Deconvolution of Nascent Sequencing Data Using Transcriptional Regulatory Elements.

ABSTRACT

The problem of microdissection of heterogeneous tissue samples is of great interest for both fundamental biology and biomedical research. Until now, microdissection in the form of supervised deconvolution of mixed sequencing samples has been limited to assays measuring gene expression (RNA-seq) or chromatin accessibility (ATAC-seq). We present here the first attempt at solving the supervised deconvolution problem for run-on nascent sequencing data (GRO-seq and PRO-seq), a readout of active transcription. Then, we develop a novel filtering method suited to the mixed set of promoter and enhancer regions provided by nascent sequencing, and apply best-practice standards from the RNA-seq literature, using in-silico mixtures of cells. Using these methods, we find that enhancer RNAs are highly informative features for supervised deconvolution. In most cases, simple deconvolution methods perform better than more complex ones for solving the nascent deconvolution problem. Furthermore, undifferentiated cell types confound deconvolution of nascent sequencing data, likely as a consequence of transcriptional activity over the highly open chromatin regions of undifferentiated cell types. Our results suggest that while the problem of nascent deconvolution is generally tractable, stronger approaches integrating other sequencing protocols may be required to solve mixtures containing undifferentiated celltypes.