MPRAnator: a web-based tool for the design of Massively Parallel Reporter Assay experiments

DNA regulatory elements contain short motifs where transcription factors (TFs) can bind to modulate gene expression. Although the broad principles of TF regulation are well understood, the rules that dictate how combinatorial TF binding translates into transcriptional activity remain largely unknown. With the rapid advances in DNA synthesis and sequencing technologies and the continuing decline in the associated costs, high-throughput experiments can be performed to investigate the regulatory role of thousands of oligonucleotide sequences simultaneously. Nevertheless, designing high-throughput reporter assay experiments such as Massively Parallel Reporter Assays (MPRAs) and similar methods remains challenging. We introduce MPRAnator, a set of tools that facilitate rapid design of MPRA experiments. With MPRA Motif design, a set of variables provides fine control of how motifs are placed into sequences therefore allowing the user to investigate the rules that govern TF occupancy. MPRA SNP design can be used to investigate the functional effects of single or combinations of SNPs at regulatory sequences. Finally, the Transmutation tool allows for the design of negative controls by permitting scrambling, reversing, complementing or introducing multiple random mutations in the input sequences or motifs.

[1]  N. Ahituv,et al.  Decoding enhancers using massively parallel reporter assays. , 2015, Genomics.

[2]  A. Stark,et al.  Uncovering cis-regulatory sequence requirements for context-specific transcription factor binding , 2012, Genome research.

[3]  Shane J. Neph,et al.  Systematic Localization of Common Disease-Associated Variation in Regulatory DNA , 2012, Science.

[4]  T. Mikkelsen,et al.  Systematic dissection of regulatory motifs in 2000 predicted human enhancers using a massively parallel reporter assay. , 2013, Genome research.

[5]  M. Berger,et al.  Universal protein-binding microarrays for the comprehensive characterization of the DNA-binding specificities of transcription factors , 2009, Nature Protocols.

[6]  R. Rohs,et al.  A widespread role of the motif environment in transcription factor binding across diverse protein families , 2015, Genome research.

[7]  E. Segal,et al.  The grammar of transcriptional regulation , 2014, Human Genetics.

[8]  E. Segal,et al.  In pursuit of design principles of regulatory sequences , 2014, Nature Reviews Genetics.

[9]  L. Dailey,et al.  High throughput technologies for the functional discovery of mammalian enhancers: new approaches for understanding transcriptional regulatory network dynamics. , 2015, Genomics.

[10]  William Stafford Noble,et al.  Sequence features and chromatin structure around the genomic regions bound by 119 human transcription factors , 2012, Genome research.

[11]  Z. Yakhini,et al.  Inferring gene regulatory logic from high-throughput measurements of thousands of systematically designed promoters , 2012, Nature Biotechnology.

[12]  Hao-Geng Hung,et al.  Discovering gapped binding sites of yeast transcription factors , 2008, Proceedings of the National Academy of Sciences.

[13]  J. Shendure,et al.  Massively parallel decoding of mammalian regulatory sequences supports a flexible organizational model , 2013, Nature Genetics.

[14]  Juan M. Vaquerizas,et al.  DNA-Binding Specificities of Human Transcription Factors , 2013, Cell.

[15]  Gautier Koscielny,et al.  Analysis of variation at transcription factor binding sites in Drosophila and humans , 2012, Genome Biology.

[16]  B. Cohen,et al.  Massively parallel synthetic promoter assays reveal the in vivo effects of binding site variants , 2013, Genome research.

[17]  B. Cohen,et al.  Massively parallel in vivo enhancer assay reveals that highly local features determine the cis-regulatory function of ChIP-seq peaks , 2013, Proceedings of the National Academy of Sciences.

[18]  E. Furlong,et al.  Combinatorial binding predicts spatio-temporal cis-regulatory activity , 2009, Nature.

[19]  Z. Yakhini,et al.  Unraveling determinants of transcription factor binding outside the core binding site , 2015, Genome research.

[20]  Byrappa Venkatesh,et al.  Mouse Transgenesis Identifies Conserved Functional Enhancers and cis-Regulatory Motif in the Vertebrate LIM Homeobox Gene Lhx2 Locus , 2011, PloS one.

[21]  Eran Segal,et al.  Probing the effect of promoters on noise in gene expression using thousands of designed sequences , 2014, Genome research.

[22]  S. Batzoglou,et al.  Linking disease associations with regulatory information in the human genome , 2012, Genome research.

[23]  Andrew R. Gehrke,et al.  Genome-wide analysis of ETS-family DNA-binding in vitro and in vivo , 2010, The EMBO journal.

[24]  Matthew Slattery,et al.  Absence of a simple code: how transcription factors read the genome. , 2014, Trends in biochemical sciences.