BACKGROUND: GABA A receptor deficits that are induced by global or forebrain-specific heterozygous inactivation of the γ2 subunit gene in mouse embryos result in behavior indicative of trait anxiety and depressive states. By contrast, a comparable deficit that is delayed to adolescence is without these behavioral consequences. Here we characterized γ2-deficient mice with respect to HPA axis abnormalities and antidepressant drug responses.
Edited by F. Anne StephensonThe ␥2 subunit of GABA type A receptors (GABA A Rs) is thought to be subject to palmitoylation by both Golgi-associated DHHC-type zinc finger protein (GODZ; also known as DHHC3) and its paralog Sertoli cell gene with a zinc finger domain- (SERZ-; DHHC7) based on overexpression of enzymes and substrates in heterologous cells. Here we have further investigated the substrate specificity of these enzymes by characterization of GODZ and SERZ- knock-out (KO) mice as well as double KO (DKO) neurons. Palmitoylation of ␥2 and a second substrate, growth-associated protein of 43 kDa, that is independently implicated in trafficking of GABA A Rs was significantly reduced in brain of GODZ KO versus wild-type (WT) mice but unaltered in SERZ- KO mice. Accumulation of GABA A Rs at synapses, GABAergic innervation, and synaptic function were reduced in GODZ KO and DKO neurons to a similar extent, indicating that SERZ- does not contribute to palmitoylation or trafficking of GABA A Rs even in the absence of GODZ. Notably, these effects were seen only when mutant neurons were grown in competition with WT neurons, thereby mimicking conditions of shRNA-transfected neurons previously used to characterize GODZ. However, GABA-evoked whole-cell currents of DKO neurons and the GABA A R cell surface expression in DKO neurons and GODZ or SERZ- KO brain slices were unaltered, indicating that GODZ-mediated palmitoylation selectively controls the pool of receptors at synapses. The different substrate specificities of GODZ and SERZ- in vivo were correlated with their differential localization to cis-versus trans-Golgi compartment, a mechanism that was compromised by overexpression of GODZ.S-Palmitoylation is an important posttranslational modification that involves the addition of the 16-carbon fatty acid chain palmitate via a thioester bond to Cys residues (1). This modification in turn can alter a protein's conformational state, membrane association, and complex formation as well as its susceptibility to other posttranslational modifications (2-4). Global analyses of rat synaptosomal fractions led to the discovery of nearly 300 palmitoylated synaptosomal protein candidates that illustrate the particular importance of palmitoylation in regulating the function of neuronal synapses (5).In mammalian cells, the palmitoylation reaction is catalyzed principally by a super gene family of 23 palmitoyl acyltransferases containing a DHHC motif in a cysteine-rich domain (DHHC-CRD) 3 that are both essential for enzyme function (6 -10). The mechanisms that determine substrate specificity of PATs remain poorly understood, although some specificity is observed upon overexpression of substrates and PATs in heterologous cells. However, increasing evidence suggests that the substrate specificity of DHHC-type PATs in vivo is much more stringent than in vitro. For example, the postsynaptic density (PSD) protein of 95-kDa (PSD-95) can be palmitoylated in heterologous cells by at least five members of the DHHC family of PATs (11), w...
Mice that were rendered heterozygous for the γ2 subunit of GABAA receptors (γ2+/− mice) have been characterized extensively as a model for major depressive disorder. The phenotype of these mice includes behavior indicative of heightened anxiety, despair, and anhedonia, as well as defects in hippocampus-dependent pattern separation, HPA axis hyperactivity and increased responsiveness to antidepressant drugs. The γ2+/− model thereby provides strong support for the GABAergic deficit hypothesis of major depressive disorder. Here we show that γ2+/− mice additionally exhibit specific defects in late stage survival of adult-born hippocampal granule cells, including reduced complexity of dendritic arbors and impaired maturation of synaptic spines. Moreover, cortical γ2+/− neurons cultured in vitro show marked deficits in GABAergic innervation selectively when grown under competitive conditions that may mimic the environment of adult-born hippocampal granule cells. Finally, brain extracts of γ2+/− mice show a numerical but insignificant trend (p = 0.06) for transiently reduced expression of brain derived neurotrophic factor (BDNF) at three weeks of age, which might contribute to the previously reported developmental origin of the behavioral phenotype of γ2+/− mice. The data indicate increasing congruence of the GABAergic, glutamatergic, stress-based and neurotrophic deficit hypotheses of major depressive disorder.
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