Cytosolic iron-sulfur (Fe-S) cluster assembly (CIA) pathway delivers Fe-S clusters to nuclear and cytosolic Fe-S proteins involved in essential cellular functions. This delivery process is regulated by availability of iron and oxygen, but it remains unclear how CIA components orchestrate the cluster transfer under varying cellular environments. Here, we investigated the organization of CIA machinery under various conditions. We developed a targeted proteomics assay monitoring known CIA factors. Using this assay, we were able to detect NUBP1, CIAO3 and CIA substrates in immunoprecipitates of NUBP2, a component of the CIA scaffold complex. We also revealed that NUBP2 transiently associates with the CIA targeting complex (MMS19, CIAO1, CIAO2B), indicating the possible existence of CIA metabolons. We observed stronger interactions between CIAO3 and the CIA scaffold complex upon iron supplementation or low oxygen tension, while iron chelation and reactive oxygen species weaken CIAO3 interactions with CIA components. We further demonstrated that CIAO3 mutant with defective Fe-S cluster binding failed to integrate into the metabolons. However, these mutants unexpectedly exhibit stronger association with CIA substrates regardless of their reduced association with the CIA targeting complex, implicating that CIAO3 and CIA substrates may present in complexes independent of the CIA targeting complex. Together, our data suggested that CIA components potentially form metabolons whose assembly are regulated by environmental cues and require Fe-S cluster incorporation in CIAO3. These findings provided additional evidence that the CIA pathway adapts to changes in cellular environment through complex reorganization.
[doi:10.25345/C5T85M]
[dataset license: CC0 1.0 Universal (CC0 1.0)]
Keywords: iron-sulfur cluster biogenesis ; targeted proteomics assay ; protein-protein interaction
Principal Investigators: (in alphabetical order) |
James A Wohlschlegel, UCLA Biological Chemistry, United States |
Submitting User: | xyztl |
Xiaorui Fan, William D. Barshop, Ajay A. Vashisht, Vijaya Pandey, Stephanie Leal, Shima Rayatpisheh, Yasaman Jami-Alahmadi, Jihui Sha, James A. Wohlschlegel.
Iron-Regulated Assembly of the Cytosolic Iron-Sulfur Cluster Biogenesis Machinery.
Fan X, Barshop WD, Vashisht AA, Pandey V, Leal S, Rayatpisheh S, Jami-Alahmadi Y, Sha J, Wohlschlegel JA (2022). Iron-Regulated Assembly of the Cytosolic Iron-Sulfur Cluster Biogenesis Machinery. J Biol Chem. DOI: 10.1016/j.jbc.2022.102094.
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