frameshift variants impair GABAA receptor proteostasis.

The gamma-aminobutyric acid type A receptor (GABA R) is the most common inhibitory neurotransmitter-gated ion channel in the central nervous system. Pathogenic variants in genes encoding GABA R subunits can cause receptor dysfunction and lead to genetic epilepsy. Frameshift variants in these genes can result in a premature termination codon, producing truncated receptor subunit variants.

Pharmacological chaperones restore proteostasis of epilepsy-associated GABA receptor variants.

Recent advances in genetic diagnosis identified variants in genes encoding GABA receptors as causative for genetic epilepsy. Here, we selected eight disease-associated variants in the α1 subunit of GABA receptors causing mild to severe clinical phenotypes and showed that they are loss of function, mainly by reducing the folding and surface trafficking of the α1 protein. Furthermore, we sought client protein-specific pharmacological chaperones to restore the function of pathogenic receptors.

Hsp47 promotes biogenesis of multi-subunit neuroreceptors in the endoplasmic reticulum.

Protein homeostasis (proteostasis) deficiency is an important contributing factor to neurological and metabolic diseases. However, how the proteostasis network orchestrates the folding and assembly of multi-subunit membrane proteins is poorly understood. Previous proteomics studies identified Hsp47 (Gene: ), a heat shock protein in the endoplasmic reticulum lumen, as the most enriched interacting chaperone for gamma-aminobutyric acid type A (GABA) receptors.

Pathogenicity Prediction of GABA Receptor Missense Variants.

Variants in the genes encoding the subunits of gamma-aminobutyric acid type A (GABA) receptors are associated with epilepsy. To date, over 1000 clinical variants have been identified in these genes. However, the majority of these variants lack functional studies and their clinical significance is uncertain although accumulating evidence indicates that proteostasis deficiency is the major disease-causing mechanism for GABA receptor variants.

Adapting the endoplasmic reticulum proteostasis rescues epilepsy-associated NMDA receptor variants.

The GRIN genes encoding N-methyl-D-aspartate receptor (NMDAR) subunits are remarkably intolerant to variation. Many pathogenic NMDAR variants result in their protein misfolding, inefficient assembly, reduced surface expression, and impaired function on neuronal membrane, causing neurological disorders including epilepsy and intellectual disability. Here, we investigated the proteostasis maintenance of NMDARs containing epilepsy-associated variations in the GluN2A subunit, including M705V and A727T.

Pharmacological chaperones restore proteostasis of epilepsy-associated GABA receptor variants.

Recent advances in genetic diagnosis identified variants in genes encoding GABA receptors as causative for genetic epilepsy. Here, we selected eight disease-associated variants in the subunit of GABA receptors causing mild to severe clinical phenotypes and showed that they are loss of function, mainly by reducing the folding and surface trafficking of the protein. Furthermore, we sought client protein-specific pharmacological chaperones to restore the function of pathogenic receptors.

Anti-seizure mechanisms of midazolam and valproate at the β2(L51M) variant of the GABA receptor.

Genetic sequencing is identifying an expanding number of variants of GABA receptors associated with human epilepsies. We identified a new de novo variant of the β2 subunit (β2L51M) of the inhibitory GABA receptor associated with seizures. Our analysis determined the pathogenicity of the variant and the effects of anti-seizure medications.

Quantitative interactome proteomics identifies a proteostasis network for GABA receptors.

Gamma-aminobutyric acid type A (GABA) receptors are the primary inhibitory neurotransmitter-gated ion channels in the mammalian central nervous system. Maintenance of GABA receptor protein homeostasis (proteostasis) in cells utilizing its interacting proteins is essential for the function of GABA receptors. However, how the proteostasis network orchestrates GABA receptor biogenesis in the endoplasmic reticulum is not well understood.

The endoplasmic reticulum membrane complex promotes proteostasis of GABA receptors.

The endoplasmic reticulum membrane complex (EMC) plays a critical role in the biogenesis of tail-anchored proteins and a subset of multi-pass membrane proteins in the endoplasmic reticulum (ER). However, because of nearly exclusive expression of neurotransmitter-gated ion channels in the central nervous system (CNS), the role of the EMC in their biogenesis is not well understood. In this study, we demonstrated that the EMC positively regulates the surface trafficking and thus function of endogenous γ-aminobutyric acid type A (GABA) receptors, the primary inhibitory ion channels in the mammalian brain.

