Cyclic AMP-dependent protein kinase A and EPAC mediate VIP and secretin stimulation of PAK4 and activation of Na+,K+-ATPase in pancreatic acinar cells

Rat pancreatic acinar cells possess only the p21-activated kinase (PAKs), PAK4 of the group II PAK, and it is activated by gastrointestinal hormones/neurotransmitters stimulating PLC and by a number of growth factors. However, little is known generally of cAMP agents causing PAK4 activation, and there are no studies with gastrointestinal hormones/neurotransmitters activating cAMP cascades. In the present study, we examined the ability of VIP and secretin, which stimulate cAMP generation in pancreatic acini, to stimulate PAK4 activation, the signaling cascades involved, and their possible role in activating sodium-potassium adenosine triphosphatase (Na+,K+-ATPase). PAK4 activation was compared with activation of the well-established cAMP target, cyclic AMP response element binding protein (CREB). Secretin-stimulated PAK4 activation was inhibited by KT-5720 and PKA Type II inhibitor (PKI), protein kinase A (PKA) inhibitors, whereas VIP activation was inhibited by ESI-09 and HJC0197, exchange protein directly activated by cAMP (EPAC) inhibitors. In contrast, both VIP/secretin-stimulated phosphorylation of CREB (pCREB) via EPAC activation; however, it was inhibited by the p44/42 inhibitor PD98059 and the p38 inhibitor SB202190. The specific EPAC agonist 8-CPT-2- O-Me-cAMP as well 8-Br-cAMP and forskolin stimulated PAK4 activation. Secretin/VIP activation of Na+,K+-ATPase, was inhibited by PAK4 inhibitors (PF-3758309, LCH-7749944). These results demonstrate PAK4 is activated in pancreatic acini by stimulation of both VIP-/secretin-preferring receptors, as is CREB. However, they differ in their signaling cascades. Furthermore, PAK4 activation is needed for Na+,K+ATPase activation, which mediates pancreatic fluid secretion. These results, coupled with recent studies reporting PAKs are involved in both pancreatitis/pancreatic cancer growth/enzyme secretion, show that PAK4, similar to PAK2, likely plays an important role in both pancreatic physiological/pathological responses. NEW & NOTEWORTHY Pancreatic acini possess only the group II p21-activated kinase, PAK4, which is activated by PLC-stimulating agents/growth factors and is important in enzyme-secretion/growth/pancreatitis. Little information exists on cAMP-activating agents stimulating group II PAKs. We studied ability/effect of cyclic AMP-stimulating agents [vasoactive intestinal polypeptide (VIP), secretin] on PAK4 activity in rat pancreatic-acini. Both VIP/secretin activated PAK4/CREB, but the cAMP signaling cascades differed for EPAC, MAPK, and PKA pathways. Both hormones require PAK4 activation to stimulate sodium-potassium adenosine triphosphatase activity. This study shows PAK4 plays an important role in VIP-/secretin-stimulated pancreatic fluid secretion and suggests it plays important roles in pancreatic acinar physiological/pathophysiological responses mediated by cAMP-activating agents.

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P21-activated kinase 4 in pancreatic acinar cells is activated by numerous gastrointestinal hormones/neurotransmitters and growth factors by novel signaling, and its activation stimulates secretory/growth cascades

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00005.2018 ◽

2018 ◽

Vol 315 (2) ◽

pp. G302-G317 ◽

Cited By ~ 4

Author(s):

Irene Ramos-Alvarez ◽

R. T. Jensen

Keyword(s):

