Analysis of Wild Type and Variant B Cystatin C Interactome in Retinal Pigment Epithelium Cells Reveals Variant B Interacting Mitochondrial Proteins
Issued Date
2023-03-01
Resource Type
eISSN
20734409
Scopus ID
2-s2.0-85149801903
Pubmed ID
36899848
Journal Title
Cells
Volume
12
Issue
5
Rights Holder(s)
SCOPUS
Bibliographic Citation
Cells Vol.12 No.5 (2023)
Suggested Citation
Carlsson E., Sharif U., Supharattanasitthi W., Paraoan L. Analysis of Wild Type and Variant B Cystatin C Interactome in Retinal Pigment Epithelium Cells Reveals Variant B Interacting Mitochondrial Proteins. Cells Vol.12 No.5 (2023). doi:10.3390/cells12050713 Retrieved from: https://repository.li.mahidol.ac.th/handle/20.500.14594/81638
Title
Analysis of Wild Type and Variant B Cystatin C Interactome in Retinal Pigment Epithelium Cells Reveals Variant B Interacting Mitochondrial Proteins
Author(s)
Author's Affiliation
Other Contributor(s)
Abstract
Cystatin C, a secreted cysteine protease inhibitor, is abundantly expressed in retinal pigment epithelium (RPE) cells. A mutation in the protein’s leader sequence, corresponding to formation of an alternate variant B protein, has been linked with an increased risk for both age-related macular degeneration (AMD) and Alzheimer’s disease (AD). Variant B cystatin C displays intracellular mistrafficking with partial mitochondrial association. We hypothesized that variant B cystatin C interacts with mitochondrial proteins and impacts mitochondrial function. We sought to determine how the interactome of the disease-related variant B cystatin C differs from that of the wild-type (WT) form. For this purpose, we expressed cystatin C Halo-tag fusion constructs in RPE cells to pull down proteins interacting with either the WT or variant B form, followed by identification and quantification by mass spectrometry. We identified a total of 28 interacting proteins, of which 8 were exclusively pulled down by variant B cystatin C. These included 18 kDa translocator protein (TSPO) and cytochrome B5 type B, both of which are localized to the mitochondrial outer membrane. Variant B cystatin C expression also affected RPE mitochondrial function with increased membrane potential and susceptibility to damage-induced ROS production. The findings help us to understand how variant B cystatin C differs functionally from the WT form and provide leads to RPE processes adversely affected by the variant B genotype.