Our research seeks to define the function of the dystrophin in normal tissues in order to explain how its absence or abnormality leads to the pathologies observed in several muscular dystrophies and some cardiomyopathies. Dystrophin has been speculatively modeled as an antiparallel, rod-shaped dirner that crosslinks actin filaments by way of a putative actin-binding domain located at the amino-terminus of each dystrophin monomer. However, we have obtained biochemical evidence indicating that dystrophin exists as a monomer that binds alongside an actin filament via multiple contact points distributed throughout the amino-terminal and rod domains (Rybakova et al., 1996 J. Cell BioL, In Press). Our findings suggest that although dystrophin is -most closely related to the actin crosslinking superfamily of proteins based on sequence homology, dystrophin functions more like an actin side binding protein such as tropomyosin. To further test the validity of this novel model for the interaction of dystrophin with F-actin, it is now necessary to obtain ultrastructural data on dystrophin when complexed with F-actin. Thus, we propose to visualize dystrophin/F-actin complexes by high resolution cryo scanning electron microscopy. We also propose to tag purified dystrophin with colloidal gold or with available site-specific antibodies in order to identify it in EM images. The results of this study are expected to provide the first convincing images of intact dystrophin and provide an independent test of our novel side-binding model for the interaction of dystrophin with F-actin.

Agency
National Institute of Health (NIH)
Institute
National Center for Research Resources (NCRR)
Type
Biotechnology Resource Grants (P41)
Project #
3P41RR000570-28
Application #
6278473
Study Section
Project Start
1998-07-01
Project End
2000-06-30
Budget Start
1997-10-01
Budget End
1998-09-30
Support Year
28
Fiscal Year
1998
Total Cost
Indirect Cost
Name
University of Wisconsin Madison
Department
Type
DUNS #
161202122
City
Madison
State
WI
Country
United States
Zip Code
53715
Malecki, Marek; Putzer, Emily; Sabo, Chelsea et al. (2014) Directed cardiomyogenesis of autologous human induced pluripotent stem cells recruited to infarcted myocardium with bioengineered antibodies. Mol Cell Ther 2:
Malecki, Marek (2014) 'Above all, do no harm': safeguarding pluripotent stem cell therapy against iatrogenic tumorigenesis. Stem Cell Res Ther 5:73
Mavroudi, Maria; Zarogoulidis, Paul; Porpodis, Konstantinos et al. (2014) Stem cells' guided gene therapy of cancer: New frontier in personalized and targeted therapy. J Cancer Res Ther (Manch) 2:22-33
Malecki, Marek; LaVanne, Christine; Alhambra, Dominique et al. (2013) Safeguarding Stem Cell-Based Regenerative Therapy against Iatrogenic Cancerogenesis: Transgenic Expression of DNASE1, DNASE1L3, DNASE2, DFFB Controlled By POLA1 Promoter in Proliferating and Directed Differentiation Resisting Human Autologous Pluripotent J Stem Cell Res Ther Suppl 9:
Malecki, Marek; Tombokan, Xenia; Anderson, Mark et al. (2013) TRA-1-60(+), SSEA-4(+), POU5F1(+), SOX2(+), NANOG(+) Clones of Pluripotent Stem Cells in the Embryonal Carcinomas of the Testes. J Stem Cell Res Ther 3:
Malecki, Marek (2013) Improved targeting and enhanced retention of the human, autologous, fibroblast-derived, induced, pluripotent stem cells to the sarcomeres of the infarcted myocardium with the aid of the bioengineered, heterospecific, tetravalent antibodies. J Stem Cell Res Ther 3:
Malecki, Marek; Dahlke, Jessica; Haig, Melissa et al. (2013) Eradication of Human Ovarian Cancer Cells by Transgenic Expression of Recombinant DNASE1, DNASE1L3, DNASE2, and DFFB Controlled by EGFR Promoter: Novel Strategy for Targeted Therapy of Cancer. J Genet Syndr Gene Ther 4:152
Zarogoulidis, Paul; Darwiche, Kaid; Sakkas, Antonios et al. (2013) Suicide Gene Therapy for Cancer - Current Strategies. J Genet Syndr Gene Ther 4:
Malecki, Marek; Sabo, Chelsea; Putzer, Emily et al. (2013) Recruitment and retention of human autologous CD34+ CD117+ CD133+ bone marrow stem cells to infarcted myocardium followed by directed vasculogenesis: Novel strategy for cardiac regeneration. Mol Cell Ther 1:
Malecki, Marek; Malecki, Bianca (2012) Routing of Biomolecules and Transgenes' Vectors in Nuclei of Oocytes. J Fertili In Vitro 2012:108-118

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