Regulation of CFTR activity (norm and CF)

Regulation of CFTR activity (norm and CF)

The cystic fibrosis transmembrane conductance regulator (CFTR) is a member of the ATP-binding cassette (ABC) transporter superfamily. It acts in apical part of the epithelial cells as a plasma-membrane, cyclic AMP-activated chloride anion, bicarbonate anion and glutathione channel [1], [2], [3]. CFTR is required for cell surface water-salt homeostasis and normal function of epithelia lining the airways, intestinal tract, ducts in the pancreas, salivary and sweat glands, liver and others [3], [4].

CFTR is an ATP-dependent membrane transporter which is activated by directly binding to ATP. Opening of the CFTR is initiated by ATP binding at the NBD2 site of this channel [5], [6].

Posttranslational modifications and interactions with several proteins are main regulatory events affecting activity and stabilizing membrane expression of the CFTR channel [4].

Cyclic adenosine monophosphate (cAMP)/ cAMP-dependent protein kinase A (PKA) pathway is a dominant cascade which affects CFTR channel activity [4]. Adenosine is a mediator which activates CFTR channel via cAMP/ PKA. Activation of the Adenosine A2B receptor by physiological ligands such as Adenosine leads to stimulation of Adenylate cyclase by G-protein alpha-s leading to an increase in concentration of highly compartmentalized cAMP, and subsequent activation of the PKA [7], [8].

Phosphorylation of CFTR by PKA-cat is mediated by PRKAR2A which is linked physically and functionally to CFTR by a Villin 2 (VIL2 (ezrin)). The latter serves as an anchoring protein for PKA-cat-mediated phosphorylation of CFTR. Anchoring protein VIL2 (ezrin) promotes PKA-to-CFTR interaction [9]. Moreover VIL2 (ezrin) itself exists in a complex with CFTR. This interaction is mediated by Solute carrier family 9 member 3 regulator 2 (E3KARP (NHERF2)) - a PDZ-containing binding partner of CFTR [10]. Formation of a VIL2 (ezrin)/ E3KARP (NHERF2)/ CFTR complex enhances the efficacy of cAMP-mediated CFTR activation [10]. In addition, Protein phosphatase 3 catalytic subunit (Calcineurin A)/ Annexin A2 (Annexin II)/ S100 calcium binding protein A10 (S100A10) complex participate in PKA-dependent CFTR activation [11].

It is shown, that Annexin A5 (Annexin V) is necessary for normal CFTR chloride channel activity as well, but exactly mechanism it acting is unknown [12].

Interestingly, activation of PKA by Adenosine may also increase the activity of Phosphodiesterase 4D (PDE4D) leading to attenuation of the cAMP signal. The by-product of cAMP degradation - Adenosine monophosphate (AMP) can activate AMP-activated protein kinase (AMPK) [13]. AMPK can also phosphorylate CFTR, but unlike PKA, AMPK-dependent phosphorylation has a negative effect on CFTR channel activity [14], [15], [16]. The exact molecular events leading to AMPK-dependent phosphorylation of CFTR are still elusive. Most likely, it starts with activation of PDE4D by PKA-cat followed by conversion of cAMP to AMP [13]. AMP binds to and activates regulatory AMPK gamma 1 (this isoform predominantly and functionally associates with CFTR [14]) and initiates formation of a complex consisting of regulatory AMPK gamma and beta subunits and catalytic AMPK alpha 1 subunit. In turn, AMPK alpha 1 subunit binds to CFTR. This interaction might be essential for AMPK mediated phosphorylation of CFTR, which reduces chloride anion secretion by inhibiting channel activity without affecting the number of CFTR channels in the plasma membrane [14], [15], [16].

In addition, enterotoxins released by Vibrio cholerae (cholera toxin) and Escherichia coli (heat stable enterotoxin) activate intracellular cAMP/ PKA and cGMP/ Protein kinase G 2 and signal CFTR on the apical plasma membrane [17].

CFTR membrane expression is also regulated by the Tubulin/ Solute carrier family 9 member 3 regulator 1 (EBP50)/ Guanine nucleotide binding protein beta polypeptide 2-like 1 (RACK1)/ Protein kinase C epsilon (PKC-epsilon) pathway. PKC-epsilon phosphorylates CFTR and thus stabilizes expression of CFTR in the apical plasma membrane of epithelial cells. [18], [19]. Apparently, constitutive phosphorylation by PKC-epsilon is essential for the acute activation of CFTR by PKA-cat, since phosphorylation by PKA-cat alone is not a sufficient stimulus to open the CFTR [20].

