Ubiquitination drives COPI priming and Golgi SNARE localization
- Vanderbilt University, United States
- University of Massachusetts Amherst, United States
Abstract
Deciphering mechanisms controlling SNARE localization within the Golgi complex is crucial to understanding protein trafficking patterns within the secretory pathway. SNAREs are also thought to prime COPI assembly to ensure incorporation of these essential cargoes into vesicles, but the regulation of these events is poorly understood. Here we report roles for ubiquitin recognition by COPI in SNARE trafficking and in stabilizing interactions between Arf, COPI, and Golgi SNAREs in Saccharomyces cerevisiae. The ability of COPI to bind ubiquitin, but not the dilysine motif, through its N-terminal WD repeat domain of β'COP or through an unrelated ubiquitin-binding domain (UBD) is essential for the proper localization of Golgi SNAREs Bet1 and Gos1. We find that COPI, the ArfGAP Glo3 and multiple Golgi SNAREs are ubiquitinated. Notably, the binding of Arf and COPI to Gos1 is markedly enhanced by ubiquitination of these components. Glo3 is proposed to prime COPI-SNARE interactions; however, Glo3 is not enriched in the ubiquitin-stabilized SNARE-Arf-COPI complex but is instead enriched with COPI complexes that lack SNAREs. These results support a new model for how posttranslational modifications drive COPI priming events crucial for Golgi SNARE localization.
Data availability
All data generated or analyzed during this study are included in the manuscript and supporting file; Source Data files have been provided for all figures.
Article and author information
Author details
Funding
National Institutes of Health (R35GM144123-01)
- Todd R Graham
National Institutes of Health (1R35GM119525)
- Lauren P Jackson
National Institutes of Health (R35GM144112)
- Jason A MacGurn
Pew Charitable Trusts
- Lauren P Jackson
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2022, Date 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
-
- 1,897
- views
-
- 467
- downloads
-
- 8
- 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)
Ubiquitination drives COPI priming and Golgi SNARE localization
eLife 11:e80911.
https://doi.org/10.7554/eLife.80911
Further reading
-
- Cell Biology
- Chromosomes and Gene Expression
We investigated the role of the nucleolar protein Treacle in organizing and regulating the nucleolus in human cells. Our results support Treacle’s ability to form liquid-like phase condensates through electrostatic interactions among molecules. The formation of these biomolecular condensates is crucial for segregating nucleolar fibrillar centers from the dense fibrillar component and ensuring high levels of ribosomal RNA (rRNA) gene transcription and accurate rRNA processing. Both the central and C-terminal domains of Treacle are required to form liquid-like condensates. The initiation of phase separation is attributed to the C-terminal domain. The central domain is characterized by repeated stretches of alternatively charged amino acid residues and is vital for condensate stability. Overexpression of mutant forms of Treacle that cannot form liquid-like phase condensates compromises the assembly of fibrillar centers, suppressing rRNA gene transcription and disrupting rRNA processing. These mutant forms also fail to recruit DNA topoisomerase II binding protein 1 (TOPBP1), suppressing the DNA damage response in the nucleolus.
-
- Cell Biology
The primary cilium is a microtubule-based organelle that cycles through assembly and disassembly. In many cell types, formation of the cilium is initiated by recruitment of preciliary vesicles to the distal appendage of the mother centriole. However, the distal appendage mechanism that directly captures preciliary vesicles is yet to be identified. In an accompanying paper, we show that the distal appendage protein, CEP89, is important for the preciliary vesicle recruitment, but not for other steps of cilium formation (Kanie et al., 2025). The lack of a membrane-binding motif in CEP89 suggests that it may indirectly recruit preciliary vesicles via another binding partner. Here, we identify Neuronal Calcium Sensor-1 (NCS1) as a stoichiometric interactor of CEP89. NCS1 localizes to the position between CEP89 and the centriole-associated vesicle marker, RAB34, at the distal appendage. This localization was completely abolished in CEP89 knockouts, suggesting that CEP89 recruits NCS1 to the distal appendage. Similar to CEP89 knockouts, preciliary vesicle recruitment as well as subsequent cilium formation was perturbed in NCS1 knockout cells. The ability of NCS1 to recruit the preciliary vesicle is dependent on its myristoylation motif and NCS1 knockout cells expressing a myristoylation defective mutant failed to rescue the vesicle recruitment defect despite localizing properly to the centriole. In sum, our analysis reveals the first known mechanism for how the distal appendage recruits the preciliary vesicles.
