Structural and Functional Modulation of Inhibitory Synapses by GODZ-mediated Palmitoylation of GABAA Receptors

Open Access
Author:
Fang, Cheng
Graduate Program:
Biology
Degree:
Doctor of Philosophy
Document Type:
Dissertation
Date of Defense:
March 09, 2007
Committee Members:
  • Bernhard Luscher, Committee Chair
  • Douglas Cavener, Committee Member
  • Gong Chen, Committee Member
  • Erin Elizabeth Sheets, Committee Member
  • Wendy Hanna Rose, Committee Member
Keywords:
  • trafficking
  • synaptogenesis
  • inhibitory synapses
  • palmitoylation
  • GABAA receptors
  • DHHC-CRD proteins
Abstract:
GABAA receptors (GABAARs) mediate most of the fast inhibitory neurotransmission in the central nervous system. Clustering of GABAARs at postsynaptic sites is critical for the function of inhibitory synapses. Moreover, changes in synaptic receptor concentration are believed to contribute to functional plasticity of neurons. GABAARs are important for normal brain function as evidenced by their critical role in a wide range of neurological and neuropsychiatric diseases including epilepsy, anxiety, drug addiction, and diverse forms of mental retardation. Furthermore, GABAARs are a major site of action of clinically relevant CNS drugs, including the benzodiazepines, barbiturates, general anesthetics, and ethanol, as well as endogenous neurosteroids. Structurally, GABAARs represent heteropentameric ligand-gated Cl- channels, typically composed of 2?, 2? and 1?2 subunit. The ?2 subunit is specifically required for proper trafficking and clustering of the major subset of GABAARs that function at inhibitory synapses. GABAARs are subject to many posttranslational modifications, including palmitoylation, phosphorylation and ubiquitination. Of particular relevance for this thesis, palmitoylation is implicated in regulated trafficking of the ?2 subunit containing GABAARs to the plasma membrane and, by extension, in synaptic plasticity of inhibitory synapses. The ?2 subunit of GABAARs is palmitoylated at multiple Cys residues in the cytoplasmic loop region. Furthermore, GODZ has recently been isolated as a ?2 subunit interacting protein that can palmitoylate the ?2 subunit in vitro. The main objective of this doctoral thesis was to address the function of GODZ-mediated palmitoylation of the ?2 subunit in neurons. Here, I have shown that GODZ and its close paralog, SERZ-?, are the only enzymes among 23 DHHC family members of palmitoyl acyltransferases (PATs) that exhibit significant palmitoyltransferase activity for the ?2 subunit as a substrate in vitro. Moreover, GODZ and SERZ-? show indistinguishable expression patterns in brain as well as palmitoylation-dependent interaction with the ?2 subunit. Moreover, they form homo- and heteromultimers on expression in heterologous cells. Analyses in neurons indicate that GODZ is critically important for trafficking of GABAARs to synapses, for whole cell GABAergic inhibitory function and for normal amplitude and frequency of GABAergic miniature inhibitory synaptic currents. Indirectly, GODZ mediated palmitoylation in postsynaptic neurons is required for presynaptic GABAergic innervation. In contrast, GODZ is dispensable for AMPA receptor-mediated whole cell currents as well as miniature excitatory postsynaptic currents. Thus, GODZ-mediated palmitoylation of GABAARs is important selectively for structural and functional plasticity of inhibitory synapses.