Lasp-1 is a regulated phosphoprotein within the cAMP  signaling pathway in the gastric parietal cell

Activation of the cAMP signaling pathway is correlated with increased secretory-related events in a wide variety of cell types including the gastric parietal cell. Within this pathway, as well as in other intracellular signaling pathways, protein phosphorylation serves as a major downstream regulatory mechanism. However, although agonist and cAMP-dependent activation of cAMP-dependent protein kinase (PKA) has been demonstrated, little is currently known about the downstream in vivo phosphoprotein substrates of this enzyme. Here we report the isolation, microsequencing, and cloning of a LIM and SH3 domain-containing, cAMP-responsive, 40-kDa phosphoprotein (pp40) from rabbit gastric parietal cells. The deduced amino acid sequence for pp40 is 93.5%, homologous with the putative protein product of the human gene lasp-1, which was recently identified based on its overexpression in some breast carcinomas. In addition to LIM and SH3 domains, the rabbit homolog contains two highly conserved PKA consensus sequences as well as two conserved SH2 binding motifs and several other putative protein kinase phosphorylation sites, including two for tyrosine kinase(s). Combined Northern and Western blot analyses indicate that pp40/lasp-1 is widely expressed (through a single 3.3-kb message) not only in epithelial tissues but also in muscle and brain. Furthermore, stimulation of isolated parietal cells, distal colonic crypts, and pancreatic cells with the adenylyl cyclase activator forskolin leads to the appearance of a higher molecular weight form of pp40/lasp-1, a finding which is consistent with an increase in protein phosphorylation. Thus pp40/lasp-1 appears to be regulated within the cAMP signaling pathway in a wide range of epithelial cell types. Because the cAMP-dependent increase in pp40 phosphorylation is correlated with secretory responses in the parietal cell and because pp40 appears to be widely distributed among various secretory tissues, this newly defined signaling protein may play an important role in modulating ionic transport or other secretory-related activities in many different cell types.

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Parietal cell MAP kinases: multiple activation pathways

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.1996.271.4.g640 ◽

1996 ◽

Vol 271 (4) ◽

pp. G640-G649 ◽

Cited By ~ 10

Author(s):

K. Nakamura ◽

C. J. Zhou ◽

J. Parente ◽

C. S. Chew

Keyword(s):

