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Gene modification in primary human NK cells with Cas9 RNP electroporation - very useful preprint @biorxivpreprints - biorxiv.org/content/early/… Highlights and thoughts… #Immunology #GenomeEditing #CRISPR #PreReview #ASAPBio
The bottom line: Best RNP electroporation conditions resulted in up to 80% deletion, consistently >50%, of 3 tested genes for activated primary human NK cells. Validation of Ncr1 and Cish biology from mouse studies in human NK cells.
Paper starts by demonstrating issues with lentiviral transduction in human NK cells. Electroporation was optimized very thoroughly (>10 buffers and pulse conditions), with ~doubling in efficiency (viability and uptake) to >80% transformed and >80% viable
Very cool pair of new preprints @biorxivpreprints from @barton_lab on TLR regulation by Unc93b1 - biorxiv.org/content/early/… and biorxiv.org/content/early/… Here's my summary and a couple thoughts... #TLR #CellBiology #Immunology #PreReview #ASAPBio
First one, on TLR9 regulation by Unc93b1. Triple-alanine scanning mutagenesis library revealed mutant of Unc93b1 (SKN) that abolishes TLR9 signaling, although leaves endosomal trafficking and maturation intact. Single serine mutant (S282A) is responsible for full SKN phenotype
SKN or S282A both reduce amount of TLR9 binding to CpG. Both also increase amount of TLR9 bound by Unc93b1. Other mutations in same region of Unc93b1 (loop 5) either enhance or block Unc93b1 binding but all impair TLR9 activation, with much less effect on TLR7