SH3/ankyrin domain gene 2 (SHANK2) inhibits CFTR activity by breaching the CFTR-EBP50 association and by bringing PDE4D, which precludes cAMP/ PKA signaling [21].

Dephosphorylation also affects activity of CFTR channel. For instance, Protein phosphatase 2 (PP2A) and PP2C domain containing protein phosphatases (PP2C) inhibit CFTR activity [22], [23], [24].

In addition to posttranscriptional modifications, the binding partners (especially PDZ-domain containing proteins) can also modulate CFTR activity [4]. These are EBP50, E3KARP (NHERF2), PDZ domain containing (PDZK1) 1 and others.

E3KARP (NHERF2) functions as a scaffold (see above [10]). PDZK1 is capable of linking CFTR molecules to form dimers. In this dimeric form, CFTR channel activity is enhanced. Disrupting PDZK1/ CFTR complex abrogates the functional coupling of cAMP transporter activity to CFTR function [25], [26]. EBP50, which exists in a complex with CFTR at the apical surface of epithelial cells, is a main PDZ-domain containing binding partner which positively regulates CFTR channel activity [4].

EBP50 may stimulate CFTR expression on apical membrane in receptor-dependent fashion - mainly with Beta-2 adrenergic receptor. Beta-2 adrenergic receptor and CFTR are physically and functionally coupled into a macromolecular signaling complex via interactions with EBP50 [27]. Importantly, this process is independent of the agonist-mediated cAMP / PKA pathway [28]. On the other hand, PKA-cat-mediated phosphorylation of CFTR strongly inhibits formation of the macromolecular complex consisting of Beta-2 adrenergic receptor/ EBP50 / CFTR [27]. Functional consequences of the disruption of this complex are elusive.

Copper metabolism domain containing 1 (COMMD1) (Drevillion, L et al., The 21st annual north American cystic fibrosis conference, California, 2007), Filamin A and Filamin B [29] stabilize expression of CFTR in the apical plasma membrane.

In addiction to positive regulation of CFTR by PDZ-containing scaffold proteins other binding partners such as Synaptosomal-associated protein 23kDa (SNAP-23) - Syntaxin 1A complex can sterically interfere with CFTR. This results in a decrease of channel activity, although inhibitory influences of Syntaxin binding protein 1 (MUNC18) can be diminished by its binding to Syntaxin 1A [30], [31].

In addition, Endothelial differentiation lysophosphatidic acid G-protein-coupled receptor 4 (EDG4) activated by a Lysophosphatidic acid rapidly inhibits CFTR channel activity through G-protein alpha-i family by suppressing PKA-cat-mediated activation of CFTR. EDG4 is most typical for gut but EDG4/ E3KARP (NHERF2)/ CFTR macromolecular complex may be form in different cell as HT29-CL19A (colonic epithelial cells) as Calu-3 (airway serous gland epithelial cells). And so it is possibly, that EDG4 participates in CFTR regulation in airway cells too [32].

The most common CFTR mutation is loss of a Phe residue at position 508 (deltaF508-CFTR). Majority of regulators-to-CFTR are equal interactions for wtCFTR and deltaF508-CFTR. One of the exclusion is a Casein kinase II. Casein kinase II associates with and phosphorylates wtCFTR but not deltaF508-CFTR. This interaction activates CFTR-dependent chloride transport [33].

References:

