Molecular Systems Biology Peer Review Process File Regulation of Alternative Splicing at the Single-cell Level Transaction Report

(Note: With the exception of the correction of typographical or spelling errors that could be a source of ambiguity, letters and reports are not edited. The original formatting of letters and referee reports may not be reflected in this compilation.) Thank you for submitting your work to Molecular Systems Biology. We have now heard back from the three referees who agreed to evaluate your manuscript. As you will see from the reports below, the referees find the topic of your study of potential interest. They raise however several points, which should be convincingly addressed in a revision of this work. The comments made by the reviewers are very clear in this regard and it will be particularly important to address point 1 of reviewer #1 to ascertain the quality of the data and the correct interpretation of the source of variability observed in your data. If you feel you can satisfactorily deal with these points and those listed by the referees, you may wish to submit a revised version of your manuscript. Please attach a covering letter giving details of the way in which you have handled each of the points raised by the referees. A revised manuscript will be once again subject to review and you probably understand that we can give you no guarantee at this stage that the eventual outcome will be favorable. In the manuscript by Faigenbloom, using single cell isoform-specific qRT-PCR, the authors quantified the splicing pattern of 44 alternative exons in single cells from three distinct human cell lines, and correlated the variance of inclusion levels of alternative exons among individual cells with different features. Their results demonstrated that the precision of exon inclusion is mostly determined by the degree of evolutionary conservation of flanking intronic sequences. In addition, the precision is also affected by the inclusion level itself as well as the expression level of the specific transcripts. Finally, based on published single cell RNA sequencing data from human ES