A microscopy-based kinetic analysis of yeast vacuolar protein sorting

Casler, Jason C.; Glick, Benjamin S.

doi:10.7554/elife.56844

Cited by 32 publications

(35 citation statements)

References 87 publications

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“…This effect highlights a limitation of ESCargo: as previously documented for yeast cells, there is a kinetic competition between aggregation in the ER and signal-dependent ER export ( Casler et al. , 2019 ; Casler and Glick, 2020 ). In mammalian cells, ER export tends to win the race, and so much of the ESCargo accumulates in post-ER compartments.…”

Section: Resultsmentioning

confidence: 57%

“…First, when ESCargo molecules containing the APV tripeptide enter the ER, they experience a kinetic competition between aggregation and signal-dependent ER export ( Casler et al. , 2019 ; Casler and Glick, 2020 ). As a result, a substantial fraction of the newly synthesized ESCargo molecules are either secreted or delivered to post-ER compartments.…”

Section: Resultsmentioning

confidence: 99%

“…, 2019 ). This approach was subsequently extended by adding a C-terminal vacuolar targeting peptide to ESCargo, thereby enabling a kinetic analysis of biosynthetic traffic to the yeast vacuole ( Casler and Glick, 2020 ).…”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

ESCargo: a regulatable fluorescent secretory cargo for diverse model organisms

Casler

Zajac

Valbuena

et al. 2020

MBoC

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Membrane traffic can be studied by imaging a cargo protein as it transits the secretory pathway. The best tools for this purpose initially block export of the secretory cargo from the endoplasmic reticulum (ER), and then release the block to generate a cargo wave. However, previously developed regulatable secretory cargoes are often tricky to use or specific for a single model organism. To overcome these hurdles for budding yeast, we recently optimized an artificial fluorescent secretory protein that exits the ER with the aid of the Erv29 cargo receptor, which is homologous to mammalian Surf4. The fluorescent secretory protein forms aggregates in the ER lumen and can be rapidly disaggregated by addition of a ligand to generate a nearly synchronized cargo wave. Here we term this regulatable secretory protein ESCargo (Erv29/Surf4-dependent Secretory Cargo) and demonstrate its utility not only in yeast cells, but also in cultured mammalian cells, Drosophila cells, and the ciliate Tetrahymena thermophila. Kinetic studies indicate that rapid export from the ER requires recognition by Erv29/Surf4. By choosing an appropriate ER signal sequence and expression vector, this simple technology can likely be used with many model organisms. [Media: see text] [Media: see text] [Media: see text]

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Section: Resultsmentioning

confidence: 57%

Section: Resultsmentioning

confidence: 99%

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ESCargo: a regulatable fluorescent secretory cargo for diverse model organisms

Casler

Zajac

Valbuena

et al. 2020

MBoC

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“…Microscopy-based kinetic analysis provides information on the dynamic accumulation of a protein at a given subcellular location, which can be very sensitive to environmental and genetic perturbations. This approach has proven powerful in understanding the fine control of timely SS formation by the F-BAR protein Hof1 ( Oh, et al., 2017 ), the CDK regulation of IQGAP Iqg1 in AMR assembly ( Naylor and Morgan, 2014 ), and vacuolar protein sorting in budding yeast ( Casler and Glick, 2020 ). This approach has also provided mechanistic insights into the roles of the myosin-II isoforms Myo2 and Myp2 in cytokinesis under normal and stressed conditions in fission yeast ( Okada, et al., 2019 ).…”

Section: Introductionmentioning

confidence: 99%

The kinetic landscape and interplay of protein networks in cytokinesis

et al. 2021

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Summary Cytokinesis is executed by protein networks organized into functional modules. Individual proteins within each module have been characterized to various degrees. However, the collective behavior and interplay of the modules remain poorly understood. In this study, we conducted quantitative time-lapse imaging to analyze the accumulation kinetics of more than 20 proteins from different modules of cytokinesis in budding yeast. This analysis has led to a comprehensive picture of the kinetic landscape of cytokinesis, from actomyosin ring (AMR) assembly to cell separation. It revealed that the AMR undergoes biphasic constriction and that the switch between the constriction phases is likely triggered by AMR maturation and primary septum formation. This analysis also provided further insights into the functions of actin filaments and the transglutaminase-like protein Cyk3 in cytokinesis and, in addition, defined Kre6 as the likely enzyme that catalyzes β-1,6-glucan synthesis to drive cell wall maturation during cell growth and division.

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“…In the final step, late endosomes fuse with the highly acidic vacuoles that contain proteases for degradation of the endosomal contents. Recently, very elegant experiments using an engineered fluorescent vacuolar cargo and 4D microscopy have suggested that transfer of material from late endosomes to the vacuole most likely involves “ kiss-and-run ” fusion events [ 90 , 92 ].…”

Section: Sorting Nexin Cargo Recognition In Yeastmentioning

confidence: 99%

Sorting Nexins in Protein Homeostasis

Hanley

Cooper

2020

Cells

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Protein homeostasis is maintained by removing misfolded, damaged, or excess proteins and damaged organelles from the cell by three major pathways; the ubiquitin-proteasome system, the autophagy-lysosomal pathway, and the endo-lysosomal pathway. The requirement for ubiquitin provides a link between all three pathways. Sorting nexins are a highly conserved and diverse family of membrane-associated proteins that not only traffic proteins throughout the cells but also provide a second common thread between protein homeostasis pathways. In this review, we will discuss the connections between sorting nexins, ubiquitin, and the interconnected roles they play in maintaining protein quality control mechanisms. Underlying their importance, genetic defects in sorting nexins are linked with a variety of human diseases including neurodegenerative, cardiovascular diseases, viral infections, and cancer. This serves to emphasize the critical roles sorting nexins play in many aspects of cellular function.

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A microscopy-based kinetic analysis of yeast vacuolar protein sorting

Cited by 32 publications

References 87 publications

ESCargo: a regulatable fluorescent secretory cargo for diverse model organisms

ESCargo: a regulatable fluorescent secretory cargo for diverse model organisms

The kinetic landscape and interplay of protein networks in cytokinesis

Sorting Nexins in Protein Homeostasis

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