SC COBRE in Oxidants, Redox Balance and Stress Signaling
|2012 - 2013|
| Sherine Chan, Ph.D.|
The role of mitochondrial DNA stability in the development of mitochondrial diseases
1. Opa1 is required for proper mitochondrial metabolism in early development
We recently published our work on a new zebrafish model of Opa1 mitochondrial dysfunction (Rahn et al., 2013), which was also chosen for an abstract talk at the Gordon Research Conference on Mitochondria and Chloroplasts. Opa1 catalyzes fusion of inner mitochondrial membranes and formation of the cristae. OPA1 mutations in humans lead to autosomal dominant optic atrophy. OPA1 knockout mice lose viability around embryonic day 9, indicating that OPA1 is essential for embryonic development. Zebrafish are an attractive model for studying vertebrate development and have been used for many years to describe developmental events that are difficult or impractical to view in mammalian models. In this study, Opa1 was successfully depleted in zebrafish embryos using antisense morpholinos, which resulted in disrupted mitochondrial morphology. Phenotypically, these embryos exhibited abnormal blood circulation and heart defects, as well as small eyes and small pectoral fin buds. Additionally, startle response was reduced and locomotor activity was impaired. Furthermore, Opa1 depletion caused defects in overall respiration without impairing mitochondrial efficiency. These results not only reveal a new Opa1-associated phenotype in a vertebrate model system, but also further elucidate the absolute requirement of Opa1 for successful vertebrate development.
|James Chou, Ph.D.|
Novel AML therapy targeting hdac6 and hsp90 chaperone complex
1. We have developed the first HDAC6 catalytic site 1 inhibitor targeting Hsp90 acetylation and its activity. Unlike classical HDAC inhibitors which only target the tubulin deacetylation site of HDAC6. The HDAC6 site 1 inhibitor increases Hsp90 acetylation which renders Hsp90 inactive resulting in Hsp90 oncogenic client protein degradation. The results were published in ACS Chemical Biology Feb 17, 2012.
2. We obtained R01 grant from NIH/NCI starting April 1st, 2013.
3. Provisional patent application filed. MUSC Foundation for Research Development on Nov. 11, 2012 (P1225 ; MESC.P0071US.P1) “Vitamin K derivatives for neuroprotection and treatment of epileptic seizures” Chou CJ, Chan SSL, Rahn J, and Josey BJ
Marcelo Vargas, Ph.D.
Although not directly related with the core subject of the project the following publications were made possible in part due to the financial support form the COBRE grant.
Vargas MR, Burton NC, Gan L, Johnson DA, Schäfer M, Werner S, Johnson JA. Absence of Nrf2 or Its Selective Overexpression in Neurons and Muscle Does Not Affect Survival in ALS-Linked Mutant hSOD1 Mouse Models. PLoS One. 2013;8(2):e56625. PMID: 23418589;
Pehar M, Puglielli L. Lysine acetylation in the lumen of the ER: a novel and essential function under the control of the UPR. Biochim Biophys Acta. 2013 Mar;1833(3):686-97. PMID: 23247107
Joachim Uys, Ph.D.
The National Survey on Drug Use and Health (NSDUH) estimates that in 2008 there were 1.9 million current (past-month) cocaine users. Cocaine use was responsible for 1 in 4 drug abuse related visits to the emergency room and a contributing factor in over 4800 deaths per year in the US. We have recently shown that the lack of the redox-sensitive enzyme, GSTP, contributes to cocaine sensitivity and cocaine preference in a preclinical model of cocaine sensitization. Interestingly, the gene encoding GSTP is known to be polymorphic in humans at exon 5 and exon 6, which has functional consequences resulting in reduced GST activity. In fact, one of these polymorphic variants, Ile105Val, has been associated with cocaine dependence. Our current studies are focusing on the mechanisms through which GSTP regulates cocaine sensitivity and modulates cocaine-induced redox sensitive proteins.
|2011 - 2012|
James Chou, Ph.D.
Dr. Chou has developed the first HDAC6 catalytic site 1 inhibitor targeting Hsp90 acetylation and its activity. Unlike classical HDAC inhibitors that only target the tubulin deacetylation site of HDAC6, the HDAC6 site 1 inhibitor increases Hsp90 acetylation rendering Hsp90 inactive resulting in Hsp90 oncogenic client protein degradation. Initial results were published in ACS Chemical Biology Feb 17, 2012.
Danyelle Townsend, Ph.D.
S-glutathionylation of protein disulfide isomerase regulates estrogen receptor α stability and function. S-glutathionylation of cysteine residues within target proteins is a post-translational modification that alters structure and function. Dr. Townsend showed that S-glutathionylation of protein disulfide isomerase (PDI) disrupts protein folding and leads to the activation of the unfolded protein response (UPR). PDI is a molecular chaperone for estrogen receptor alpha (ERα). Her present data show in breast cancer cells that S-glutathionylation of PDI interferes with its chaperone activity and abolishes its capacity to form a complex with ERα. Such drug treatments also reverse estradiol-induced up-regulation of c-Myc, cyclinD1 and p21Cip, - all gene products involved in cell proliferation. Expression of an S-glutathionylation refractory PDI mutant diminished the toxic effects of a nitric oxide releasing pro-drug. Her conclusions suggest that redox regulation of PDI causes its S-glutathionylation, thereby mediating cell death through activation of the UPR and abrogation of ERα stability and signaling. These data suggest that targeting PDI in ERα positive, tamoxifen resistant breast cancer patients might be a promising therapeutic approach.
Sherine Chan, Ph.D.
In the award period, Dr. Chan was invited by several universities in Australia to discuss her new method for measuring bioenergetics in a whole organism, the developing zebrafish embryo. This method offers researchers a new way to measure mitochondrial function in a whole animal model and is invaluable for identifying mitochondrial dysfunction in toxicological and genetic studies. As embryos are used, she can now use this method to answer questions in developmental biology, drug development and toxicology. She is also using this assay as one of the fundamental assays for her mitochondrial disease zebrafish model. This method was published during the COBRE award period (Stackley, 2011).
Craig Beeson, Ph.D.
Screening bioenergetics in developing zebrafish embryos
A central feature of the research endeavors of Sherine Chan, a COBRE investigator, has been to study the effects of mitochondrial-DNA (mtDNA) mutations on early zebrafish development. The zebrafish embryo is a good model for understanding events during an animal’s early development can result in neurodegeneration and cancer. Because mitochondrial dysfunction in the developing embryo was expected to manifest as aberrations in energy metabolism, the Metabolomics Core worked extensively to develop a metabolic assay using developing zebrafish embryos. To enable the assay, we used special “islet plates” that have a mini-well at the bottom and the mini-well has a lip upon which a small screen can be attached to prevent loss of contents. The oxygen consumption and acidification were measured at multiple time points from 3-48 h post fertilization and the data were subsequently analyzed to reveal, for example, ATP-coupled mitochondrial respiration and proton leak, which describe the efficiency of the mitochondria. All of these parameters were ultimately related back to mitochondrial protein expression levels and the embryo developmental state. The protocol and results of this novel assay have been recently published.