Protein Kinase ◽

Map Kinase ◽

Parietal Cell ◽

Intracellular Signaling ◽

Map Kinases ◽

Sodium Dodecyl ◽

Cell Types ◽

Parietal Cells ◽

Gastric Parietal Cell ◽

Cell Acid

Epidermal growth factor (EGF) is a potent mitogen for many cell types; however, the best known effect of EGF on gastric parietal cell HCl secretion is inhibition of this response. Using rabbit parietal cells in primary culture, we recently showed that the effect of EGF is biphasic with acute inhibition followed by sustained enhancement of acid secretory-related responses. We hypothesized that EGF might activate a mitogen-activated protein (MAP) kinase signaling pathway in parietal cells, and this pathway might play a role in mediating sustained and/or acute effects of EGF on parietal cell acid secretory-related functions [C. S. Chew, K. Nakamura, and A. C. Petropolous. Am. J. Physiol. 267 (Gastrointest. Liver Physiol. 30): G818-G826, 1994]. We used several methodological approaches to demonstrate the presence of MAP kinase (MAPK) isoforms, extracellular signal-regulated kinases (ERKs) 1 and 2, in parietal cells and to begin to characterize their mechanisms of activation in this highly differentiated cell type. In acutely isolated, 90-98% enriched parietal cells, EGF biphasically activated ERK-1 and ERK-2, with peak response occurring at approximately 5 min followed by a sustained lower level of activation for at least 2 h. The EC50 for EGF (1.2 +/- 0.4 nM) was similar to the previously determined EC50 for the stimulatory effect of EGF on acid secretory responses. In contrast to EGF, the phorbol ester protein kinase C activator 12-O-tetradecanoylphorbol 13-acetate (TPA) induced a sustained activation of ERK-1 and ERK-2 for at least 2 h. Carbachol also activated ERK-1 and ERK-2; however, this response was weaker and monophasic. Neither the Ca2+ ionophore ionomycin nor the adenylyl cyclase activator forskolin altered basal or stimulated ERK activity. Carbachol, but not EGF or TPA, also activated an unidentified 70-kDa protein kinase as detected with in-gel myelin basic protein (MBP) kinase renaturation assays. Parietal cell MAPK activation was not correlated to a shift in apparent relative molecular mass on sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels, suggesting that basal phosphorylation of ERK isoforms may be higher in parietal cells compared with actively proliferating cell lines. Also, in contrast to observations in neutrophils, the phosphatidylinositol 3-kinase (PtdIns 3-kinase) inhibitor, wortmannin (0.3-3 microM), failed to inhibit ERK activation in response to EGF, carbachol, or TPA. The combined data indicate that 1) EGF, TPA, and carbachol activate overlapping as well as distinct intracellular signaling pathways in gastric parietal cells, 2) EGF activates ERKs and enhances parietal cell acid secretory related functions via receptors with similar affinities, and 3) in contrast to some cell types, the parietal cell ERK-signaling cascade does not appear to be directly modulated by the PtdIns 3-kinase pathway or by elevated intracellular free Ca2+ or adenosine 3',5'-cyclic monophosphate concentrations.

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Drebrin E2 is differentially expressed and phosphorylated in parietal cells in the gastric mucosa

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00002.2005 ◽

2005 ◽

Vol 289 (2) ◽

pp. G320-G331 ◽

Cited By ~ 19

Author(s):

Catherine S. Chew ◽

Curtis T. Okamoto ◽

Xunsheng Chen ◽

Ruby Thomas

Keyword(s):

Gastric Mucosa ◽

Actin Filament ◽

Parietal Cell ◽

Active Role ◽

Cell Types ◽

Parietal Cells ◽

Dominant Negative ◽

Filament Formation ◽

Differential Distribution ◽

Gastric Parietal Cell

Developmentally regulated brain proteins (drebrins) are highly expressed in brain where they may regulate actin filament formation in dendritic spines. Recently, the drebrin E2 isoform was detected in certain epithelial cell types including the gastric parietal cell. In gastric parietal cells, activation of HCl secretion is correlated with actin filament formation and elongation within intracellular canaliculi, which are the sites of acid secretion. The aim of this study was to define the pattern of drebrin expression in gland units in the intact rabbit oxyntic gastric mucosa and to initiate approaches to define the functions of this protein in parietal cells. Drebrin E2 expression was limited entirely or almost entirely to parietal cells and depended upon the localization of parietal cells along the gland axis. Rabbit drebrin E2 was cloned and found to share 86% identity with human drebrin 1a and to possess a number of cross-species conserved protein-protein interaction and phosphorylation consensus sites. Two-dimensional Western blot and phosphoaffinity column analyses confirmed that drebrin is phosphorylated in parietal cells, and several candidate phosphorylation sites were identified by mass spectrometry. Overexpression of epitope-tagged drebrin E2 led to the formation of microspikes and F-actin-rich ring-like structures in cultured parietal cells and suppressed cAMP-dependent acid secretory responses. In Madin-Darby canine kidney cells, coexpression of epitope-tagged drebrin and the Rho family GTPase Cdc42, which induces filopodial extension, produced an additive increase in the length of microspike projections. Coexpression of dominant negative Cdc42 with drebrin E2 did not prevent drebrin-induced microspike formation. These findings suggest that 1) drebrin can induce the formation of F-actin-rich membrane projections by Cdc42-dependent and -independent mechanisms; and that 2) drebrin plays an active role in directing the secretagogue-dependent formation of F-actin-rich filaments on the parietal cell canalicular membrane. Finally, the differential distribution of drebrin in parietal cells along the gland axis suggests that drebrin E2 may be an important marker of parietal cell differentiation and functionality.

