High-resolution secretory timeline from vesicle formation at the Golgi to fusion at the plasma membrane in S. cerevisiae
- Cornell University, United States
Abstract
Most of the components in the yeast secretory pathway have been studied, yet a high-resolution temporal timeline of their participation is lacking. Here we define the order of acquisition, lifetime, and release of critical components involved in late secretion from the Golgi to the plasma membrane. Of particular interest is the timing of the many reported effectors of the secretory vesicle Rab protein Sec4, including the myosin-V Myo2, the exocyst complex, the lgl homolog Sro7, and the small yeast-specific protein Mso1. At the trans-Golgi network (TGN) Sec4's GEF, Sec2, is recruited to Ypt31-positive compartments, quickly followed by Sec4 and Myo2 and vesicle formation. While transported to the bud tip, the entire exocyst complex, including Sec3, is assembled on to the vesicle. Before fusion, vesicles tether for 5s, during which the vesicle retains the exocyst complex and stimulates lateral recruitment of Rho3 on the plasma membrane. Sec2 and Myo2 are rapidly lost, followed by recruitment of cytosolic Sro7, and finally the SM protein Sec1, which appears for just 2 seconds prior to fusion. Perturbation experiments reveal an ordered and robust series of events during tethering that provide insights into the function of Sec4 and effector exchange.
Data availability
Details of each yeast strain used is in Table 1.All plasmids used are listed in Table 2.All DNA oligos used are listed in Table 3.The unique sequence used for the Myo2 marker is in Table 4.All sample sizes are provided in Table 5.All raw component data as well as mean and median is provided in Table 6.
Article and author information
Author details
Funding
National Institute of General Medical Sciences (5RO1GM039066)
- Anthony Bretscher
National Institute of General Medical Sciences (5R35GM131751)
- Anthony Bretscher
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Copyright
© 2022, Gingras 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.
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High-resolution secretory timeline from vesicle formation at the Golgi to fusion at the plasma membrane in S. cerevisiae
eLife 11:e78750.
https://doi.org/10.7554/eLife.78750
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