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Dissecting splicing decisions and cell-to-cell variability with designed sequence libraries

By Martin Mikl, Amit Hamburg, Yitzhak Pilpel, Eran Segal

Posted 16 Aug 2018
bioRxiv DOI: 10.1101/392605 (published DOI: 10.1038/s41467-019-12642-3)

Most human genes are alternatively spliced, allowing for a large expansion of the proteome. The multitude of regulatory inputs to splicing limits the potential to infer general principles from investigating native sequences. Here, we created a rationally designed library of >32,000 splicing events to dissect the complexity of splicing regulation through systematic sequence alterations. Measuring RNA and protein splice isoforms allowed us to investigate both cause and effect of splicing decisions, quantify diverse regulatory inputs and accurately predict (R2=0.75-0.85) isoform ratios from sequence and secondary structure. By profiling individual cells, we measure the cell-to-cell variability of splicing decisions and show that it can be encoded in the DNA and influenced by regulatory inputs, opening the door for a novel, single-cell perspective on splicing regulation.

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