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Post‐translational modifications frequently modulate protein functions. Lysine acetylation in particular plays a key role in interactions between respiratory cytochrome c and its metabolic partners. To date, in vivo acetylation of lysines at positions 8 and 53 has specifically been identified in mammalian cytochrome c, but little is known about the structural basis of acetylation‐induced functional changes. Here, we independently replaced these two residues in recombinant human cytochrome c with glutamine to mimic lysine acetylation and then characterized the structure and function of the resulting K8Q and K53Q mutants. We found that the physicochemical features were mostly unchanged in the two acetyl‐mimetic mutants, but their thermal stability was significantly altered. NMR chemical shift perturbations of the backbone amide resonances revealed local structural changes, and the thermodynamics and kinetics of electron transfer in mutants immobilized on gold electrodes showed an increase in both protein dynamics and solvent involvement in the redox process. We also observed that the K8Q (but not the K53Q) mutation slightly increased the binding affinity of cytochrome c to its physiological electron donor, cytochrome c1—which is a component of mitochondrial complex III, or cytochrome bc1—thus suggesting that Lys8 (but not Lys53) is located in the interaction area. Finally, the K8Q and K53Q mutants exhibited reduced efficiency as electron donors to complex IV, or cytochrome c oxidase. [ABSTRACT FROM AUTHOR]

Despite mitochondria being key for the control of cell homeostasis and fate, their role in DNA damage response is usually just regarded as an apoptotic trigger. However, growing evidence points to mitochondrial factors modulating nuclear functions. Remarkably, after DNA damage, cytochrome c (Cc) interacts in the cell nucleus with a variety of well‐known histone chaperones, whose activity is competitively inhibited by the haem protein. As nuclear Cc inhibits the nucleosome assembly/disassembly activity of histone chaperones, it might indeed affect chromatin dynamics and histone deposition on DNA. Several histone chaperones actually interact with Cc Lys residues through their acidic regions, which are also involved in heterotypic interactions leading to liquid–liquid phase transitions responsible for the assembly of nuclear condensates, including heterochromatin. This relies on dynamic histone–DNA interactions that can be modulated by acetylation of specific histone Lys residues. Thus, Cc may have a major regulatory role in DNA repair by fine‐tuning nucleosome assembly activity and likely nuclear condensate formation. [ABSTRACT FROM AUTHOR]

SUMMARY: Programmed cell death (PCD) is crucial for development and homeostasis of all multicellular organisms. In human cells, the double role of extra‐mitochondrial cytochrome c in triggering apoptosis and inhibiting survival pathways is well reported. In plants, however, the specific role of cytochrome c upon release from the mitochondria remains in part veiled yet death stimuli do trigger cytochrome c translocation as well. Here, we identify an Arabidopsis thaliana 14‐3‐3ι isoform as a cytosolic cytochrome c target and inhibitor of caspase‐like activity. This finding establishes the 14‐3‐3ι protein as a relevant factor at the onset of plant H2O2‐induced PCD. The in vivo and in vitro studies herein reported reveal that the interaction between cytochrome c and 14‐3‐3ι exhibits noticeable similarities with the complex formed by their human orthologues. Further analysis of the heterologous complexes between human and plant cytochrome c with plant 14‐3‐3ι and human 14‐3‐3ε isoforms corroborated common features. These results suggest that cytochrome c blocks p14‐3‐3ι so as to inhibit caspase‐like proteases, which in turn promote cell death upon H2O2 treatment. Besides establishing common biochemical features between human and plant PCD, this work sheds light onto the signaling networks of plant cell death. Significance Statement: Common features of the cytochrome c‐dependent pathways leading to programmed cell death in plants and humans are herein revealed. In response to oxidative stress, cytochrome c is released from mitochondria to the cytoplasm to hamper the iota isoform of the 14‐3‐3 protein family, thereby decreasing the inhibition of caspase‐like activity and likely contributing to promote cell death in plants. [ABSTRACT FROM AUTHOR]

Cytochrome c (Cc) is a protein that functions as an electron carrier in the mitochondrial respiratory chain. However, Cc has moonlighting roles outside mitochondria driving the transition of apoptotic cells from life to death. When living cells are damaged, Cc escapes its natural mitochondrial environment and, once in the cytosol, it binds other proteins to form a complex named the apoptosome—a platform that triggers caspase activation and further leads to controlled cell dismantlement. Early released Cc also binds to inositol 1,4,5‐triphosphate receptors on the ER membrane, which stimulates further massive Cc release from mitochondria. Besides the well‐characterized binding proteins contributing to the proapoptotic functions of Cc, many novel protein targets have been recently described. Among them, histone chaperones were identified as key partners of Cc following DNA breaks, indicating that Cc might modulate chromatin dynamics through competitive binding to histone chaperones. In this article, we review the ample set of recently discovered antiapoptotic proteins—involved in DNA damage, transcription, and energetic metabolism—reported to interact with Cc in the cytoplasm and even the nucleus upon DNA breaks. [ABSTRACT FROM AUTHOR]