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The cAMP Signaling Pathway and Direct Protein Kinase A Phosphorylation Regulate Polycystin-2 (TRPP2) Channel Function

Journal of Biological Chemistry ◽

10.1074/jbc.m115.661082 ◽

2015 ◽

Vol 290 (39) ◽

pp. 23888-23896 ◽

Cited By ~ 13

Author(s):

María del Rocío Cantero ◽

Irina F. Velázquez ◽

Andrew J. Streets ◽

Albert C. M. Ong ◽

Horacio F. Cantiello

Keyword(s):

Protein Kinase ◽

Protein Kinase A ◽

Signaling Pathway ◽

Camp Signaling ◽

Channel Function ◽

Camp Signaling Pathway ◽

Kinase A

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Cyclic AMP Stimulates SF-1-Dependent CYP11A1 Expression through Homeodomain-Interacting Protein Kinase 3-Mediated Jun N-Terminal Kinase and c-Jun Phosphorylation

Molecular and Cellular Biology ◽

10.1128/mcb.02253-06 ◽

2007 ◽

Vol 27 (6) ◽

pp. 2027-2036 ◽

Cited By ~ 44

Author(s):

Hsin-Chieh Lan ◽

Hua-Jung Li ◽

Guang Lin ◽

Pao-Yen Lai ◽

Bon-chu Chung

Keyword(s):

Gene Expression ◽

Protein Kinase ◽

Cyclic Amp ◽

Signaling Pathway ◽

Camp Signaling ◽

Functional Interaction ◽

Interacting Protein ◽

Surface Receptors ◽

Camp Signaling Pathway ◽

Camp Stimulation

ABSTRACT Steroids are synthesized in adrenal glands and gonads under the control of pituitary peptides. These peptides bind to cell surface receptors to activate the cyclic AMP (cAMP) signaling pathway leading to an increase of steroidogenic gene expression. Exactly how cAMP activates steroidogenic gene expression is not clear, except for the knowledge that transcription factor SF-1 plays a key role. Investigating the factors participating in SF-1 action, we found that c-Jun and homeodomain-interacting protein kinase 3 (HIPK3) were required for basal and cAMP-stimulated expression of one major steroidogenic gene, CYP11A1. HIPK3 enhanced SF-1 activity, and c-Jun was required for the functional interaction of HIPK3 with SF-1. Furthermore, after cAMP stimulation, both c-Jun and Jun N-terminal kinase (JNK) were phosphorylated through HIPK3. These phosphorylations were important for SF-1 activity and CYP11A1 expression. Thus, we have defined HIPK3-mediated JNK activity and c-Jun phosphorylation as important events that increase SF-1 activity for CYP11A1 transcription in response to cAMP. This finding has linked three common factors, HIPK3, JNK, and c-Jun, to the cAMP signaling pathway leading to increased steroidogenic gene expression.

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The RII regulatory subunit of protein kinase A binds to cAMP response element: An alternative cAMP signaling pathway

Proceedings of the National Academy of Sciences ◽

10.1073/pnas.95.12.6687 ◽

1998 ◽

Vol 95 (12) ◽

pp. 6687-6692 ◽

Cited By ~ 16

Author(s):

R. K. Srivastava ◽

Y. N. Lee ◽

K. Noguchi ◽

Y. G. Park ◽

M. J. C. Ellis ◽

...

Keyword(s):

Protein Kinase ◽

Protein Kinase A ◽

Signaling Pathway ◽

Camp Signaling ◽

Regulatory Subunit ◽

Camp Response Element ◽

Response Element ◽

Camp Signaling Pathway ◽

Kinase A

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cAMP-signaling pathway acts in selective synergism with glucose or tolbutamide to increase cytosolic Ca2+ in rat pancreatic beta-cells

Diabetes ◽

10.2337/diabetes.45.3.295 ◽

1996 ◽

Vol 45 (3) ◽

pp. 295-301 ◽

Cited By ~ 8

Author(s):

