Correction of multi-gene deficiency in vivo using a single 'self-cleaving' 2A peptide-based retroviral vector.
Attempts to generate reliable and versatile vectors for gene therapy and biomedical research that express multiple genes have met with limited success. Here we used Picornavirus 'self-cleaving' 2A peptides, or 2A-like sequences from other viruses, to generate multicistronic retroviral vectors with efficient translation of four cistrons. Using the T-cell receptor:CD3 complex as a test system, we show that a single 2A peptide-linked retroviral vector can be used to generate all four CD3 proteins (CD3epsilon, gamma, delta, zeta), and restore T-cell development and function in CD3-deficient mice. We also show complete 2A peptide-mediated 'cleavage' and stoichiometric production of two fluorescent proteins using a fluorescence resonance energy transfer-based system in multiple cell types including blood, thymus, spleen, bone marrow and early stem cell progenitors.
Szymczak
Andrea L
AL
Workman
Creg J
CJ
Wang
Yao
Y
Vignali
Kate M
KM
Dilioglou
Smaroula
S
Vanin
Elio F
EF
Vignali
Dario A A
DA
D000595:Amino Acid Sequence; D000818:Animals; D002460:Cell Line; D005822:Genetic Vectors; D006801:Humans; D051379:Mice; D008810:Mice, Inbred C57BL; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D012190:Retroviridae / Q000235:genetics*; D017386:Sequence Homology, Amino Acid
D010455:Peptides