Decoding polyubiquitin regulation of KV7. 1 functional expression with engineered linkage-selective deubiquitinases
Sri Karthika Shanmugam1, Scott A Kanner2, Xinle Zou3
1Department of Physiology and Cellular Biophysics.
Biorxiv : the Preprint Server for Biology
|September 30, 2024
Summary
Engineered deubiquitinases (enDUBs) reveal how specific polyubiquitin chains regulate protein function. This study introduces a novel tool for dissecting the ubiquitin code and its role in ion channel regulation.
Area of Science:
- Cellular Biology
- Biochemistry
- Molecular Biology
Background:
- Protein posttranslational modification by polyubiquitin chains forms the 'ubiquitin code,' regulating protein function.
- Understanding the specific roles of diverse polyubiquitin linkages remains challenging due to limited methods for selective manipulation in live cells.
Purpose of the Study:
- To develop and utilize engineered deubiquitinases (enDUBs) for selective manipulation of polyubiquitin linkages on proteins.
- To investigate the functional consequences of distinct polyubiquitin linkages on the ion channel YFP-KCNQ1.
Main Methods:
- Engineered deubiquitinases (enDUBs) were created by fusing DUB catalytic domains to GFP-nanobodies.
- enDUBs were used to modulate polyubiquitin linkages on YFP-KCNQ1 in HEK293 cells and cardiomyocytes.
- Mass spectrometry was employed to identify polyubiquitin linkages on YFP-KCNQ1.
Main Results:
- K48 and K63 linkages were dominant on YFP-KCNQ1.
- Different enDUBs induced unique effects on YFP-KCNQ1 expression, localization, and function.
- Specific linkages (K11, K29/K33, K63) were found to mediate intracellular retention through distinct mechanisms.
- enDUBs showed differential efficacy in rescuing trafficking-deficient long QT syndrome type 1 mutations.
Conclusions:
- Distinct polyubiquitin chains differentially regulate various aspects of KCNQ1 ion channel function.
- The study highlights the plasticity of the ubiquitin code.
- Linkage-selective enDUBs represent a powerful new tool for deciphering the ubiquitin code.
Related Concept Videos
02:04Covalently Linked Protein Regulators
6.8K
Proteins can undergo many types of post-translational modifications, often in response to changes in their environment. These modifications play an important role in the function and stability of these proteins. Covalently linked molecules include functional groups, such as methyl, acetyl, and phosphate groups, and also small proteins, such as ubiquitin. There are around 200 different types of covalent regulators that have been identified.
These groups modify specific amino acids in a protein....
These groups modify specific amino acids in a protein....
6.8K
02:58Regulated Protein Degradation
7.2K
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...
Protein degradation plays two important roles in the cells. It helps to protect cells from misfolded or damaged proteins before they lead to a...
7.2K
01:13The Proteasome
817
Eukaryotic cells can degrade proteins through several pathways. One of the most important among these is the ubiquitin-proteasome pathway. It helps the cell eliminate the misfolded, damaged, or unwarranted cytoplasmic proteins in a highly specific manner.
In this pathway, the target proteins are first tagged with small proteins called ubiquitin. This involves participation of a series of enzymes including— E1 (ubiquitin-activating enzyme), E2 (ubiquitin-conjugating enzyme), and E3...
In this pathway, the target proteins are first tagged with small proteins called ubiquitin. This involves participation of a series of enzymes including— E1 (ubiquitin-activating enzyme), E2 (ubiquitin-conjugating enzyme), and E3...
817
01:57Protein Complexes with Interchangeable Parts
2.5K
Groups of proteins may form a complex where each protein in this complex has a different role in the overall execution of the complex’s function. Often some of the proteins in the complex can be replaced by a closely related variant to give a complex that contains many of the same components yet is functionally distinct.
The SCF ubiquitin ligase is a protein complex of five individual proteins. This complex attaches ubiquitin to other target proteins to mark them for degradation. In order...
The SCF ubiquitin ligase is a protein complex of five individual proteins. This complex attaches ubiquitin to other target proteins to mark them for degradation. In order...
2.5K
01:15Receptor Downregulation in MVBs
2.0K
Multivesicular bodies (MVBs) are mature endosomes that sort ubiquitinated proteins and then fuse with lysosomes to degrade the sorted proteins. Epidermal growth factor (EGF) and its receptor (EGFR) form a complex that can be internalized through endocytosis, sorted into an MVB, and later degraded.
The EGFR can initiate signaling pathways that lead to cell proliferation, migration, and differentiation. Overexpression of EGFR stimulates cells to proliferate. Excessive EGFR...
The EGFR can initiate signaling pathways that lead to cell proliferation, migration, and differentiation. Overexpression of EGFR stimulates cells to proliferate. Excessive EGFR...
2.0K
01:23Regulation of Expression at Multiple Steps
875
The gene expression in cells is regulated at different stages: (i) transcription, (ii) RNA processing, (iii) RNA localization, and (iv) translation. Transcriptional regulation is mediated by regulatory proteins such as transcription factors, activators, or repressors—these control gene expression by initiating or inhibiting the transcription of genes. Once a precursor or pre-mRNA is produced, it undergoes post-transcriptional modification, including 5' capping, splicing, and the...
875