Growth Factors ◽

Growth Factor ◽

Gastrointestinal Hormones ◽

Acinar Cells ◽

Pancreatic Acinar Cells ◽

Amylase Release ◽

Pancreatic Acini ◽

Group Ii ◽

P21 Activated Kinase ◽

Signal Cascades

p21-activated kinases (PAKs) are highly conserved serine/threonine protein kinases, which are divided into two groups: group-I (PAKs1–3) and group-II (PAKs4–6). In various tissues, Group-II PAKs play important roles in cytoskeletal dynamics and cell growth as well as neoplastic development/progression. However, little is known about Group-II PAK’s role in a number of physiological events, including their ability to be activated by gastrointestinal (GI) hormones/neurotransmitters/growth factors (GFs). We used rat pancreatic acini to explore the ability of GI hormones/neurotransmitters/GFs to activate Group-II-PAKs and the signaling cascades involved. Only PAK4 was detected in pancreatic acini. PAK4 was activated by endothelin, secretagogues-stimulating phospholipase C (bombesin, CCK-8, and carbachol), by pancreatic GFs (insulin, insulin-like growth factor 1, hepatocyte growth factor, epidermal growth factor, basic fibroblast growth factor, and platelet-derived growth factor), and by postreceptor stimulants (12-O-tetradecanoylphobol-13-acetate and A23187 ). CCK-8 activation of PAK4 required both high- and low-affinity CCK1-receptor state activation. It was reduced by PKC-, Src-, p44/42-, or p38-inhibition but not with phosphatidylinositol 3-kinase-inhibitors and only minimally by thapsigargin. A protein kinase D (PKD)-inhibitor completely inhibited CCK-8-stimulated PKD-activation; however, stimulated PAK4 phosphorylation was only inhibited by 60%, demonstrating that it is both PKD-dependent and PKD-independent. PF-3758309 and LCH-7749944, inhibitors of PAK4, decreased CCK-8-stimulated PAK4 activation but not PAK2 activation. Each inhibited ERK1/2 activation and amylase release induced by CCK-8 or bombesin. These results show that PAK4 has an important role in modulating signal cascades activated by a number of GI hormones/neurotransmitters/GFs that have been shown to mediate both physiological/pathological responses in acinar cells. Therefore, in addition to the extensive studies on PAK4 in pancreatic cancer, PAK4 should also be considered an important signaling molecule for pancreatic acinar physiological responses and, in the future, should be investigated for a possible role in pancreatic acinar pathophysiological responses, such as in pancreatitis. NEW & NOTEWORTHY This study demonstrates that the only Group-II p21-activated kinase (PAK) in rat pancreatic acinar cells is PAK4, and thus differs from islets/pancreatic cancer. Both gastrointestinal hormones/neurotransmitters stimulating PLC and pancreatic growth factors activate PAK4. With cholecystokinin (CCK), activation is PKC-dependent/-independent, requires both CCK1-R affinity states, Src, p42/44, and p38 activation. PAK4 activation is required for CCK-mediated p42/44 activation/amylase release. These results show PAK4 plays an important role in mediating CCK physiological signal cascades and suggest it may be a target in pancreatic acinar diseases besides cancer.

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Pancreas-specific aquaporin 12 null mice showed increased susceptibility to caerulein-induced acute pancreatitis

AJP Cell Physiology ◽

10.1152/ajpcell.00117.2009 ◽

2009 ◽

Vol 297 (6) ◽

pp. C1368-C1378 ◽

Cited By ~ 36

Author(s):

Eriko Ohta ◽

Tomohiro Itoh ◽

Tomomi Nemoto ◽

Jiro Kumagai ◽

Shigeru B. H. Ko ◽

...

Keyword(s):

Blood Chemistry ◽

Pancreatic Acinar Cells ◽

Imaging Method ◽

Pancreatic Acini ◽

Two Photon ◽

Pancreatic Fluid ◽

Photon Excitation ◽

Expression Studies ◽

Increased Susceptibility

Aquaporin 12 (AQP12) is the most recently identified member of the mammalian AQP family and is specifically expressed in pancreatic acinar cells. In vitro expression studies have revealed that AQP12 is localized at intracellular sites. To determine the physiological roles of AQP12 in the pancreas, we generated knockout mice for this gene (AQP12-KO). No obvious differences were observed under normal conditions between wild-type (WT) and AQP12-KO mice in terms of growth, blood chemistry, pancreatic fluid content, or histology. However, when we induced pancreatitis through the administration of a cholecystokinin-8 (CCK-8) analog, the AQP12-KO mice showed more severe pathological damage to this organ than WT mice. Furthermore, when we analyzed exocytosis in the pancreatic acini using a two-photon excitation imaging method, the results revealed larger exocytotic vesicles (vacuoles) in the acini of AQP12-KO mice at a high CCK-8 dose (100 nM). From these results, we conclude that AQP12 may function in the mechanisms that control the proper secretion of pancreatic fluid following rapid and intense stimulation.

