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The gamma2 subunit of GABA(A) receptors is a substrate for palmitoylation by GODZ.

ABSTRACT: The neurotransmitter GABA activates heteropentameric GABA(A) receptors, which are composed mostly of alpha, beta, and gamma2 subunits. Regulated membrane trafficking and subcellular targeting of GABA(A) receptors is important for determining the efficacy of GABAergic inhibitory function. Of special interest is the gamma2 subunit, which is mostly dispensable for assembly and membrane insertion of functional receptors but essential for accumulation of GABA(A) receptors at synapses. In a search for novel receptor trafficking proteins, we have used the SOS-recruitment system and isolated a Golgi-specific DHHC zinc finger protein (GODZ) as a novel gamma2 subunit-interacting protein. GODZ is a member of the superfamily of DHHC cysteine-rich domain (DHHC-CRD) polytopic membrane proteins shown recently in yeast to represent palmitoyltransferases. GODZ mRNA is found in many tissues; however, in brain the protein is detected in neurons only and highly concentrated and asymmetrically distributed in the Golgi complex. GODZ interacts with a cysteine-rich 14-amino acid domain conserved specifically in the large cytoplasmic loop of gamma1-3 subunits but not in other GABA(A) receptor subunits. Coexpression of GODZ and GABA(A) receptors in heterologous cells results in palmitoylation of the gamma2 subunit in a cytoplasmic loop domain-dependent manner. Neuronal GABA(A) receptors are similarly palmitoylated. Thus, GODZ-mediated palmitoylation represents a novel posttranslational modification that is selective for gamma subunit-containing GABA(A) receptor subtypes, a mechanism that is likely to be important for regulated trafficking of these receptors in the secretory pathway.

SUBMITTER: Keller CA

PROVIDER: S-EPMC2366890 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BACKGROUND AND PURPOSE: Heterologous expression of alpha1, beta2 and gamma2S(gamma1) subunits produces a mixed population of GABA(A) receptors containing alpha1beta2 or alpha1beta2gamma2S(gamma1) subunits. GABA sensitivity (lower in receptors containing gamma1 or gamma2S subunits) and the potentiation of GABA-activated chloride currents (I(GABA)) by benzodiazepines (BZDs) are dependent on gamma2S(gamma1) incorporation. A variable gamma subunit incorporation may affect the estimation of I(GABA) potentiation by BZDs. We propose an approach for estimation of BZD efficiency that accounts for mixed population of alpha1beta2 and alpha1beta2gamma2S(gamma1) receptors. EXPERIMENTAL APPROACH: We investigated the relation between GABA sensitivity (EC50) and BZD modulation by analysing triazolam-, clotiazepam- and midazolam-induced potentiation of I(GABA) in Xenopus oocytes under two-microelectrode voltage clamp. KEY RESULTS: Plotting EC50 versus BZD-induced shifts of GABA concentration-response curves (DeltaEC50(BZD)) of oocytes injected with different amounts of alpha1, beta2 and gamma2S(gamma1) cRNA (1:1:1-1:1:10) revealed a linear regression between gamma2S(gamma1)-mediated reduction of GABA sensitivity (EC50) and DeltaEC50(BZD). The slope factors of the regression were always higher for oocytes expressing alpha1beta2gamma1 subunit receptors (1.8 +/- 0.1 (triazolam), 1.6 +/- 0.1 (clotiazepam), 2.3 +/- 0.2 (midazolam)) than for oocytes expressing alpha1beta2gamma2S receptors (1.4 +/- 0.1 (triazolam), 1.4 +/- 0.1 (clotiazepam), 1.3 +/- 0.1 (midazolam)). Mutant GABA(A) receptors (alpha1beta2-R207Cgamma2S) with lower GABA sensitivity showed higher drug efficiencies (slope factors=1.1 +/- 0.1 (triazolam), 1.1 +/- 0.1 (clotiazepam), 1.2 +/- 0.1 (midazolam)). CONCLUSIONS AND IMPLICATIONS: Regression analysis enabled the estimation of BZD efficiency when variable mixtures of alpha1beta2 and alpha1beta2gamma2S(gamma1) receptors are expressed and provided new insights into the gamma2S(gamma1) dependency of BZD action.

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The gamma2 subunit of GABA(A) receptors is a substrate for palmitoylation by GODZ.

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