Structure of the gene therapy vector, adeno-associated virus with its cell receptor, AAVR
- Oregon Health and Science University, United States
- Florida State University, United States
- University of Missouri, United States
Abstract
Adeno-associated virus (AAV) vectors are preeminent in emerging clinical gene therapies. Generalizing beyond the most tractable genetic diseases will require modulation of cell specificity and immune neutralization. Interactions of AAV with its cellular receptor, AAVR, are key to understanding cell-entry and trafficking with the rigor needed to engineer tissue-specific vectors. Cryo-electron tomography shows ordered binding of part of the flexible receptor to the viral surface, with distal domains in multiple conformations. Regions of the virus and receptor in close physical proximity can be identified by cross-linking / mass spectrometry. Cryo-electron microscopy with a two-domain receptor fragment reveals the interactions at 2.4 Å resolution. AAVR binds between AAV's spikes on a plateau that is conserved, except in one clade whose structure is AAVR-incompatible. AAVR's footprint overlaps the epitopes of several neutralizing antibodies, prompting a re-evaluation of neutralization mechanisms. The structure provides a roadmap for experimental probing and manipulation of viral-receptor interactions.
Data availability
Electron microscopy maps and atomic coordinates will be available from the electron microscopy and protein data banks (https://www.ebi.ac.uk/pdbe/emdb/ & https://www.rcsb.org/). For the high resolution PKD1-2/AAV2 complex, the accession numbers are EMD-0553, PDB ID 6NZ0, respectively. Reconstructions for the 4 tomographic classes have accession numbers of EMD-0621, EMD-0622, EMD-0623 and EMD-0624.
-
Cryo-EM structure of AAV-2 in complex with AAVR PKD domains 1 and 2Protein Data Bank, 6NZ0.
-
Cryo-EM structure of AAV-2 in complex with AAVR PKD domains 1 and 2Unified Data Resource for 3DEM, EMD-0553.
-
Structure of the AAV2 with its Cell Receptor AAVRUnified Data Resource for 3DEM, EMD-0621.
-
Structure of the AAV2 with its Cell Receptor AAVRUnified Data Resource for 3DEM, EMD-0622.
-
Structure of the AAV2 with its Cell Receptor AAVRUnified Data Resource for 3DEM, EMD-0623.
-
Structure of the AAV2 with its Cell Receptor AAVRUnified Data Resource for 3DEM, EMD-0624.
Article and author information
Author details
Michael Stewart Chapman
Funding
National Institutes of Health (R35GM122564)
- Andrew Trzynka
National Institutes of Health (R01GM066875)
- Michael Stewart Chapman
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2019, Meyer 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
-
- 12,209
- views
-
- 1,795
- downloads
-
- 64
- 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)
Structure of the gene therapy vector, adeno-associated virus with its cell receptor, AAVR
eLife 8:e44707.
https://doi.org/10.7554/eLife.44707
Further reading
-
- Microbiology and Infectious Disease
The drivers of tissue necrosis in Mycobacterium ulcerans infection (Buruli ulcer disease) have historically been ascribed solely to the directly cytotoxic action of the diffusible exotoxin, mycolactone. However, its role in the clinically evident vascular component of disease aetiology remains poorly explained. We have now dissected mycolactone’s effects on human primary vascular endothelial cells in vitro. We show that mycolactone-induced changes in endothelial morphology, adhesion, migration, and permeability are dependent on its action at the Sec61 translocon. Unbiased quantitative proteomics identified a profound effect on proteoglycans, driven by rapid loss of type II transmembrane proteins of the Golgi, including enzymes required for glycosaminoglycan (GAG) synthesis, combined with a reduction in the core proteins themselves. Loss of the glycocalyx is likely to be of particular mechanistic importance, since knockdown of galactosyltransferase II (beta-1,3-galactotransferase 6; B3GALT6), the GAG linker-building enzyme, phenocopied the permeability and phenotypic changes induced by mycolactone. Additionally, mycolactone depleted many secreted basement membrane components and microvascular basement membranes were disrupted in vivo during M. ulcerans infection in the mouse model. Remarkably, exogenous addition of laminin-511 reduced endothelial cell rounding, restored cell attachment and reversed the defective migration caused by mycolactone. Hence supplementing mycolactone-depleted extracellular matrix may be a future therapeutic avenue, to improve wound healing rates.
-
- Biochemistry and Chemical Biology
- Microbiology and Infectious Disease
Mycofactocin is a redox cofactor essential for the alcohol metabolism of mycobacteria. While the biosynthesis of mycofactocin is well established, the gene mftG, which encodes an oxidoreductase of the glucose-methanol-choline superfamily, remained functionally uncharacterized. Here, we show that MftG enzymes are almost exclusively found in genomes containing mycofactocin biosynthetic genes and are present in 75% of organisms harboring these genes. Gene deletion experiments in Mycolicibacterium smegmatis demonstrated a growth defect of the ∆mftG mutant on ethanol as a carbon source, accompanied by an arrest of cell division reminiscent of mild starvation. Investigation of carbon and cofactor metabolism implied a defect in mycofactocin reoxidation. Cell-free enzyme assays and respirometry using isolated cell membranes indicated that MftG acts as a mycofactocin dehydrogenase shuttling electrons toward the respiratory chain. Transcriptomics studies also indicated remodeling of redox metabolism to compensate for a shortage of redox equivalents. In conclusion, this work closes an important knowledge gap concerning the mycofactocin system and adds a new pathway to the intricate web of redox reactions governing the metabolism of mycobacteria.
