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

Abstract

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.

Melatonin ameliorates endoplasmic reticulum stress in N2a neuroblastoma cell hypoxia-reoxygenation injury by activating the AMPK-Pak2 pathway

“…Several studies have reported the protective actions of Pak2 under various stress conditions. For example, in pancreatic acinar cells, ATP production is affected by Pak2 (Ramos-Alvarez et al 2018). In claudinlow breast cancer, Pak2 affects cell migration and epithelialto-mesenchymal transition (Matsunuma et al 2018).…”

“…A recent study identified Pak2 as a novel therapeutic target for ER stress response (Binder et al 2019). In addition, in pancreatic acinar cells, Pak2 activation is required for Na + -K + ATPase activation and pancreatic fluid secretion (Ramos-Alvarez et al 2018). Pak2 also modulates cancer migration (Matsunuma et al 2018) and endothelial cell death (Chelvanambi et al 2018).…”

The Role of p21-Activated Kinases in Cancer and Beyond: Where Are We Heading?

“…This activation of PAK2 stimulates several signal cascades including MAPK, FAK, and PI3K–Akt pathways to mediate physiological or pathophysiological responses related to the onset of pancreatitis, which can be inhibited by a PAK inhibitor, suggesting that PAK2 can act as a new therapeutic target for the treatment of pancreatic acinar diseases ( Nuche-Berenguer et al, 2016 ). Except the PAK2-mediated signaling pathways in pancreatic acinar cells, PAK4 can also be activated by stimulation of CREB, the VIP-/secretin-preferring receptors via EPAC, as well as regulate β-catenin signaling pathway to participate in Na + and K + -ATPase activation, mediating the pancreatic fluid secretion ( Supplementary Figure 3D ; Ramos-Alvarez and Jensen, 2018 ; Ramos-Alvarez et al, 2019 , 2020 ). These results together illustrate that PAKs play important roles in pancreatitis and pancreatic cancer growth, as well as enzyme secretion.…”

Na+/K+-ATPase Revisited: On Its Mechanism of Action, Role in Cancer, and Activity Modulation

“…The first one involves the activation of protein kinase A, while the second one activates the exchange of protein directly activated by cAMP (EPAC) 1 and 2. In both pathways, the next step is switching on the p21-activated kinase 4, which in turn triggers NKA [ 161 ]. Apart from cAMP, another endogenous NKA modulator is epinephrine (adrenaline), which induces the production of prostaglandin E2 (PGE2).…”