Amphiphilic and hydrophilic forms of choline-O-acetyltransferase in cholinergic nerve endings of the Torpedo

154

It is well known that the regulation of choline acetyltransferase (ChAT) activity under physiological and pathological conditions is important for the development and neuronal activities of cholinergic systems involved in many fundamental brain functions. This review focuses on recent progress in understanding the regulation of ChAT at the levels of both the protein and the mRNA. A deficiency in ChAT activity has been reported for neurodegenerative conditions such as Alzheimer's disease, amyotrophic lateral sclerosis, and schizophrenia. Although a major feature of ChAT regulation is likely to involve the spatial and temporal control of transcription, regulation of expression can also be at the level of RNA processing, transport/ translocation, turnover, or translation. In addition, there is increasing evidence that ChAT might be regulated at the posttranslational level by compartmentation and/or covalent modification, i.e., phosphorylation, as well as noncovalent modification (protein‐protein interaction, etc.). Synaptic activity and the state of neuronal transmission may also involve the regulation of ChAT at different levels via both positive and negative feedback loops, as was demonstrated in the characterization of two ChAT mutant Drosophila strains. Clearly, identification of cholinergic‐specific elements and the characterization of the trans‐acting factors that bind to them represent an important area of future research. Equally important is research on the mechanisms governing ChAT as an enzymatic entity. The future should be an exciting time during which we look forward to the elucidation of the cholinergic signal and its regulation as well as the determination of the three‐dimensional structure of the enzyme.

Section: Intracellular Localization Of Chatmentioning

confidence: 99%

Choline Acetyltransferase: Celebrating Its Fiftieth Year

Hersh

1994

154

“…Various investigators have suggested that ChAT may be associated with neuronal plasma membranes both in brain tissue (Docherty et al, 1985(Docherty et al, , 1987 and in the electric organ of Torpedo (Eder-Colli and Amato, 1985;Eder-Colli et al, 1986). Thus, it appeared as though the ChAT and the occluded ACh in the 0.6 A4 sucrose fraction might be functionally similar to the ChAT and occluded ACh associated with the 0.4 M sucrose fraction.…”

Section: Subcsllulor Fractionsmentioning

confidence: 99%

“…In contrast, the detergent-soluble ChAT fraction, which is purported to be closely associated with vesicular and/or neuronal plasma membranes, may play a crit-ical role in the formation of that ACh released on depolarization (Benishin and Carroll, 1983;Eder-Colli and Amato, 1985;Eder-Colli et al, 1986). For example, the water-soluble ChAT fraction, which differs physically from the detergent-soluble ChAT fraction , may float freely in the cytosol and form that ACh released under resting conditions.…”

mentioning

confidence: 99%

Effect of 2‐(4‐Phenylpiperidino)cyclohexanol (AH 5183) on the Veratridine‐Induced Increase in Acetylcholine Synthesis by Rat Hippocampal Tissue

Carroll

Ivy

1988

The intent of this study was to determine whether the drug 2-(4-phenylpiperidino)cyclohexanol (AH 5183 or vesamicol) might inhibit the veratridine-induced increase in acetylcholine (ACh) synthesis by reducing the veratridine-induced activation of a detergent-soluble choline-O-acetyltransferase (EC 2.3.1.6; ChAT) fraction associated with a vesicle-bound store of ACh. When minces of rat hippocampal tissue were loaded with [14C]choline and subsequently depolarized with veratridine, an increase in the synthesis of [14C]ACh occurred that could be abolished by L-AH 5183 (75 nM). When minces were depolarized with veratridine in the presence of L-AH 5183 (75 nM), the depolarization-induced activation of a detergent-soluble ChAT fraction associated with a vesicle-bound store of ACh was blocked. Conversely, the veratridine-induced activation of a water-soluble ChAT fraction believed to be cytosolic was not. AH 5183 also blocked the repletion of the vesicle-bound store with newly synthesized ACh following veratridine-induced depletion of ACh, a result that appeared to be mediated by an effect on the synthesis of ACh at the vesicular surface. These results suggest that veratridine depolarization of rat hippocampal nerve terminals stimulates the synthesis of ACh by activating a detergent-soluble fraction of ChAT closely associated with synaptic vesicle release sites. ACh synthesis and transport at the vesicular surface may be influenced by a common AH 5183-sensitive regulatory protein.

“…This membrane-bound ChAT activity was mainly amphiphilic enzyme. Actually, ChAT was found to exist in a hydrophilic (88-90%) and in an amphiphilic form (10-12%) in the nerve terminals isolated from the Torpedo electric organ (Eder-Colli et al, 1986).…”

mentioning

confidence: 99%

“…Of a series of monoclonal antibodies we could develop against purified SPMs of the Torpedo (Eder-Colli and Amato, 1986), two were found to inhibit the activity of Torpedo ChAT (Eder-Colii et al, 1987). Recently, we found that these two antibodies had very similar properties and that most probably it was one and not two monoclonal antibodies we obtained that reacted against ChAT activity.…”

mentioning

confidence: 99%

A Monoclonal Antibody Raised Against Plasma Membrane of Cholinergic Nerve Terminals of the Torpedo Inhibits Choline Acetyltransferase Activity and Acetylcholine Release

Eder-Colli¹,

Briand²,

Pellegrinelli³

et al. 1989

Self Cite

Monoclonal antibodies were raised against the synaptosomal plasma membranes (SPMs) purified from the electric organ of the Torpedo. One antibody that reacts preferentially with SPMs rather than with other membrane fractions isolated from this tissue was previously found to inhibit hydrophilic and amphiphilic choline-O-acetyltransferase (ChAT) activity. On immunoblots of SPMs, this antibody recognizes two polypeptides of 135 and 66 kilodaltons that are related; the 66-kilodalton polypeptide appears to exist as a monomer and as a dimer in SPMs. The antibody was also able to inhibit the calcium-dependent release of acetylcholine in Torpedo synaptosomes without affecting the total neurotransmitter content. This inhibition was dependent on the antibody concentration and was observed when the release was elicited by either KCl depolarization or the calcium ionophore A23187; this suggests that inhibition was not mediated by a blockage of the depolarization-activated calcium influx. The inhibition could not be prevented by atropine, a result indicating that the antibody does not block release by mimicking the action of acetylcholine on presynaptic muscarinic autoreceptors. Thus, the antigen recognized by this antibody appeared to be involved in acetylcholine release; this antigen could be membrane-bound ChAT, another protein of the SPMs, or both.