Browsing by Author "Palatinus, Joseph A."

Now showing 1 - 3 of 3
  • Thumbnail Image
    Diabetes Increases Cryoinjury Size with Associated Effects on Cx43 Gap Junction Function and Phosphorylation in the Mouse Heart
    Palatinus, Joseph A.; Gourdie, Robert G. (2016)
    Diabetic patients develop larger myocardial infarctions and have an increased risk of death following a heart attack. The poor response to myocardial injury in the diabetic heart is likely related to the many metabolic derangements from diabetes that create a poor substrate in general for wound healing, response to injury and infection. Studies in rodents have implicated a role for the gap junction protein connexin 43 (Cx43) in regulating the injury response in diabetic skin wounds. In this study, we sought to determine whether diabetes alters Cx43 molecular interactions or intracellular communication in the cryoinjured STZ type I diabetic mouse heart. We found that epicardial cryoinjury size is increased in diabetic mice and this increase is prevented by preinjury insulin administration. Consistent with these findings, we found that intercellular coupling via gap junctions is decreased after insulin administration in diabetic and nondiabetic mice. This decrease in coupling is associated with a concomitant increase in phosphorylation of Cx43 at serine 368, a residue known to decrease channel conductance. Taken together, our results suggest that insulin regulates both gap junction-mediated intercellular communication and injury propagation in the mouse heart.
  • Thumbnail Image
    Interaction of α Carboxyl Terminus 1 Peptide With the Connexin 43 Carboxyl Terminus Preserves Left Ventricular Function After Ischemia‐Reperfusion Injury
    Jiang, Jingbo; Hoagland, Daniel T.; Palatinus, Joseph A.; He, Huamei; Iyyathurai, Jegan; Jourdan, L. Jane; Bultynck, Geert; Wang, Zhen; Zhang, Zhiwei; Schey, Kevin; Poelzing, Steven; McGowan, Francis X.; Gourdie, Robert G. (American Heart Association, 2019-08-19)
    Background α Carboxyl terminus 1 (αCT1) is a 25–amino acid therapeutic peptide incorporating the zonula occludens‐1 (ZO‐1)–binding domain of connexin 43 (Cx43) that is currently in phase 3 clinical testing on chronic wounds. In mice, we reported that αCT1 reduced arrhythmias after cardiac injury, accompanied by increases in protein kinase Cε phosphorylation of Cx43 at serine 368. Herein, we characterize detailed molecular mode of action of αCT1 in mitigating cardiac ischemia‐reperfusion injury. Methods and Results To study αCT1‐mediated increases in phosphorylation of Cx43 at serine 368, we undertook mass spectrometry of protein kinase Cε phosphorylation assay reactants. This indicated potential interaction between negatively charged residues in the αCT1 Asp‐Asp‐Leu‐Glu‐Iso sequence and lysines (Lys345, Lys346) in an α‐helical sequence (helix 2) within the Cx43‐CT. In silico modeling provided further support for this interaction, indicating that αCT1 may interact with both Cx43 and ZO‐1. Using surface plasmon resonance, thermal shift, and phosphorylation assays, we characterized a series of αCT1 variants, identifying peptides that interacted with either ZO‐1–postsynaptic density‐95/disks large/zonula occludens‐1 2 or Cx43‐CT, but with limited or no ability to bind both molecules. Only peptides competent to interact with Cx43‐CT, but not ZO‐1–postsynaptic density‐95/disks large/zonula occludens‐1 2 alone, prompted increased pS368 phosphorylation. Moreover, in an ex vivo mouse model of ischemia‐reperfusion injury, preischemic infusion only with those peptides competent to bind Cx43 preserved ventricular function after ischemia‐reperfusion. Interestingly, a short 9–amino acid variant of αCT1 (αCT11) demonstrated potent cardioprotective effects when infused either before or after ischemic injury. Conclusions Interaction of αCT1 with the Cx43, but not ZO‐1, is correlated with cardioprotection. Pharmacophores targeting Cx43‐CT could provide a translational approach to preserving heart function after ischemic injury.
  • Thumbnail Image
    Protective mitochondrial fission induced by stress-responsive protein GJA1-20k
    Shimura, Daisuke; Nuebel, Esther; Baum, Rachel; Valdez, Steven E.; Xiao, Shaohua; Warren, Junco S.; Palatinus, Joseph A.; Hong, TingTing; Rutter, Jared; Shaw, Robin M. (eLife, 2021-10-05)
    The Connexin43 gap junction gene GJA1 has one coding exon, but its mRNA undergoes internal translation to generate N-terminal truncated isoforms of Connexin43 with the predominant isoform being only 20 kDa in size (GJA1-20k). Endogenous GJA1-20k protein is not membrane bound and has been found to increase in response to ischemic stress, localize to mitochondria, and mimic ischemic preconditioning protection in the heart. However, it is not known how GJA1-20k benefits mitochondria to provide this protection. Here, using human cells and mice, we identify that GJA1-20k polymerizes actin around mitochondria which induces focal constriction sites. Mitochondrial fission events occur within about 45 s of GJA1-20k recruitment of actin. Interestingly, GJA1-20k mediated fission is independent of canonical Dynamin-Related Protein 1 (DRP1). We find that GJA1-20k-induced smaller mitochondria have decreased reactive oxygen species (ROS) generation and, in hearts, provide potent protection against ischemia-reperfusion injury. The results indicate that stress responsive internally translated GJA1-20k stabilizes polymerized actin filaments to stimulate non-canonical mitochondrial fission which limits ischemic-reperfusion induced myocardial infarction.