Abstract
AbstractCytochromec(Cytc) is a multifunctional protein, acting as an electron carrier in the electron transport chain (ETC), where it shuttles electrons frombc1complex to cytochromecoxidase (COX), and as a trigger of type II apoptosis when released from the mitochondria. We previously showed that Cytcis regulated in a highly tissue-specific manner: Cytcisolated from heart, liver, and kidney is phosphorylated on Y97, Y48, and T28, respectively. Here, we have analyzed the effect of a new Cytcphosphorylation site, threonine 58, which we mapped in rat kidney Cytcby mass spectrometry. We generated and overexpressed wild-type, phosphomimetic T58E, and two controls, T58A and T58I Cytc; the latter replacement is found in human and testis-specific Cytc.In vitro, COX activity, caspase-3 activity, and heme degradation in the presence of H2O2were decreased with phosphomimetic Cytccompared to wild-type. Cytc-knockout cells expressing T58E or T58I Cytcshowed a reduction in intact cell respiration, mitochondrial membrane potential (∆Ψm), ROS production, and apoptotic activity compared to wild-type. We propose that, under physiological conditions, Cytcis phosphorylated, which controls mitochondrial respiration and apoptosis. Under conditions of stress Cytcphosphorylations are lost leading to maximal respiration rates, ∆Ψmhyperpolarization, ROS production, and apoptosis.
Funder
U.S. Department of Health & Human Services | NIH | National Institute of General Medical Sciences
Publisher
Springer Science and Business Media LLC
Reference49 articles.
1. Li, K. et al. Cytochrome c deficiency causes embryonic lethality and attenuates stress-induced apoptosis. Cell 101, 389–399 (2000).
2. Morriss, G. M. & New, D. A. Effect of oxygen concentration on morphogenesis of cranial neural folds and neural crest in cultured rat embryos. J Embryol Exp Morphol 54, 17–35 (1979).
3. Green, D. R. & Reed, J. C. Mitochondria and apoptosis. Science 281, 1309–1312 (1998).
4. Zou, H., Li, Y., Liu, X. & Wang, X. An APAF-1.cytochrome c multimeric complex is a functional apoptosome that activates procaspase-9. J Biol Chem 274, 11549–11556 (1999).
5. Kagan, V. E. et al. Cytochrome c acts as a cardiolipin oxygenase required for release of proapoptotic factors. Nat Chem Biol 1, 223–232, https://doi.org/10.1038/nchembio727 (2005).