1. Structural Biology and Molecular Biophysics
Download icon

A critical residue in the α1M2-M3 linker regulating mammalian GABAA receptor pore gating by diazepam

  1. Joseph W Nors
  2. Shipra Gupta
  3. Marcel P Goldschen-Ohm  Is a corresponding author
  1. University of Texas at Austin, United States
Research Article
  • Cited 0
  • Views 206
  • Annotations
Cite this article as: eLife 2021;10:e64400 doi: 10.7554/eLife.64400

Abstract

Benzodiazepines (BZDs) are a class of widely prescribed psychotropic drugs that modulate activity of GABAA receptors (GABAARs), neurotransmitter-gated ion channels critical for synaptic transmission. However, the physical basis of this modulation is poorly understood. We explore the role of an important gating domain, the a1M2-M3 linker, in linkage between the BZD site and pore gate. To probe energetics of this coupling without complication from bound agonist we use a gain of function mutant (a1L9'Tb2g2L) directly activated by BZDs. We identify a specific residue whose mutation (a1V279A) more than doubles the energetic contribution of the BZD positive modulator diazepam (DZ) to pore opening and also enhances DZ-potentiation of GABA-evoked currents in a wild-type background. In contrast, other linker mutations have little effect on DZ efficiency, but generally impair unliganded pore opening. Our observations reveal an important residue regulating BZD-pore linkage, thereby shedding new light on the molecular mechanism of these drugs.

Article and author information

Author details

  1. Joseph W Nors

    Neuroscience, University of Texas at Austin, Austin, United States
    Competing interests
    The authors declare that no competing interests exist.

  2. Shipra Gupta

    Neuroscience, University of Texas at Austin, Austin, United States
    Competing interests
    The authors declare that no competing interests exist.

  3. Marcel P Goldschen-Ohm

    Neuroscience, University of Texas at Austin, Austin, United States
    For correspondence
    marcel.goldschen-ohm@austin.utexas.edu
    Competing interests
    The authors declare that no competing interests exist.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-1466-9808

Funding

University of Texas at Austin (Department of Neuroscience Startup)

  • Marcel P Goldschen-Ohm

University of Texas at Austin (STARS)

  • Marcel P Goldschen-Ohm

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Leon D Islas, Universidad Nacional Autónoma de México, Mexico

Publication history

  1. Received: October 27, 2020
  2. Accepted: February 15, 2021
  3. Accepted Manuscript published: February 16, 2021 (version 1)
  4. Version of Record published: February 22, 2021 (version 2)

Copyright

© 2021, Nors et al.

This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.

Metrics

  • 206
    Page views
  • 32
    Downloads
  • 0
    Citations

Article citation count generated by polling the highest count across the following sources: Crossref, PubMed Central, Scopus.

Download links

A two-part list of links to download the article, or parts of the article, in various formats.

Downloads (link to download the article as PDF)

Download citations (links to download the citations from this article in formats compatible with various reference manager tools)

Open citations (links to open the citations from this article in various online reference manager services)

Categories and tags

Research organism

Further reading

    1. Cell Biology
    2. Structural Biology and Molecular Biophysics

    Structural basis of Stu2 recruitment to yeast kinetochores

    Jacob A Zahm et al.
    Research Article Updated

    Chromosome segregation during cell division requires engagement of kinetochores of sister chromatids with microtubules emanating from opposite poles. As the corresponding microtubules shorten, these ‘bioriented’ sister kinetochores experience tension-dependent stabilization of microtubule attachments. The yeast XMAP215 family member and microtubule polymerase, Stu2, associates with kinetochores and contributes to tension-dependent stabilization in vitro. We show here that a C-terminal segment of Stu2 binds the four-way junction of the Ndc80 complex (Ndc80c) and that residues conserved both in yeast Stu2 orthologs and in their metazoan counterparts make specific contacts with Ndc80 and Spc24. Mutations that perturb this interaction prevent association of Stu2 with kinetochores, impair cell viability, produce biorientation defects, and delay cell cycle progression. Ectopic tethering of the mutant Stu2 species to the Ndc80c junction restores wild-type function in vivo. These findings show that the role of Stu2 in tension-sensing depends on its association with kinetochores by binding with Ndc80c.

    1. Neuroscience
    2. Structural Biology and Molecular Biophysics

    Molecular basis for functional connectivity between the voltage sensor and the selectivity filter gate in Shaker K+ channels

    Carlos A Z Bassetto Jnr et al.
    Research Article

    In Shaker K+ channels, the S4-S5 linker couples the voltage sensor (VSD) and pore domain (PD). Another coupling mechanism is revealed using two W434F-containing channels: L361R:W434F and L366H:W434F. In L361R:W434F, W434F affects the L361R VSD seen as a shallower Q-V curve that crosses the G-V. In L366H:W434F, L366H relieves the W434F effect converting a non-conductive channel in a conductive one. We report a chain of residues connecting the VSD (S4) to the selectivity filter (SF) in the PD of an adjacent subunit as the molecular basis for voltage-sensor selectivity filter gate (VS-SF) coupling. Single alanine substitutions in this region (L409A, S411A, S412A or F433A) are enough to disrupt the VS-SF coupling, shown by the absence of Q-V and G-V crossing in L361R:W434F mutant and by the lack of ionic conduction in the L366H:W434F mutant. This residue chain defines a new coupling between the VSD and the PD in voltage-gated channels.

    1. Cell Biology
    2. Structural Biology and Molecular Biophysics

    Pre-existing bilayer stresses modulate triglyceride accumulation in the ER versus lipid droplets

    Valeria Zoni et al.
    Research Article Updated

    Cells store energy in the form of neutral lipids (NLs) packaged into micrometer-sized organelles named lipid droplets (LDs). These structures emerge from the endoplasmic reticulum (ER) at sites marked by the protein seipin, but the mechanisms regulating their biogenesis remain poorly understood. Using a combination of molecular simulations, yeast genetics, and fluorescence microscopy, we show that interactions between lipids’ acyl-chains modulate the propensity of NLs to be stored in LDs, in turn preventing or promoting their accumulation in the ER membrane. Our data suggest that diacylglycerol, which is enriched at sites of LD formation, promotes the packaging of NLs into LDs, together with ER-abundant lipids, such as phosphatidylethanolamine. On the opposite end, short and saturated acyl-chains antagonize fat storage in LDs and promote accumulation of NLs in the ER. Our results provide a new conceptual understanding of LD biogenesis in the context of ER homeostasis and function.