosphorylation of calmodulin

Benaím

Villalobo

2002

European Journal of Biochemistry

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134

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Calmodulin (CaM) is phosphorylated in vitro and in vivo by multiple protein-serine/threonine and protein-tyrosine kinases. Casein kinase II and myosin light-chain kinase are two of the well established protein-serine/threonine kinases implicated in this process. On the other hand, within the protein-tyrosine kinases involved in the phosphorylation of CaM are receptors with tyrosine kinase activity, such as the insulin receptor and the epidermal growth factor receptor, and nonreceptor protein-tyrosine kinases, such as several members of the Src family kinases, Janus kinase 2, and p38Syk. The phosphorylation of CaM brings important physiological consequences for the cell as the diverse phosphocalmodulin species have differential actions as compared to nonphosphorylated CaM when acting on different CaM-dependent systems. In this review we will summarize the progress made on this topic as the first report on phosphorylation of CaM was published almost two decades ago. We will emphasize the description of the phosphorylation events mediated by the different protein kinases not only in the test tube but in intact cells, the phosphorylation-mediated changes of CaM activity, its action on CaM-dependent systems, and the functional repercussion of these phosphorylation processes in the physiology of the cell.Keywords: calmodulin; calmodulin-dependent systems; cellular signalling; phosphocalmodulin; protein kinases; phosphoprotein phosphatases. I N T R O D U C T I O N Calmodulin as a Ca 2+ sensorThe average cytosolic concentration of free Ca 2+ in resting cells ranges from 20 to 50 nM, reaching values close to 1 lM when the cell is stimulated by a variety of physiological stimuli, while in the extracellular milieu this concentration is about 1 mM. This large concentration gradient allows intracellular Ca 2+ to work as an useful second messenger [1,2]. The cytosolic Ca 2+ concentration is exquisitely regulated by the operation of transport systems responsible for its increase, represented by different types of Ca 2+ channels and Na + (H + )/Ca 2+ exchangers located in the plasma membrane, the endo(sarco)plasmic reticulum, and/or the mitochondria; and extrusion transport systems represented by Ca 2+ -ATPases located in both the plasma membrane and the endo(sarco)plasmic reticulum, and the Na + /Ca 2+ exchanger also located within the plasma membrane [1,3]. The operation of theses transporters gives rise to oscillations in the concentration of Ca 2+ not only in the cytosol but in the nucleus and intracellular organelles [1,4,5]. It has been possible to observe in living cells inhomogeneities in these transient changes inCorrespondence to A. Villalobo, Instituto de Investigaciones Biome´dicas, Consejo Superior de Investigaciones Cientı´ficas and Universidad Auto´noma de Madrid, Arturo Duperier 4, E-28029 Madrid, Spain. E-mail: antonio.villalobo@iib.uam.es Abbreviations: AdeCycl, adenylate cyclase; CaM, calmodulin; CaM-BD, calmodulin-binding domain; CaMPK-II, calmodulin-dependent protein kinase II; CK-II, casein kinase II; CKR,...

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Section: Phosphorylation Of Calmodulin By Nonreceptor Protein‐tyrosinmentioning

confidence: 77%

Section: Phosphorylation Of Calmodulin By Nonreceptor Protein‐tyrosinmentioning

confidence: 99%

Section: Phosphorylation Of Calmodulin By Nonreceptor Protein‐tyrosinmentioning

confidence: 99%

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Phosphorylation of calmodulin

Benaím

Villalobo

2002

European Journal of Biochemistry

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134

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“…The amount of phosphocalmodulin isolated by the purification procedure was determined by immunoblot performed as described above except that the PVDF membrane was probed with an anti-calmodulin monoclonal antibody (1/1,000 dilution) that recognizes both phosphorylated and nonphosphorylated calmodulin and a secondary anti-mouse IgGs antibody coupled to alkaline phosphatase (1/1000 dilution) (19). Color development was done with 0.32 mg/ml NBT and 0.16 mg/ml BCIP in 100 mM Tris-HCl (pH 9.5), 100 mM NaCl, and 5 mM MgCl 2 .…”

Section: Preparation Of Liver Plasma Membrane Fractionsmentioning

confidence: 99%

A Method for the Purification of Phospho(Tyr)calmodulin Free of Nonphosphorylated Calmodulin