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Sa1815 p21-Activated Kinase, PAK2, Is an Important Mediator of Numerous Signaling Cascades Mediating CCS Physiological and Pathophysiological Effects in Pancreatic Acinar Cells

Gastroenterology ◽

10.1016/s0016-5085(15)31131-8 ◽

2015 ◽

Vol 148 (4) ◽

pp. S-339

Author(s):

Bernardo Nuche-Berenguer ◽

Paola Moreno ◽

Robert T. Jensen

Keyword(s):

Acinar Cells ◽

Signaling Cascades ◽

Pancreatic Acinar Cells ◽

Important Mediator ◽

P21 Activated Kinase

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Su1096 ROLE OF P21-ACTIVATED KINASE, PAK4, IN ACTIVATION OF NUMEROUS IMPORTANT CELLULAR SIGNALING CASCADES IN PANCREATIC ACINAR CELLS

Gastroenterology ◽

10.1016/s0016-5085(20)31948-x ◽

2020 ◽

Vol 158 (6) ◽

pp. S-508

Author(s):

Irene Ramos ◽

Lingaku Lee ◽

Samuel A. Mantey ◽

Tatiana Iordanskaia ◽

Robert T. Jensen

Keyword(s):

Cellular Signaling ◽

Acinar Cells ◽

Signaling Cascades ◽

Pancreatic Acinar Cells ◽

P21 Activated Kinase

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PKCθ activation in pancreatic acinar cells by gastrointestinal hormones/neurotransmitters and growth factors is needed for stimulation of numerous important cellular signaling cascades

Biochimica et Biophysica Acta (BBA) - Molecular Cell Research ◽

10.1016/j.bbamcr.2011.07.007 ◽

2011 ◽

Vol 1813 (12) ◽

pp. 2145-2156 ◽

Cited By ~ 9

Author(s):

Veronica Sancho ◽

Marc J. Berna ◽

Michelle Thill ◽

R.T. Jensen

Keyword(s):

Growth Factors ◽

Cellular Signaling ◽

Gastrointestinal Hormones ◽

Acinar Cells ◽

Signaling Cascades ◽

Pancreatic Acinar Cells ◽

Stimulation Of

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Group II p21-activated kinase, PAK4, is needed for activation of focal adhesion kinases, MAPK, GSK3, and β-catenin in rat pancreatic acinar cells

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.00229.2019 ◽

2020 ◽

Vol 318 (3) ◽

pp. G490-G503

Author(s):

Irene Ramos-Álvarez ◽

Lingaku Lee ◽

Robert T. Jensen

Keyword(s):

Focal Adhesion ◽

Acinar Cell ◽

Cellular Signaling ◽

Acinar Cells ◽

Signaling Cascades ◽

Pancreatic Acinar Cells ◽

Adapter Proteins ◽

Group I ◽

Group Ii

PAK4 is the only member of the Group II p21-activated kinases (PAKs) present in rat pancreatic acinar cells and is activated by gastrointestinal hormones/neurotransmitters stimulating PLC/cAMP and by various pancreatic growth factors. However, little is known of the role of PAK4 activation in cellular signaling cascades in pancreatic acinar cells. In the present study, we examined the role of PAK4’s participation in five different cholecystokinin-8 (CCK-8)-stimulated signaling pathways (PI3K/Akt, MAPK, focal adhesion kinase, GSK3, and β-catenin), which mediate many of its physiological acinar-cell effects, as well as effects in pathophysiological conditions. To define PAK4’s role, the effect of two different PAK4 inhibitors, PF-3758309 and LCH-7749944, was examined under experimental conditions that only inhibited PAK4 activation and not activation of the other pancreatic PAK, Group I PAK2. The inhibitors’ effects on activation of these five signaling cascades by both physiological and pathophysiological concentrations of CCK, as well as by 12- O-tetradecanoylphobol-13-acetate (TPA), a PKC-activator, were examined. CCK/TPA activation of focal adhesion kinases(PYK2/p125FAK) and the accompanying adapter proteins (paxillin/p130CAS), Mek1/2, and p44/42, but not c-Raf or other MAPKs (JNK/p38), were mediated by PAK4. Activation of PI3K/Akt/p70s6K was independent of PAK4, whereas GSK3 and β-catenin stimulation was PAK4-dependent. These results, coupled with recent studies showing PAK4 is important in pancreatic fluid/electrolyte/enzyme secretion and acinar cell growth, show that PAK4 plays an important role in different cellular signaling cascades, which have been shown to mediate numerous physiological and pathophysiological processes in pancreatic acinar cells. NEW & NOTEWORTHY In pancreatic acinar cells, cholecystokinin (CCK) or 12- O-tetradecanoylphobol-13-acetate (TPA) activation of focal adhesion kinases (p125FAK,PYK2) and its accompanying adapter proteins, p130CAS/paxillin; Mek1/2, p44/42, GSK3, and β-catenin are mediated by PAK4. PI3K/Akt/p70s6K, c-Raf, JNK, or p38 pathways are independent of PAK4 activation.

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