Cryo-EM structures and functional properties of CALHM channels of the human placenta
- University of Zürich, Switzerland
- University of Bern, Switzerland
Abstract
The transport of substances across the placenta is essential for the development of the fetus. Here, we were interested in the role of channels of the calcium homeostasis modulator (CALHM) family in the human placenta. By transcript analysis, we found the paralogs CALHM2, 4, and 6 to be highly expressed in this organ and upregulated during trophoblast differentiation. Based on electrophysiology, we found that activation of these paralogs differs from the voltage- and calcium-gated channel CALHM1. Cryo-EM structures of CALHM4 display decameric and undecameric assemblies with large cylindrical pore, while in CALHM6 a conformational change has converted the pore shape into a conus that narrows at the intracellular side, thus describing distinct functional states of the channel. The pore geometry alters the distribution of lipids, which occupy the cylindrical pore of CALHM4 in a bilayer-like arrangement whereas they have redistributed in the conical pore of CALHM6 with potential functional consequences.
Data availability
cryo-EM data were deposited with the PDB and cryo-EM densiities were deposited with the Electron Microscopy databank
-
Cryo-EM structure of decameric human CALHM6 in the presence of Ca2+Protein Data Bank, 6YTV.
-
Cryo-EM structure of undecameric human CALHM6 in the presence of Ca2+Protein Data Bank, 6YTX.
Article and author information
Author details
Katarzyna Drożdżyk
Funding
Schweizerischer Nationalfonds zur Förderung der Wissenschaftlichen Forschung (31003A_163421)
- Raimund Dutzler
Stiftung Lindenhof Bern
- Christiane Albrecht
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Ethics
Human subjects: Human placental tissues were collected from the Division of Obstetrics and Gynecology, Lindenhofgruppe Bern, Switzerland, under approval by the ethical commission of the Canton of Bern (approval No Basec 2016-00250). Written informed consent was obtained from all participants.
Copyright
© 2020, Drożdżyk 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
-
- 3,420
- views
-
- 517
- downloads
-
- 39
- citations
Views, downloads and citations are aggregated across all versions of this paper published by eLife.
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)
Open citations (links to open the citations from this article in various online reference manager services)
Cite this article (links to download the citations from this article in formats compatible with various reference manager tools)
Cryo-EM structures and functional properties of CALHM channels of the human placenta
eLife 9:e55853.
https://doi.org/10.7554/eLife.55853
Further reading
-
- Immunology and Inflammation
- Structural Biology and Molecular Biophysics
Antibodies are a major component of adaptive immunity against invading pathogens. Here, we explore possibilities for an analytical approach to characterize the antigen-specific antibody repertoire directly from the secreted proteins in convalescent serum. This approach aims to perform simultaneous antibody sequencing and epitope mapping using a combination of single particle cryo-electron microscopy (cryoEM) and bottom-up proteomics techniques based on mass spectrometry (LC-MS/MS). We evaluate the performance of the deep-learning tool ModelAngelo in determining de novo antibody sequences directly from reconstructed 3D volumes of antibody-antigen complexes. We demonstrate that while map quality is a critical bottleneck, it is possible to sequence antibody variable domains from cryoEM reconstructions with accuracies of up to 80–90%. While the rate of errors exceeds the typical levels of somatic hypermutation, we show that the ModelAngelo-derived sequences can be used to assign the used V-genes. This provides a functional guide to assemble de novo peptides from LC-MS/MS data more accurately and improves the tolerance to a background of polyclonal antibody sequences. Following this proof-of-principle, we discuss the feasibility and future directions of this approach to characterize antigen-specific antibody repertoires.
-
- Biochemistry and Chemical Biology
- Structural Biology and Molecular Biophysics
The two identical motor domains (heads) of dimeric kinesin-1 move in a hand-over-hand process along a microtubule, coordinating their ATPase cycles such that each ATP hydrolysis is tightly coupled to a step and enabling the motor to take many steps without dissociating. The neck linker, a structural element that connects the two heads, has been shown to be essential for head–head coordination; however, which kinetic step(s) in the chemomechanical cycle is ‘gated’ by the neck linker remains unresolved. Here, we employed pre-steady-state kinetics and single-molecule assays to investigate how the neck-linker conformation affects kinesin’s motility cycle. We show that the backward-pointing configuration of the neck linker in the front kinesin head confers higher affinity for microtubule, but does not change ATP binding and dissociation rates. In contrast, the forward-pointing configuration of the neck linker in the rear kinesin head decreases the ATP dissociation rate but has little effect on microtubule dissociation. In combination, these conformation-specific effects of the neck linker favor ATP hydrolysis and dissociation of the rear head prior to microtubule detachment of the front head, thereby providing a kinetic explanation for the coordinated walking mechanism of dimeric kinesin.
