Use of Mutated Self-Cleaving 2A Peptides as a Molecular Rheostat to Direct Simultaneous Formation of Membrane and Secreted Anti-HIV ImmunoglobulinsReportar como inadecuado




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In nature, B cells produce surface immunoglobulin and secreted antibody from the same immunoglobulin gene via alternative splicing of the pre-messenger RNA. Here we present a novel system for genetically programming B cells to direct the simultaneous formation of membrane-bound and secreted immunoglobulins that we term a -Molecular Rheostat-, based on the use of mutated -self-cleaving- 2A peptides. The Molecular Rheostat is designed so that the ratio of secreted to membrane-bound immunoglobulins can be controlled by selecting appropriate mutations in the 2A peptide. Lentiviral transgenesis of Molecular Rheostat constructs into B cell lines enables the simultaneous expression of functional b12-based IgM-like BCRs that signal to the cells and mediate the secretion of b12 IgG broadly neutralizing antibodies that can bind and neutralize HIV-1 pseudovirus. We show that these b12-based Molecular Rheostat constructs promote the maturation of EU12 B cells in an in vitro model of B lymphopoiesis. The Molecular Rheostat offers a novel tool for genetically manipulating B cell specificity for B-cell based gene therapy.



Autor: Kenneth K. Yu, Kiefer Aguilar, Jonathan Tsai, Rachel Galimidi, Priyanthi Gnanapragasam, Lili Yang, David Baltimore

Fuente: http://plos.srce.hr/



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