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Related Concept Videos

Regulated Protein Degradation02:58

Regulated Protein Degradation

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It is vital to regulate the activity of enzymatic as well as non-enzymatic proteins inside the cell. This can be achieved either through creating a balance between their rate of synthesis and degradation or regulating the intrinsic activity of the protein. Both these regulation mechanisms play an essential role in the normal functioning of cells.
Protein degradation plays two important roles in the cells. It helps to protect cells from misfolded or damaged proteins before they lead to a...
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Regulated Protein Degradation02:58

Regulated Protein Degradation

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High-Throughput Cellular Profiling of Targeted Protein Degradation Compounds Using HiBiT CRISPR Cell Lines
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A cell based screening approach for identifying protein degradation regulators.

Scott Simanski1, Marie E Maloof2, Trey K Sato3

  • 1a Department of Cancer Biology , Scripps Florida, The Scripps Research Institute , Jupiter , FL , USA.

Cell Cycle (Georgetown, Tex.)
|March 16, 2017
PubMed
Summary
This summary is machine-generated.

Regulated protein degradation drives cell cycle transitions. Researchers identified Cdh1 and Fbxl15 as key regulators of cyclin B1 turnover using a novel luciferase reporter assay for protein degradation pathways.

Keywords:
cell-based screeningdegradationluciferasesubstrateubiquitin

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Area of Science:

  • Cell Biology
  • Molecular Biology
  • Biochemistry

Background:

  • Cellular transitions depend on regulated protein degradation pathways, particularly ubiquitin-mediated degradation, which ensures unidirectional cell cycle progression.
  • The turnover of cell cycle regulators like cyclin B1 is critical for processes such as mitotic exit.

Purpose of the Study:

  • To develop and apply a screening approach using luciferase fusion proteins to identify specific regulators of protein turnover.
  • To investigate the pathways controlling cyclin B1 turnover by adapting a reporter system.

Main Methods:

  • Created an N-cyclin B1-luciferase fusion protein as a reporter for protein turnover.
  • Overexpressed approximately 14,000 cDNAs with the N-cyclin B1-luciferase fusion protein.
  • Analyzed the steady-state levels of the reporter protein relative to other luciferase fusion proteins to identify turnover regulators.

Main Results:

  • Identified the known Anaphase-Promoting Complex/Cyclosome (APC/C) regulator Cdh1 as a modulator of N-cyclin B1-luciferase steady-state levels.
  • Identified the F-box protein Fbxl15 as a novel specific modulator of N-cyclin B1-luciferase turnover.
  • Demonstrated that parallel analysis of luciferase fusion protein steady-state levels facilitates the identification of specific protein turnover regulators.

Conclusions:

  • The developed screening approach is effective for identifying specific regulators of protein turnover.
  • Cdh1 and Fbxl15 play significant roles in regulating cyclin B1 turnover.
  • This method provides a powerful tool for dissecting protein degradation pathways crucial for cell cycle control.