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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 , , .
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 , .
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
Posttranslational modifications and interactions with several proteins are main
regulatory events affecting activity and stabilizing membrane expression of the
CFTR channel .
Cyclic adenosine monophosphate (cAMP)/ cAMP-dependent
protein kinase A (PKA) pathway is a dominant cascade which
affects CFTR channel activity .
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 , .
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
PKA-to-CFTR interaction . 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 .
Formation of a VIL2 (ezrin)/ E3KARP
(NHERF2)/ CFTR complex enhances the efficacy
of cAMP-mediated CFTR activation . 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
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 .
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) . AMPK
can also phosphorylate CFTR, but unlike
phosphorylation has a negative effect on CFTR channel
activity , , . 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 . AMP binds to and activates regulatory
AMPK gamma 1 (this isoform predominantly and functionally
associates with CFTR )
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 , , .
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 .
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 phosphorylates CFTR
and thus stabilizes expression of CFTR in the apical plasma
membrane of epithelial cells. , . 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 .
SH3/ankyrin domain gene 2 (SHANK2) inhibits
CFTR activity by breaching the
CFTR-EBP50 association and by
bringing PDE4D, which precludes
cAMP/ PKA signaling .
Dephosphorylation also affects activity of CFTR channel.
For instance, Protein phosphatase 2 (PP2A) and PP2C domain
containing protein phosphatases (PP2C) inhibit
CFTR activity , , .
In addition to posttranscriptional modifications, the binding partners (especially
PDZ-domain containing proteins) can also modulate CFTR
activity . These are EBP50, E3KARP (NHERF2),
PDZ domain containing (PDZK1) 1 and
E3KARP (NHERF2) functions as a scaffold (see above ). 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 , . 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 .
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
. Importantly, this process is independent of the
agonist-mediated cAMP / PKA
pathway . 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 .
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  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 , .
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 .
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 .
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