title: Direct reprogramming of fibroblasts to myocytes via bacterial injection of MyoD protein.
abstract: Forced exogenous gene expression has been well characterized as an effective method for directing both cellular differentiation and dedifferentiation. However, transgene expression is not amenable for therapeutic application due to potential insertional mutagenesis. Protein-based techniques provide a safe alternative, but current protein delivery methods are quite limited by labor-intensive purification processes, low protein yield, and inefficient intracellular targeting. Such limitations may be overcome by using a naturally occurring bacterial protein injection system, called the type III secretion system (T3SS), which injects bacterial proteins directly into the eukaryotic cell cytoplasm. Using a genetically attenuated strain of Pseudomonas aeruginosa, we have previously described the ability of this system to easily deliver a high quantity of protein to both differentiated and pluripotent cells. MyoD is a key muscle regulatory factor, the overexpression of which is able to induce transdifferentiation of numerous cell types into functional myocytes. Here we demonstrate transient injection of MyoD protein by P. aeruginosa to be sufficient to induce myogenic conversion of mouse embryonic fibroblasts. In addition to clear morphological changes, muscle-specific gene expression has been observed both at mRNA and protein levels. These studies serve as a foundation for the bacterial delivery of transcription factors to efficiently modulate concentration-dependent and temporal activation of gene expression that directs cell fate without jeopardizing genomic integrity.
authors: Department of Molecular Genetics and Microbiology, College of Medicine, University of Florida, Gainesville, FL 32610, USA.
authors: Candace
authors: C
authors: Bichsel
authors: Dennis
authors: D
authors: Neeld
authors: Takashi
authors: T
authors: Hamazaki
authors: Lung-Ji
authors: LJ
authors: Chang
authors: Li-Jun
authors: LJ
authors: Yang
authors: Naohiro
authors: N
authors: Terada
authors: Shouguang
authors: S
authors: Jin
mesh_terms: D000818:Animals; D058947:Bacterial Secretion Systems / Q000502:physiology; D002460:Cell Line; D054338:Cell Transdifferentiation; D005347:Fibroblasts / Q000378:metabolism* / Q000382:microbiology; D006801:Humans; D051379:Mice; D032342:Muscle Cells / Q000378:metabolism* / Q000382:microbiology; D017570:MyoD Protein / Q000096:biosynthesis* / Q000235:genetics; D011552:Pseudomonas Infections / Q000235:genetics / Q000378:metabolism*; D011550:Pseudomonas aeruginosa / Q000235:genetics / Q000378:metabolism*; D014161:Transduction, Genetic
chemical_list: D058947:Bacterial Secretion Systems; D017570:MyoD Protein; C057363:MyoD1 myogenic differentiation protein