**10. Adenine nucleotide translocase (ANT)**

ANT, an antiporter embedded in the inner mitochondrial membrane that facilitates the exchange of ADP and ATP, is one of the most abundant mitochondrial proteins and a primary target of ischemia [5, 53]. The four human isoforms, ANT1, 2, 3, and 4, are phosphorylated on tyrosines [49]. Phosphorylation of ANT on Y194 and Y190 alters the activity of ADP/ATP translocase and the transport of both nucleotides in the brain [49]. Phosphorylation of ANT1 at Y194 in cardiac tissue is diminished by I/R but maintained by pre- and post-conditioning, which suggests that this phosphorylation plays a protective role against ischemia in the heart and could improve tolerance against injury [54]. Although this phosphorylation improves respiration and mitochondrial function, it is not known whether phosphorylation of Y194 is sufficient for protection of cardiomyocytes from I/R-induced injury [54]. A binding of phosphorylated (inactive) glycogen synthase kinase-3β (GSK-3β) to ANT was also shown, but it is unclear whether GSK-3β directly phosphorylates ANT [55].

a large complex. PDC is the rate-limiting enzyme in the oxidative metabolism of all carbohydrates and its activity is tightly regulated by multiple mechanisms including phosphorylation. Phosphorylation of the PDC by the pyruvate dehydrogenase kinase (PDK) inactivates the complex [64]. The α-subunit of the E1 is phosphorylated at multiple sites, but the most known are phosphorylations of three distinct serines [64]. Although phosphorylation at any of these serines is sufficient to inhibit the activity of PDC, the S293 residue has the highest affinity for phosphate, and phosphorylation of this serine has the greatest impact on the inhibition of activity of PDC [64, 65]. I/R-induced injury in cardiac tissue is associated with a 4–5 fold decrease in the phosphorylation of S293, which results in the activation of PDC [64]. PDC activation protects against ischemic injury and improves cardiac efficiency and contractile

Protein Kinases and Regulation of Mitochondrial Function in Ischemia/Reperfusion Injury

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Mitochondrial morphology is dynamically changed by the balance between fusion and fission (fragmentation). Phosphorylations of GTPases, Drp1, Mfn1/2, and Opa1 regulate mitochondrial fission and fusion [66]. Phosphorylation of fission-inducing Drp1 on S616 and S637 can be mediated by cyclin-dependent kinase (cdk)-1, cdk-5, PKA, PKC-δ, and extracellular signal regulated kinase-2 (ERK-2) [66–71]. Phosphorylation of Drp1 at S616 by cdk-1, PKC-δ, and ERK-2 promotes Drp1 translocation to mitochondria and fission whereas phosphorylation of S637 by PKA inhibits Drp1 and mitochondrial fission [67–71]. We have demonstrated that activation of PKC-ε induces the translocation of Drp1 to mitochondria, mitochondrial fission and apoptosis in non-injured RPTC [72]. Drp1-dependent mitochondrial fission is associated with mitochondrial outer membrane permeabilization and apoptosis, whereas elongation of the mitochondria through fusion promotes ATP synthesis and prevents mitochondrial autophagy. When phosphorylated by ERK, Mfn1 has decreased ability to oligomerize and tether mitochondria, which inhibits their fusion, recruitment of Bak to the mitochondria, and apoptosis [73]. Mfn2 is phosphorylated on T111 and S442 by PTEN-induced putative kinase protein 1 (PINK1), which recruits the protein Parkin to depolarized mitochondria and eliminates them by mitophagy [74]. Disruption in the balance between fission and fusion occurs in pathological conditions including I/R, which increases phosphorylation of Drp1, its translocation to mitochondria, and mitochondrial fission in the heart and brain [75]. Inhibition of Drp1 protects the heart and brain against ischemia and has been proposed as a therapeutic target following cardiac arrest [76].

Mitochondrial proteins involved in the intrinsic apoptosis include Bcl-2 family members, AIF, Smac/DIABLO, cytochrome c, and Omi/Htra2. With an exception of one, all are regulated by phosphorylation [20]. Phosphorylation of Bcl-2 at S70 is required to prevent permeabilization of the mitochondrial outer membrane and for Bcl-2's anti-apoptotic activity [77]. Several protein kinases serve as Bcl-2 kinases to inhibit (JNK, p38, and GSK-3) or activate (Akt, PKA, and PKC-α) Bcl-2 [77]. Cardiac and neuronal tissues are protected against I/R when Bcl-2 is active [78–80]. Ischemia-induced acute kidney injury upregulates Bcl-2 and Bcl-XL in the distal and Bax in the proximal tubules [80]. Pro-apoptotic Bax, Bak, Bad, and Bid are phosphorylated on serines and tyrosines, which controls their insertion into the outer mitochondrial membrane

capacity in the postischemic heart [65].

**11.3. Proteins involved in mitochondrial dynamics**

**11.4. Proteins of the intrinsic apoptotic cascade**
