I received my Ph.D. from Baylor College of Medicine in Houston, TX in the Department of Cell Biology. After graduating from Baylor, I joined the laboratory of Dr. E.J. Benz, Jr., in the Division of Hematology, at Johns Hopkins University, School of Medicine in 1998, to study protein 4.1R, a peripheral membrane protein that was originally discovered in red blood cells, in skeletal muscle. In 2001, I moved to the laboratory of Dr. R.J. Bloch, in the Department of Physiology, at the University of Maryland, School of Medicine, as an Academic Fellow, to study a small form of ankyrin 1 (small ankyrin 1) and its ligands in skeletal muscle. In 2002, I was promoted to Research Associate, and in July of 2003 to Assistant Professor. My research has been funded by grants from the Muscular Dystrophy Association and the National Institutes of Health.
Obscurin (~800 kDa) is the third giant protein of the contractile apparatus identified in vertebrate striated muscle, along with titin (2-4 MDa) and nebulin (~800 kDa). Like its predecessors, obscurin is a multidomain protein composed of adhesion modules and signaling domains arranged mostly in tandem. Specifically, its NH2-terminus contains 54 Immunoglobulin-C2 (Ig-C2) and 2 Fibronectin-III (Fn-III) domains, followed by an IQ motif and a conserved SH3 domain adjacent to Rho-guanine nucleotide exchange factor (Rho-GEF) and pleckstrin homology (PH) domains (Fig. 1). The COOH-terminal end of the protein consists of 2 additional Ig domains followed by a non-modular region of ~420 amino acid residues that contains several copies of a consensus phosphorylation motif for ERK kinases, similar to that found in the NH2-terminal region of titin. The obscurin gene, obscurin-MLCK, also encodes two ser/thr kinase domains, but these are apparently not expressed as part of the ~800 kDa form of the protein, and instead are made as smaller, alternatively spliced products, mainly in heart.
- Investigation of the role of obscurin in myofibrillogenesis, using in vitro and in vivo models
- Elucidation of the role of obscurin in signaling, using functional genomics and bioinformatics tools as well as proteomics analysis
- Molecular identification and functional characterization of the small obscurin isoforms in skeletal and cardiac muscle cells, using modern molecular, cellular and biochemical methodologies
- Identification of genetic variations in the coding region of obscurin in patients with heart failure, and determination of the functional significance of potential polymorphisms in vitro using expression systems and in vivo using transgenic animal models
- Mutation screening of the small ankyrin 1 gene in patients with dilated cardiomyopathy, using Single Strand Conformation Polymorphism (SSCP) analysis (In collaboration with "The Biomedical Foundation of the Academy of Athens" and "The Onassion Hospital" Athens, Greece)
- Investigation of the functional significance of the identified mutations in vitro using expression systems, and in vivo by generating appropriate mouse models
HS-1 associated protein X-1 (HAX-1) is a ~32 kDa protein that was originally identified in a yeast two-hybrid screen on the basis of its binding to the hematopoietic cell specific protein 1 (HS-1). An NH2-terminal Bcl-Homology domain 1 (BH1) followed by a Bcl-Homology domain 2 (BH2), a PEST motif and a COOH-terminal transmembrane (TM) domain make up its structure (Fig. 4). HAX-1 is ubiquitously expressed among different tissues, although its relevant abundance varies.
- Identification of the motifs that target HAX-1 to the mitochondria versus the ER/SR membranes and evaluation of their anti-apoptotic properties
- Delineation of the molecular mechanisms through which HAX-1 exerts its anti-apoptotic activity using genomic and proteomic approaches
- Examination of the ability of endogenous HAX-1 to oscillate between the mitochondria and the ER/SR membranes under stress conditions
- Investigation of the anti-apoptotic capacity of HAX-1 in vivo using transplantation of genetically engineered myoblasts that overexpress HAX-1 in dystrophic muscles of SCID/mdx mice, followed by evaluation of their survival rate and regeneration capability
Lab Techniques and Equipment:
Molecular cloning, RT-PCR/Quantitative RT-PCR/Long Range RT-PCR/RACE-PCR, yeast two-hybrid screening, protein expression and purification, overexpression experiments via adenoviral-mediated gene delivery, small inhibitory RNA technology, immunofluorescent combined with confocal laser scanning microscopy, immunoelectron microscopy, tissue culture of skeletal and cardiac myotubes, cryosectioning of muscle tissue, in vitro binding assays, surface plasmon resonance, cDNA microarrays, mass spectrometry, generation of null and transgenic animal models, and in vivo transplantation experiments.
Postdoctoral Position Available:
A postdoctoral position is available immediately in Dr. Kontrogianni's laboratory to study the role of cytoskeletal proteins in muscle organization and activity. A variety of experimental approaches will be employed in these studies, including molecular (e.g., cloning, yeast-two-hybrid, RT-PCR/qRT-PCR, mutagenesis, siRNA, transgenic and knockdown models), cellular (tissue culture of skeletal and cardiac myotubes, cryosectioning of muscle tissue, immunocytochemistry combined with confocal microscopy and transfection/transduction), biochemical (surface plasmon resonance, in vitro binding assays, mass spectrometry) and genomic (cDNA and exon microarrays). State-of-the-art equipment and facilities are available. Experience in the techniques is highly desirable.
The postdoctoral fellow will be expected to be current in the literature in the field, to plan and conduct experiments, interpret data, and publish results. Applicants must possess a PhD degree (or equivalent), should have excellent oral and written communication skills, and display initiative as well as independence. Salary is based upon experience and NIH salary levels. Please send a statement of research interests and previous experience, curriculum vitae, and the names and contact information of three references to: firstname.lastname@example.org.
