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PreprintCited by 6Mitochondrial Function and PathologyAdvanced Proteomics Techniques and ApplicationsBiotin and Related Studies

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Let’s make it clear: Systematic exploration of mitochondrial DNA- and RNA-protein complexes by complexome profiling

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Potter, Alisa, Cabrera-Orefice, Alfredo, Spelbrink, Johannes N.·2023·Nucleic Acids Research
DOI 10.1101/2023.04.03.534993PMC PMC10602928

Preprint — published as 10.1093/nar/gkad697 · Paper page

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Abstract

Complexome profiling (CP) is a powerful tool for systematic investigation of protein interactors that has been primarily applied to study the composition and dynamics of mitochondrial protein complexes. Here, we further optimised this method to extend its application to survey mitochondrial DNA- and RNA-interacting protein complexes. We established that high-resolution clear native gel electrophoresis (hrCNE) is a better alternative to preserve DNA- and RNA-protein interactions that are otherwise disrupted when samples are separated by the widely used blue native gel electrophoresis (BNE). In combination with enzymatic digestion of DNA, our CP approach improved the identification of a wide range of protein interactors of the mitochondrial gene expression system without compromising the detection of other multi-protein complexes. The utility of this approach was particularly demonstrated by analysing the complexome changes in human mitochondria with impaired gene expression after transient, chemically-induced mtDNA depletion. Effects of RNase on mitochondrial protein complexes were also evaluated and discussed. Overall, our adaptations significantly improved the identification of mitochondrial DNA- and RNA-protein interactions by CP, thereby unlocking the comprehensive analysis of a near-complete mitochondrial complexome in a single experiment.

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Metadata

Authors
Potter, A., Cabrera-Orefice, A., & Spelbrink, J. N.
Affiliations
Radboud University Medical Center (NL)
Corresponding author
Spelbrink, Johannes N.
ORCID
Venue
Nucleic Acids Research
Publisher
openRxiv
Published
2023-04-03
Volume
51
Issue
19
Pages
10619-10641
ISSN
0305-1048 (ISSN-L)
DOI
10.1101/2023.04.03.534993
PMC
PMC10602928
OpenAlex
W4386116234
Cited by
6
References
1
Citations by year
2026: 2 · 2025: 2 · 2024: 1 · 2023: 1
FWCI
0.53
Type
journal article
Language
en
Open access
Licence
cc-by-nc
Locations
  • Nucleic Acids Research (publishedVersion, cc-by-nc)
  • bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nc-nd)
  • PubMed Central (publishedVersion, cc-by-nc)
  • PubMed (publishedVersion)
  • Publication Server of Goethe University Frankfurt am Main (Goethe University Frankfurt) (submittedVersion, cc-by-nc-nd)
  • Radboud Repository (Radboud University) (submittedVersion)
Funders
  • European Commission (721757)
  • Deutsche Forschungsgemeinschaft (TRR_267, SFB1531, 403584255, 456687919)
  • ZonMw (403584255, 91217009, ZonMW TOP 91217009, 456687919)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Mitochondrial Function and Pathology
  • Physical Sciences › Chemistry › Spectroscopy › Advanced Proteomics Techniques and Applications
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Biotin and Related Studies
Keywords
complexome profiling, mitochondrial protein complexes, mitochondrial gene expression, mitochondrial DNA depletion, human mitochondria
MeSH terms
RNA, Mitochondrial, DNA, Mitochondrial, Humans, Mitochondria, Mitochondrial Proteins, Proteomics

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