1. Kogan I, Ramjeesingh M, Li C, Kidd JF, Wang Y, Leslie EM, Cole SP, Bear CE
   CFTR directly mediates nucleotide-regulated glutathione flux. The EMBO journal 2003 May 1;22(9):1981-9
2. Chan HC, Shi QX, Zhou CX, Wang XF, Xu WM, Chen WY, Chen AJ, Ni Y, Yuan YY
   Critical role of CFTR in uterine bicarbonate secretion and the fertilizing capacity of sperm. Molecular and cellular endocrinology 2006 May 16;250(1-2):106-13
3. Gadsby DC, Vergani P, Csanady L
   The ABC protein turned chloride channel whose failure causes cystic fibrosis. Nature 2006 Mar 23;440(7083):477-83
4. Guggino WB, Stanton BA
   New insights into cystic fibrosis: molecular switches that regulate CFTR. Nature reviews. Molecular cell biology 2006 Jun;7(6):426-36
5. Zhou Z, Wang X, Liu HY, Zou X, Li M, Hwang TC
   The two ATP binding sites of cystic fibrosis transmembrane conductance regulator (CFTR) play distinct roles in gating kinetics and energetics. The Journal of general physiology 2006 Oct;128(4):413-22
6. Aleksandrov AA, Aleksandrov LA, Riordan JR
   CFTR (ABCC7) is a hydrolyzable-ligand-gated channel. Pflugers Archiv : European journal of physiology 2007 Feb;453(5):693-702
7. Clancy JP, Ruiz FE, Sorscher EJ
   Adenosine and its nucleotides activate wild-type and R117H CFTR through an A2B receptor-coupled pathway. The American journal of physiology 1999 Feb;276(2 Pt 1):C361-9
8. Huang P, Lazarowski ER, Tarran R, Milgram SL, Boucher RC, Stutts MJ
   Compartmentalized autocrine signaling to cystic fibrosis transmembrane conductance regulator at the apical membrane of airway epithelial cells. Proceedings of the National Academy of Sciences of the United States of America 2001 Nov 20;98(24):14120-5
9. Sun F, Hug MJ, Bradbury NA, Frizzell RA
   Protein kinase A associates with cystic fibrosis transmembrane conductance regulator via an interaction with ezrin. The Journal of biological chemistry 2000 May 12;275(19):14360-6
10. Sun F, Hug MJ, Lewarchik CM, Yun CH, Bradbury NA, Frizzell RA
    E3KARP mediates the association of ezrin and protein kinase A with the cystic fibrosis transmembrane conductance regulator in airway cells. The Journal of biological chemistry 2000 Sep 22;275(38):29539-46
11. Borthwick LA, McGaw J, Conner G, Taylor CJ, Gerke V, Mehta A, Robson L, Muimo R
    The formation of the cAMP/protein kinase A-dependent annexin 2-S100A10 complex with cystic fibrosis conductance regulator protein (CFTR) regulates CFTR channel function. Molecular biology of the cell 2007 Sep;18(9):3388-97
12. Trouve P, Le Drevo MA, Kerbiriou M, Friocourt G, Fichou Y, Gillet D, Ferec C
    Annexin V is directly involved in cystic fibrosis transmembrane conductance regulator's chloride channel function. Biochimica et biophysica acta 2007 Oct;1772(10):1121-33
13. Barnes AP, Livera G, Huang P, Sun C, O'Neal WK, Conti M, Stutts MJ, Milgram SL
    Phosphodiesterase 4D forms a cAMP diffusion barrier at the apical membrane of the airway epithelium. The Journal of biological chemistry 2005 Mar 4;280(9):7997-8003
14. Hallows KR, Raghuram V, Kemp BE, Witters LA, Foskett JK
    Inhibition of cystic fibrosis transmembrane conductance regulator by novel interaction with the metabolic sensor AMP-activated protein kinase. The Journal of clinical investigation 2000 Jun;105(12):1711-21
15. Hallows KR, McCane JE, Kemp BE, Witters LA, Foskett JK
    Regulation of channel gating by AMP-activated protein kinase modulates cystic fibrosis transmembrane conductance regulator activity in lung submucosal cells. The Journal of biological chemistry 2003 Jan 10;278(2):998-1004
16. Hallows KR, Kobinger GP, Wilson JM, Witters LA, Foskett JK
    Physiological modulation of CFTR activity by AMP-activated protein kinase in polarized T84 cells. American journal of physiology. Cell physiology 2003 May;284(5):C1297-308
17. Golin-Bisello F, Bradbury N, Ameen N
    STa and cGMP stimulate CFTR translocation to the surface of villus enterocytes in rat jejunum and is regulated by protein kinase G. American journal of physiology. Cell physiology 2005 Sep;289(3):C708-16
18. Liedtke CM, Yun CH, Kyle N, Wang D