Protein quality control of -methyl-D-aspartate receptors.

N-methyl-D-aspartate receptors (NMDARs) are glutamate-gated cation channels that mediate excitatory neurotransmission and are critical for synaptic development and plasticity in the mammalian central nervous system (CNS). Functional NMDARs typically form the heterotetrameric assembly of GluN1 and GluN2 subunits. Variants within genes are implicated in various neurodevelopmental and neuropsychiatric disorders.

Stress-induced perturbations in intracellular amino acids reprogram mRNA translation in osmoadaptation independently of the ISR.

The integrated stress response (ISR) plays a pivotal role in adaptation of translation machinery to cellular stress. Here, we demonstrate an ISR-independent osmoadaptation mechanism involving reprogramming of translation via coordinated but independent actions of mTOR and plasma membrane amino acid transporter SNAT2. This biphasic response entails reduced global protein synthesis and mTOR signaling followed by translation of SNAT2.

Pharmacological activation of ATF6 remodels the proteostasis network to rescue pathogenic GABA receptors.

Background: Genetic variants in the subunits of the gamma-aminobutyric acid type A (GABA) receptors are implicated in the onset of multiple pathologic conditions including genetic epilepsy. Previous work showed that pathogenic GABA subunits promote misfolding and inefficient assembly of the GABA receptors, limiting receptor expression and activity at the plasma membrane. However, GABA receptors containing variant subunits can retain activity, indicating that enhancing the folding, assembly, and trafficking of these variant receptors offers a potential opportunity to mitigate pathology associated with genetic epilepsy.

Proteostasis Regulators Restore Function of Epilepsy-Associated GABA Receptors.

Proteostasis deficiency in mutated ion channels leads to a variety of ion channel diseases that are caused by excessive endoplasmic reticulum-associated degradation (ERAD) and inefficient membrane trafficking. We investigated proteostasis maintenance of γ-aminobutyric acid type A (GABA) receptors, the primary mediators of neuronal inhibition in the mammalian central nervous system. We screened a structurally diverse, Food and Drug Administration-approved drug library and identified dinoprost (DNP) and dihydroergocristine (DHEC) as highly efficacious enhancers of surface expression of four epilepsy-causing trafficking-deficient mutant receptors.

Interactome Changes Quantified to Identify the ER Proteostasis Network to Fight Amyloid Diseases.

In this issue of Cell Chemical Biology, Plate et al. (2019) used quantitative interactome proteomics to define the molecular mechanism by which ATF6 activation reduces amyloidogenic protein secretion. These results shed light on preventing the amyloid formation at the very early step to treat devastating amyloid diseases.

LMAN1 (ERGIC-53) promotes trafficking of neuroreceptors.

The endoplasmic reticulum-Golgi intermediate compartment protein-53 (ERGIC-53, aka LMAN1), which cycles between the endoplasmic reticulum (ER) and Golgi, is a known cargo receptor for a number of soluble proteins. However, whether LMAN1 plays a role as a trafficking factor in the central nervous system is largely unknown. Here, we determined the role of LMAN1 on endogenous protein levels of the Cys-loop superfamily of neuroreceptors, including gamma-aminobutyric acid type A receptors (GABARs), 5-hydroxytryptamine (serotonin) type 3 (5-HT) receptors, and nicotinic acetylcholine receptors (nAChRs).

Remodeling the endoplasmic reticulum proteostasis network restores proteostasis of pathogenic GABAA receptors.

Biogenesis of membrane proteins is controlled by the protein homeostasis (proteostasis) network. We have been focusing on protein quality control of γ-aminobutyric acid type A (GABAA) receptors, the major inhibitory neurotransmitter-gated ion channels in mammalian central nervous system. Proteostasis deficiency in GABAA receptors causes loss of their surface expression and thus function on the plasma membrane, leading to epilepsy and other neurological diseases.

GADD34 Function in Protein Trafficking Promotes Adaptation to Hyperosmotic Stress in Human Corneal Cells.

GADD34, a stress-induced regulatory subunit of the phosphatase PP1, is known to function in hyperosmotic stress through its well-known role in the integrated stress response (ISR) pathway. Adaptation to hyperosmotic stress is important for the health of corneal epithelial cells exposed to changes in extracellular osmolarity, with maladaptation leading to dry eye syndrome. This adaptation includes induction of SNAT2, an endoplasmic reticulum (ER)-Golgi-processed protein, which helps to reverse the stress-induced loss of cell volume and promote homeostasis through amino acid uptake.