- Solomon Yap (MD/PhD; Post-doctoral Fellow)
- Maegen Borzok (PhD; Post-doctoral Fellow)
- Rebecca Hu (Graduate Student)
Kontrogianni-Konstantopoulos, A, Huang, S.-C., and Benz, E.J., Jr., "A non-erythroid isoform of protein 4.1R interacts with components of the contractile apparatus in skeletal myofibers", Mol. Biol. Cell 11, 3805-3817, 2000.
Kontrogianni-Konstantopoulos, A., Frye, C., Benz, E.J. Jr., and Hung, S.-C. "The Prototypical 4.1R-10 kDa Domain and the 4.1G-10 kDa Paralog Mediate Fodrin/Actin Complex Formation", J. Biol. Chem. 276, 20679-20687, 2001.
Kontrogianni-Konstantopoulos, A., and Bloch, R.J., "The Hydrophilic Domain of Small Ankyrin 1 Interacts with the Two NH2-terminal Immunoglobulin Domains of Titin", J. Biol. Chem. 278, 3985-3991, 2003.
Kontrogianni-Konstantopoulos, A., Jones, E., van Rossum, D., and Bloch, R.J., "Obscurin is a Ligand for Small Ankyrin 1 in Skeletal Muscle" Mol. Biol. Cell 14, 1138-1148, 2003.
Borisov, A.B., Raeker, M.O., Kontrogianni-Konstantopoulos, A., Yang, K., Kurnit D.M., Bloch, R.J., and Russel, M.W., "Rapid Response of Cardiac Obscurin Gene Cluster to Aortic Stenosis: Differential Expression of Obscurin and Obscurin-MLCK and Involvement in Hypertrophic Growth", Biochem. Biophys. Res. Commun. 310, 910-918, 2003.
Kontrogianni-Konstantopoulos, A., Catino, D.H., Strong, J.C., Randall, W.R. and Bloch, R.J., "Obscurin regulates the organization of myosin into A-bands", Am. J. Physiol. Cell Physiol., 287, C209-C217, 2004.
Borisov, A.B, Kontrogianni-Konstantopoulos, A., Bloch, R.J., Westfall, M.V. and Russell, M.W., "Dynamics of obscurin localization during differentiation and remodeling of cardiac myocytes: Obscurin as an integrator of myofibrillar structure", J. Histochem. Cytochem. 52 (9), 1117-1127, 2004.
Sanoudou, D., Vafiadaki, E., Arvanitis, D., Kranias, E.G. and Kontrogianni-Konstantopoulos, A., "Array Lessons from the Heart: Focus on the Genome and Transcriptome of Cardiomyopathies", Physiol. Genomics, 21, 131-143, 2005.
Borisov, A.B., Sutter, S.B., Kontrogianni-Konstantopoulos, A., Bloch, R.J., Westfall, M. V. and Russell, M.W., "Essential role of obscurin in cardiac myofibrillogenesis and hypertrophic response: evidence from small interfering RNA-mediated gene silencing"; Histochem. Cell Biol., 1-12, 2005.
Kontrogianni-Konstantopoulos, A., Catino, D.H., Strong, J.C., and Bloch, R.J. "De Novo Myofibrillogenesis in C2C12 Cells: Evidence for the Independent Assembly of M-lines and Z-disks" Am. J. Physiol. Cell Physiol., 290, 626-637, 2006.
Kontrogianni-Konstantopoulos, A. and Bloch, R.J., "Obscurin: a multitasking muscle giant", J. Muscle Res. Cell Motility, 26, 419-426, 2006.
Raeker, M., Su, F., Sutter, S., Kontrogianni-Konstantopoulos, A., Borisov, A.B., Lyons, S.E., Russell, M.W., "Obscurin Is Required For the Lateral Alignment of Striated Myofibrils In Zebrafish", Dev. Dynamics, 235, 2018-2029, 2006.
Kontrogianni-Konstantopoulos, A., Catino, D.H., Strong, J.C., Sutter, S., Borisov, A.B., Pumplin, D.W., Russell, M.W., and Bloch, R.J. "Obscurin Modulates the Assembly and Organization of both the Myofibril and the Sarcoplasmic Reticulum" FASEB J., 20, 2102-2111, 2006.
Vafiadaki, E., Sanoudou, D., Arvanitis, D., Catino, D.H., Kranias, E.G., and Kontrogianni-Konstantopoulos, A., "Phospholamban interacts with HAX-1, a mitochondrial protein with anti-apoptotic function". J. Mol. Biol., 367, 65-79, 2007.
Bowman, A.L., Kontrogianni-Konstantopoulos, A, Hirsch, H., Geisler, S., Gonzalez-Serratos, H., Russell, M.W. and Bloch, R.J., "Different obscurin isoforms localize to distinct sites at sarcomeres". FEBS Letters, 581, 1549-54, 2007.
Arvanitis, D.A., Vafiadaki, E., Mitton, B.A., Gregory, K.N., Kontrogianni-Konstantopoulos, A., Sanoudou, D., and Kranias E.G., "Histidine-rich calcium binding protein is a novel binding partner of SERCA2a in the heart". Am. J. Physiol.-Cell Physiol. 293(3):H1581-9, 2007.
Borzok, M.A., Catino, D.H., Nicholson, J., Kontrogianni-Konstantopoulos, A., and Bloch, R.J., "Mapping the Binding Site on Small Ankyrin 1 for Obscurin", J. Biol. Chem. 282(44), 32384-96, 2007.
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