    Protein kinase C epsilon-dependent regulation of cystic fibrosis transmembrane regulator involves binding to a receptor for activated C kinase (RACK1) and RACK1 binding to Na+/H+ exchange regulatory factor. The Journal of biological chemistry 2002 Jun 21;277(25):22925-33
19. Auerbach M, Liedtke CM
    Role of the scaffold protein RACK1 in apical expression of CFTR. American journal of physiology. Cell physiology 2007 Jul;293(1):C294-304
20. Jia Y, Mathews CJ, Hanrahan JW
    Phosphorylation by protein kinase C is required for acute activation of cystic fibrosis transmembrane conductance regulator by protein kinase A. The Journal of biological chemistry 1997 Feb 21;272(8):4978-84
21. Lee JH, Richter W, Namkung W, Kim KH, Kim E, Conti M, Lee MG
    Dynamic regulation of cystic fibrosis transmembrane conductance regulator by competitive interactions of molecular adaptors. The Journal of biological chemistry 2007 Apr 6;282(14):10414-22
22. Luo J, Pato MD, Riordan JR, Hanrahan JW
    Differential regulation of single CFTR channels by PP2C, PP2A, and other phosphatases. The American journal of physiology 1998 May;274(5 Pt 1):C1397-410
23. Vastiau A, Cao L, Jaspers M, Owsianik G, Janssens V, Cuppens H, Goris J, Nilius B, Cassiman JJ
    Interaction of the protein phosphatase 2A with the regulatory domain of the cystic fibrosis transmembrane conductance regulator channel. FEBS letters 2005 Jun 20;579(16):3392-6
24. Thelin WR, Kesimer M, Tarran R, Kreda SM, Grubb BR, Sheehan JK, Stutts MJ, Milgram SL
    The cystic fibrosis transmembrane conductance regulator is regulated by a direct interaction with the protein phosphatase 2A. The Journal of biological chemistry 2005 Dec 16;280(50):41512-20
25. Wang S, Yue H, Derin RB, Guggino WB, Li M
    Accessory protein facilitated CFTR-CFTR interaction, a molecular mechanism to potentiate the chloride channel activity. Cell 2000 Sep 29;103(1):169-79
26. Li C, Krishnamurthy PC, Penmatsa H, Marrs KL, Wang XQ, Zaccolo M, Jalink K, Li M, Nelson DJ, Schuetz JD, Naren AP
    Spatiotemporal coupling of cAMP transporter to CFTR chloride channel function in the gut epithelia. Cell 2007 Nov 30;131(5):940-51
27. Naren AP, Cobb B, Li C, Roy K, Nelson D, Heda GD, Liao J, Kirk KL, Sorscher EJ, Hanrahan J, Clancy JP
    A macromolecular complex of beta 2 adrenergic receptor, CFTR, and ezrin/radixin/moesin-binding phosphoprotein 50 is regulated by PKA. Proceedings of the National Academy of Sciences of the United States of America 2003 Jan 7;100(1):342-6
28. Taouil K, Hinnrasky J, Hologne C, Corlieu P, Klossek JM, Puchelle E
    Stimulation of beta 2-adrenergic receptor increases cystic fibrosis transmembrane conductance regulator expression in human airway epithelial cells through a cAMP/protein kinase A-independent pathway. The Journal of biological chemistry 2003 May 9;278(19):17320-7
29. Thelin WR, Chen Y, Gentzsch M, Kreda SM, Sallee JL, Scarlett CO, Borchers CH, Jacobson K, Stutts MJ, Milgram SL
    Direct interaction with filamins modulates the stability and plasma membrane expression of CFTR. The Journal of clinical investigation 2007 Feb;117(2):364-74
30. Chang SY, Di A, Naren AP, Palfrey HC, Kirk KL, Nelson DJ
    Mechanisms of CFTR regulation by syntaxin 1A and PKA. Journal of cell science 2002 Feb 15;115(Pt 4):783-91
31. Cormet-Boyaka E, Di A, Chang SY, Naren AP, Tousson A, Nelson DJ, Kirk KL
    CFTR chloride channels are regulated by a SNAP-23/syntaxin 1A complex. Proceedings of the National Academy of Sciences of the United States of America 2002 Sep 17;99(19):12477-82
32. Li C, Dandridge KS, Di A, Marrs KL, Harris EL, Roy K, Jackson JS, Makarova NV, Fujiwara Y, Farrar PL, Nelson DJ, Tigyi GJ, Naren AP
    Lysophosphatidic acid inhibits cholera toxin-induced secretory diarrhea through CFTR-dependent protein interactions. The Journal of experimental medicine 2005 Oct 3;202(7):975-86
33. Treharne KJ, Crawford RM, Xu Z, Chen JH, Best OG, Schulte EA, Gruenert DC, Wilson SM, Sheppard DN, Kunzelmann K, Mehta A
    Protein kinase CK2, cystic fibrosis transmembrane conductance regulator, and the deltaF508 mutation: F508 deletion disrupts a kinase-binding site. The Journal of biological chemistry 2007 Apr 6;282(14):10804-13