Fig. 5: co-transcriptional, isothermal formation of single-stranded RNA nanostars and peptide recruitment.
From: Modular RNA motifs for orthogonal phase separated compartments
a Scheme shows transcription and co-transcriptional folding of motif 3sv2; example images show condensate formation in T7 in vitro transcription reaction sampled at different time points. b Temporal evolution of the CLD (left) and number (N, right) of 3sv2 nanostars. Black lines indicate the mean chord length from n = 3 at given time points. c Scheme shows transcription and co-transcriptional folding of motif pairs 3sβ1 and 3sβ2; example images show 3sβ condensate growth in T7 in vitro transcription reaction. d Temporal evolution of the CLD of 3sβ condensates. Black lines indicate the mean chord length from n = 3 at given time points. (e) Example images show 3sv2 condensate growth in a cell-free PURExpress® reaction. f Temporal evolution of the CLD of 3sv2 condensate in PURExpress®. Black lines indicate the mean chord length from n = 3 at given time points. g Brightfield and fluorescence microscopy images show specific peptide recruitment to condensates formed co-transcriptionally after 2-hour incubation. 3sv2 nanostars were modified to include TAR (top) or boxB (bottom) aptamer, recruiting either TAT or p22 N peptide labeled with 6-FAM. Peptides are added at the beginning of transcription. Experiments were done in triplicates; images are representative examples. Scale bars: 10 μm.