K. Yaekura ◽

M. Kakei ◽

T. Yada

Keyword(s):

Signaling Pathway ◽

Beta Cells ◽

Pancreatic Beta Cells ◽

Camp Signaling ◽

Camp Signaling Pathway

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Role of cAMP in Double Switch of Glucagon Secretion

Cells ◽

10.3390/cells10040896 ◽

2021 ◽

Vol 10 (4) ◽

pp. 896

Author(s):

Jan Zmazek ◽

Vladimir Grubelnik ◽

Rene Markovič ◽

Marko Marhl

Keyword(s):

Glucose Metabolism ◽

Signaling Pathway ◽

Cell Model ◽

Dominant Role ◽

Glucagon Secretion ◽

Camp Signaling ◽

Independent Manner ◽

Metabolic Switch ◽

Camp Signaling Pathway ◽

Double Switch

Glucose metabolism plays a crucial role in modulating glucagon secretion in pancreatic alpha cells. However, the downstream effects of glucose metabolism and the activated signaling pathways influencing glucagon granule exocytosis are still obscure. We developed a computational alpha cell model, implementing metabolic pathways of glucose and free fatty acids (FFA) catabolism and an intrinsically activated cAMP signaling pathway. According to the model predictions, increased catabolic activity is able to suppress the cAMP signaling pathway, reducing exocytosis in a Ca2+-dependent and Ca2+ independent manner. The effect is synergistic to the pathway involving ATP-dependent closure of KATP channels and consequent reduction of Ca2+. We analyze the contribution of each pathway to glucagon secretion and show that both play decisive roles, providing a kind of “secure double switch”. The cAMP-driven signaling switch plays a dominant role, while the ATP-driven metabolic switch is less favored. The ratio is approximately 60:40, according to the most recent experimental evidence.

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Assessment of cellular mechanisms contributing to cAMP compartmentalization in pulmonary microvascular endothelial cells

AJP Cell Physiology ◽

10.1152/ajpcell.00361.2011 ◽

2012 ◽

Vol 302 (6) ◽

pp. C839-C852 ◽

Cited By ~ 38

Author(s):

Wei P. Feinstein ◽

Bing Zhu ◽

Silas J. Leavesley ◽

Sarah L. Sayner ◽

Thomas C. Rich

Keyword(s):

Endothelial Cells ◽

Adenylyl Cyclase ◽

Signaling Pathway ◽

Camp Signaling ◽

Microvascular Endothelial Cells ◽

Camp Production ◽

Barrier Integrity ◽

Pulmonary Microvascular Endothelial Cells ◽

Camp Signaling Pathway ◽

Microvascular Endothelial

Cyclic AMP signals encode information required to differentially regulate a wide variety of cellular responses; yet it is not well understood how information is encrypted within these signals. An emerging concept is that compartmentalization underlies specificity within the cAMP signaling pathway. This concept is based on a series of observations indicating that cAMP levels are distinct in different regions of the cell. One such observation is that cAMP production at the plasma membrane increases pulmonary microvascular endothelial barrier integrity, whereas cAMP production in the cytosol disrupts barrier integrity. To better understand how cAMP signals might be compartmentalized, we have developed mathematical models in which cellular geometry as well as total adenylyl cyclase and phosphodiesterase activities were constrained to approximate values measured in pulmonary microvascular endothelial cells. These simulations suggest that the subcellular localizations of adenylyl cyclase and phosphodiesterase activities are by themselves insufficient to generate physiologically relevant cAMP gradients. Thus, the assembly of adenylyl cyclase, phosphodiesterase, and protein kinase A onto protein scaffolds is by itself unlikely to ensure signal specificity. Rather, our simulations suggest that reductions in the effective cAMP diffusion coefficient may facilitate the formation of substantial cAMP gradients. We conclude that reductions in the effective rate of cAMP diffusion due to buffers, structural impediments, and local changes in viscosity greatly facilitate the ability of signaling complexes to impart specificity within the cAMP signaling pathway.

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