Grp94 Protein Delivers γ-Aminobutyric Acid Type A (GABAA) Receptors to Hrd1 Protein-mediated Endoplasmic Reticulum-associated Degradation.

Proteostasis maintenance of γ-aminobutyric acid type A (GABAA) receptors dictates their function in controlling neuronal inhibition in mammalian central nervous systems. However, as a multisubunit, multispan, integral membrane protein, even wild type subunits of GABAA receptors fold and assemble inefficiently in the endoplasmic reticulum (ER). Unassembled and misfolded subunits undergo ER-associated degradation (ERAD), but this degradation process remains poorly understood for GABAA receptors.

Proteostasis Maintenance of Cys-Loop Receptors.

The Cys-loop receptors play prominent roles in the nervous system. They include γ-aminobutyric acid type A receptors, nicotinic acetylcholine receptors, 5-hydroxytryptamine type-3 receptors, and glycine receptors. Proteostasis represents an optimal state of the cellular proteome in normal physiology.

L-type Calcium Channel Blockers Enhance Trafficking and Function of Epilepsy-associated α1(D219N) Subunits of GABA(A) Receptors.

Gamma-aminobutyric acid type A (GABAA) receptors are the primary inhibitory ion channels in the mammalian central nervous system and play an essential role in regulating inhibition-excitation balance in neural circuits. The α1 subunit harboring the D219N mutation of GABAA receptors was reported to be retained in the endoplasmic reticulum (ER) and traffic inefficiently to the plasma membrane, leading to a loss of function of α1(D219N) subunits and thus idiopathic generalized epilepsy (IGE). We present the use of small molecule proteostasis regulators to enhance the forward trafficking of α1(D219N) subunits to restore their function.

Combining valosin-containing protein (VCP) inhibition and suberanilohydroxamic acid (SAHA) treatment additively enhances the folding, trafficking, and function of epilepsy-associated γ-aminobutyric acid, type A (GABAA) receptors.

GABAA receptors are the primary inhibitory ion channels in the mammalian central nervous system. The A322D mutation in the α1 subunit results in its excessive endoplasmic reticulum-associated degradation at the expense of plasma membrane trafficking, leading to autosomal dominant juvenile myoclonic epilepsy. Presumably, valosin-containing protein (VCP)/p97 extracts misfolded subunits from the endoplasmic reticulum membrane to the cytosolic proteasome for degradation.

The association of the vanin-1 N131S variant with blood pressure is mediated by endoplasmic reticulum-associated degradation and loss of function.

High blood pressure (BP) is the most common cardiovascular risk factor worldwide and a major contributor to heart disease and stroke. We previously discovered a BP-associated missense SNP (single nucleotide polymorphism)-rs2272996-in the gene encoding vanin-1, a glycosylphosphatidylinositol (GPI)-anchored membrane pantetheinase. In the present study, we first replicated the association of rs2272996 and BP traits with a total sample size of nearly 30,000 individuals from the Continental Origins and Genetic Epidemiology Network (COGENT) of African Americans (P=0.

Using pharmacological chaperones to restore proteostasis.

Normal organismal physiology depends on the maintenance of proteostasis in each cellular compartment to achieve a delicate balance between protein synthesis, folding, trafficking, and degradation while minimizing misfolding and aggregation. Defective proteostasis leads to numerous protein misfolding diseases. Pharmacological chaperones are cell-permeant small molecules that promote the proper folding and trafficking of a protein via direct binding to that protein.

SAHA enhances Proteostasis of epilepsy-associated α1(A322D)β2γ2 GABA(A) receptors.

GABA(A) receptors are the primary inhibitory ion channels in the mammalian central nervous system. The A322D mutation in the α1 subunit of GABA(A) receptors is known to result in its degradation and reduce its cell surface expression, leading to loss of GABAA receptor function in autosomal dominant juvenile myoclonic epilepsy. Here, we show that SAHA, a FDA-approved drug, increases the transcription of the α1(A322D) subunit, enhances its folding and trafficking posttranslationally, increases its cell surface level, and restores the GABA-induced maximal current in HEK293 cells expressing α1(A322D)β2γ2 receptors to 10% of that for wild-type receptors.