Acetylcholinesterase (AChE) is known to be vital in life maintenance for
terminating cholinergic transmission. By alternative splicing, AChE is synthesized in
four different isoforms, and which are readthrough (AChE
R), hydrophobic (AChE
H),
soluble (AChE
S) and tailed (AChE
T) forms. In the brain and muscles, AChE
T is the
major form and plays an important role in nervous system. The comprehensive
function of AChE
T depends on a tight interaction between its C-terminal peptide with
the anchoring protein collagen tail (ColQ) to form asymmetric enzyme or proline-rich
membrane anchor (PRiMA) to form tetrameric membrane bond enzyme. The
expression of AChE subunits in different tissues has been studied in detail, and
transcriptional regulation appears to be the major feature in controll...[
Read more ]
Acetylcholinesterase (AChE) is known to be vital in life maintenance for
terminating cholinergic transmission. By alternative splicing, AChE is synthesized in
four different isoforms, and which are readthrough (AChE
R), hydrophobic (AChE
H),
soluble (AChE
S) and tailed (AChE
T) forms. In the brain and muscles, AChE
T is the
major form and plays an important role in nervous system. The comprehensive
function of AChE
T depends on a tight interaction between its C-terminal peptide with
the anchoring protein collagen tail (ColQ) to form asymmetric enzyme or proline-rich
membrane anchor (PRiMA) to form tetrameric membrane bond enzyme. The
expression of AChE subunits in different tissues has been studied in detail, and
transcriptional regulation appears to be the major feature in controlling the
expression associated with various cellular activities. The binding sites of different
transcriptional factors on mammalian AChE gene have proposed various functions of
this enzyme in specific cell type. However, the regulation of AChE by cAMP response
element-binding protein (CREB) and nuclear factor kappa-light-chain-enhancer of
activated B cells (NFκB) have not been fully identified, particularly the regulation
mediated by these factors in different cell types.
Genistein, 4’,5,7-trihydroxyisoflavone, a major isoflavone in soybean, is known
as phytoestrogen having known benefit to brain functions. In cultured PC12 cells,
application of genistein significantly induced the expression of neurofilaments,
markers for neuronal differentiation. In parallel, the expression of tetrameric form of
proline-rich membrane anchor (PRiMA)-linked AChE (G
4 AChE), a key isoform in the
brain, was induced in a dose-dependent manner: this induction included the
associated protein PRiMA. The genistein-induced AChE expression was fully blocked
by pre-treatment of H89 (an inhibitor of protein kinase A, PKA) and G15 (a selective
G protein-coupled receptor 30 (GPR 30) antagonist) in the cultures, suggesting a
direct involvement of a membrane-bound estrogen receptor, named as GPR 30 in
the cultures. In parallel, the estrogen-induced activation of GPR 30 induced AChE
expression in a dose-dependent manner. The genistein/estrogen-induced the
phosphorylation of CREB, which subsequently triggered a cyclic AMP responding
element (CRE) located on the ACHE gene promoter. The binding of this CRE site by
CREB induced ACHE gene transcription. Thus, the activation of GPR 30 could be
one way for estrogen or flavonoids, possessing estrogenic properties, to enhance
cholinergic functions in the brain via CREB transcription factor.
Protein assembly of oligomeric AChE could be affected by chaperons. Many
AChE inhibitors (AChEIs) could act as chemical chaperons. In cultured
neuroblastoma (NG108-15) or cortical neurons, application of AChEIs, including
tacrine (Cognex), rivastigmine (Exelon), but not donepezil (Aricept), caused a
beatable amount of AChE in NG108-15 cell surface by immunofluorescence staining.
Subcellular fractionation showed an accumulation of the unfolded AChE being
retained in endoplasmic reticulum (ER) fraction: the AChEI-bound enzyme was not
able to transport to Golgi/plasma membrane fraction. The AChEI-bound enzyme
retained intracellularly induced a result of ER stress, as indicated by an increased
mRNA of immunoglobulin heavy chain-binding protein (BiP) and CCAAT-enhancer-binding
protein homologous protein (CHOP) at ~2.5 folds and ~5 folds in NG108-15
cultures, respectively. The AChEI-induced ER stress resulted with cell apoptosis,
indicated by increased expression of apoptotic marker (cleaved caspase 3),
apoptotic cell number and mitochondrial membrane potential (MMP) in a time-dependent
manner. In parallel, the ER stress activated an activation of cAMP
signaling with an induction of CREB phosphorylation and CRE responses, which
thereafter upregulated the expression of AChE.
In immune cells, the role of AChE in cholinergic anti-inflammatory pathway
(CAP) has been demonstrated. The functions of CAP were confirmed in RAW 264.7,
a macrophage cell. Application of ACh suppressed lipopolysaccharide (LPS)-induced
TNF-α release, which was abolished by overexpression of PRiMA-linked G
4 AChE. In
parallel, the application of AChEI showed anti-inflammatory action through
suppressing TNF-α release. Activation of α7 nicotinic AChR (nAChR) by ACh or
PHA-543613, an α7 nAChR agonist, inhibited LPS-induced cell migration confirmed
by transwell® assay and pro-migratory gene expression. This suppression effect of
RAW 264.7 cell migration was enhanced by application of AChEI.
Immunofluorescence staining showed LPS significantly induced NFκB accumulation
in nuclear by ~2 folds, which can be significantly reduced by co-treatment with
AChEI. These results suggested the regulatory role of AChE in immune responses
could be mediated by α7 nAChR. Moreover, overexpression of NFκB cDNAs induced
the protein and activity of AChE in a time-dependent manner having the maximum
induction at ~2 folds. The induced AChE form was PRiMA-linked G
4 enzyme. LPS
application induced the human AChE promotor activity, showing no effects on the
promotor having a mutation of NFκB site (i.e. pAChE
ΔNFκB-Luc). The binding of NFκB
with AChE promotor was upregulated by the applied LPS, as detected by CHIP
assay. Thus, the NFκB transcriptional factor could regulate AChE expression.
The present study demonstrated the significance of transcriptional regulation
of AChE via CRE and NFκB binding sites on the AChE gene. In neurons, AChE could
be upregulated by estrogen/phytoestrogen and chemical chaperon-induced ER
stress via CREB transcriptional factor. In immune system, the regulatory role of
AChE in immune actions via α7 nAChR has been showed. Conversely, the
inflammation-induced AChE expression could be mediated by the binding site of
NFκB onto the promotor. These transcription analyses of AChE gene therefore
suggest the possible non-classical functions of AChE in different cell types.
Post a Comment