Palomo-Jiménez

Hernández-Hernando

Garciá-Nieto

et al. 1999

Protein Expression and Purification

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“…Indeed, CLP shares with CaM several serine and threonine residues (Thr 79 , Ser 101 , Thr 117 ) as well as a tyrosine residue (Tyr 138 ) known to be targets for phosphorylation in CaM (36 -38). A substantial fraction of CaM tightly associated with the plasma membrane and underlying cytoskeleton has been shown to be phosphorylated by membrane-associated protein kinases (39). Assuming that CLP can be similarly phosphorylated, and given the cytoskeletal/membrane-associated location of myosin X, it may well be that a significant fraction of the myosin X-bound CLP in vivo is phosphorylated.…”

Section: Discussionmentioning

confidence: 99%

The Tumor-sensitive Calmodulin-like Protein Is a Specific Light Chain of Human Unconventional Myosin X

Rogers¹,

Strehler

2001

Journal of Biological Chemistry

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Human calmodulin-like protein (CLP) is an epithelialspecific Ca 2؉ -binding protein whose expression is strongly down-regulated in cancers. Like calmodulin, CLP is thought to regulate cellular processes via Ca 2؉ -dependent interactions with specific target proteins. Using gel overlays, we identified a ϳ210-kDa protein binding specifically and in a Ca 2؉ -dependent manner to CLP, but not to calmodulin. Yeast two-hybrid screening yielded a CLP-interacting clone encoding the three light chain binding IQ motifs of human "unconventional" myosin X. Pull-down experiments showed CLP binding to the IQ domain to be direct and Ca 2؉-dependent. CLP interacted strongly with IQ motif 3 (K d ϳ0.5 nM) as determined by surface plasmon resonance. Epitope-tagged myosin X was localized preferentially at the cell periphery in MCF-7 cells, and CLP colocalized with myosin X in these cells. Myosin X was able to coprecipitate CLP and, to a lesser extent, calmodulin from transfected COS-1 cells, indicating that CLP is a specific light chain of myosin X in vivo. Because unconventional myosins participate in cellular processes ranging from membrane trafficking to signaling and cell motility, myosin X is an attractive CLP target. Altered myosin X regulation in (tumor) cells lacking CLP may have as yet unknown consequences for cell growth and differentiation.Human calmodulin-like protein (CLP) 1 is encoded by an intronless gene localized on chromosome 10p13-ter (1), a chromosomal region known to be lost in a number of breast and other cancer types (2-6). Initial studies failed to show expression of the CLP gene in a number of tissues and cell types (7); however, in an independent study aimed at identifying mRNAs coding for transformation-sensitive proteins, CLP was recloned as a protein named NB1 (for Normal-Breast-1) from a normal primary breast epithelial cell line that had been subtracted with RNA purified from the same cell line after chemical transformation (8). CLP was subsequently shown to be expressed in a highly tissue-specific manner in basal and suprabasal epithelial cells of the normal breast, cervix, prostate, and skin, and to be down-regulated in naturally occurring breast, cervix, and prostate tumors as well as in artificially immortalized and transformed breast epithelial cells in culture (9). Recently, CLP was found to be down-regulated in 80% of primary breast tumors of both noninvasive as well as invasive and metastatic stages (10). Given the apparent correlation between its downregulation and the tumorigenic state, the available data support the notion that CLP expression may be incompatible with the transformed state of a cell. The mechanism responsible for this incompatibility is presently unknown, as is the physiological function of CLP, though they are likely related.CLP's deduced amino acid sequence of 148 residues is identical in length and shows 85% identity to calmodulin (CaM) (7,11). Accordingly, CLP shows a number of characteristics similar to those of CaM; however, it also displays unique features that sugge...

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Phosphorylation of Calmodulin by Plasma‐Membrane‐Associated Protein Kinase(s)

Cited by 11 publications

References 46 publications

Phosphorylation of calmodulin

Phosphorylation of calmodulin

A Method for the Purification of Phospho(Tyr)calmodulin Free of Nonphosphorylated Calmodulin

The Tumor-sensitive Calmodulin-like Protein Is a Specific Light Chain of Human Unconventional Myosin X

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