Summary RNA molecules can attach to chromatin. It remains difficult to know what RNAs are associated with chromatin and where are the genomic target loci of these RNAs. Here, we present MARGI (Mapping RNA-genome interactions), a technology to massively reveal native RNA-chromatin interactions from unperturbed cells. The gist of this technology is to ligate chromatin associated RNAs (caRNAs) with their target genomic sequences by proximity ligation, forming RNA-DNA chimeric sequences, which are converted to sequencing library for paired-end sequencing. Using MARGI, we produced RNA-genome interaction maps for human embryonic stem (ES) cells and HEK cells. MARGI revealed hundreds of caRNAs including previously known XIST, SNHG1, NEAT1, MALAT1, as well as each caRNA's genomic interaction loci. Using a cross-species experiment, we estimated that approximately 2.2% of MARGI identified interactions were false positives. In ES and HEK cells, the RNA ends of more than 5% of MARGI read pairs were mapped to distal or inter-chromosomal locations as compared to the locations of their corresponding DNA ends. The majority of transcription start sites are associated with distal or inter-chromosomal caRNAs. ChIP-seq reported H3K27ac and H3K4me3 levels are positively while H3K9me3 is negatively correlated with MARGI reported RNA attachment levels. The MARGI technology should facilitate revealing novel RNA functions and their genomic target regions.

SummaryRNA molecules can attach to chromatin. It remains difficult to know what RNAs are associated with chromatin and where are the genomic target loci of these RNAs. Here, we present MARGI (Mapping RNA-genome interactions), a technology to massively reveal native RNA-chromatin interactions from unperturbed cells. The gist of this technology is to ligate chromatin associated RNAs (caRNAs) with their target genomic sequences by proximity ligation, forming RNA-DNA chimeric sequences, which are converted to sequencing library for paired-end sequencing. Using MARGI, we produced RNA-genome interaction maps for human embryonic stem (ES) cells and HEK cells. MARGI revealed hundreds of caRNAs including previously known XIST, SNHG1, NEAT1, MALAT1, as well as each caRNA's genomic interaction loci. Using a cross-species experiment, we estimated that approximately 2.2% of MARGI identified interactions were false positives. In ES and HEK cells, the RNA ends of more than 5% of MARGI read pairs were mapped to distal or inter-chromosomal locations as compared to the locations of their corresponding DNA ends. The majority of transcription start sites are associated with distal or inter-chromosomal caRNAs. ChIP-seq reported H3K27ac and H3K4me3 levels are positively while H3K9me3 is negatively correlated with MARGI reported RNA attachment levels. The MARGI technology should facilitate revealing novel RNA functions and their genomic target regions. Graphical abstractSridhar et al. develop a technology to map global RNA-chromatin interactions in unperturbed cells. They discover hundreds of chromatin associated RNAs. They find that the majority of Correspondence to: Sheng Zhong. 3 Co-first author 4 Lead Contact Supplemental Information: Supplemental Information includes Supplemental Experimental Procedures and four figures and can be found with this article online. All sequencing data are available at Gene Expression Omnibus with access number GSE92345.Author Contributions: B.S., T.C.N., and S.Z. designed the experiments. B.S., T.C.N., and L.H performed the experiments. All authors analyzed and interpreted the data.Publisher's Disclaimer: This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final citable form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain. HHS Public Access Results and Discussion Development of the MARGI technologyWe developed MARGI (Mapping RNA-genome interactions), a technology to massively reveal RNA-chromatin interactions from unperturbed cells. MARGI simultaneously identifies all caRNAs and the respective genomic target loci of each caRNA. This changes the paradigm of analyzing one-RNA-at-a-time, and enables the mapping of the native RNAchromatin interaction netwo...

No abstract

Department of Agriculture and the Florida Agricultural Experiment Stations and was released to the industry in 1957.C.P. 50-28 is an early-maturing, medium-barrel, early-flowering cultivar. It is adapted to the cold organic and sandy soils of Florida because ot~ its outstanding stubbling or ratooning ability. Although the fiber content of C.P. 50-28 is objectionable to some processors, the cold tolerance and exceptional stubbling qualities are distinct advantages of this cultivar.Seedcane of C.P. 50-28 wil be maintained by the U.S. Department of Agriculture at the U.S. Sugarcane Field Station, Canal Point, Fla. REGISTRATIONOF C.P. 56-59 SUGARCANE t (Reg. No. 7) C. O. Grassl~ E. R. Rice~ and L. P. Hebert The sugarcane cultivar "C.P. 56-59', a tri-species hybrid involving Saccharum o[[icinarum L., S. spontaneum L., and S. barberi Jeswiet, is a selection from the cross 'CI. 47-83' X 'C.P. 34-79.' The cross was made at Canal Point, Fla., during the 1956 crossing season. C.P. 56-59 was developed through the research efforts of the U.S. Department of Agriculture, the Florida Agricultural Experiment Stations, and the Florida Sugar Cane League, Inc., and was released to the industry in 1967.C.P. 56-59 is an early-maturing, medium-barrel, nonflowering cultivar that is recommended for the colddand area several miles from Lake Okeechobee and for early harvest on the warm-land areas near the Lake. The fiber content is less than from C.P. 50-28 but greater than from C1. 41-223, C.P. 56-59 produced slightly less sugar per ton of cane but 26% more sugar per acre than CI. 41-223, the most widely grown cultivar in Florida. C.P. 56-59 was moderately resistant to the mosaic disease in greenhouse inoculation tests.

Department of Agriculture and the Florida Agricultural Experiment Stations and was released to the industry in 1957.C.P. 50-28 is an early-maturing, medium-barrel, early-flowering cultivar. It is adapted to the cold organic and sandy soils of Florida because ot~ its outstanding stubbling or ratooning ability. Although the fiber content of C.P. 50-28 is objectionable to some processors, the cold tolerance and exceptional stubbling qualities are distinct advantages of this cultivar.Seedcane of C.P. 50-28 wil be maintained by the U.S. Department of Agriculture at the U.S. Sugarcane Field Station, Canal Point, Fla. REGISTRATIONOF C.P. 56-59 SUGARCANE t (Reg. No. 7) C. O. Grassl~ E. R. Rice~ and L. P. Hebert The sugarcane cultivar "C.P. 56-59', a tri-species hybrid involving Saccharum o[[icinarum L., S. spontaneum L., and S. barberi Jeswiet, is a selection from the cross 'CI. 47-83' X 'C.P. 34-79.' The cross was made at Canal Point, Fla., during the 1956 crossing season. C.P. 56-59 was developed through the research efforts of the U.S. Department of Agriculture, the Florida Agricultural Experiment Stations, and the Florida Sugar Cane League, Inc., and was released to the industry in 1967.C.P. 56-59 is an early-maturing, medium-barrel, nonflowering cultivar that is recommended for the colddand area several miles from Lake Okeechobee and for early harvest on the warm-land areas near the Lake. The fiber content is less than from C.P. 50-28 but greater than from C1. 41-223, C.P. 56-59 produced slightly less sugar per ton of cane but 26% more sugar per acre than CI. 41-223, the most widely grown cultivar in Florida. C.P. 56-59 was moderately resistant to the mosaic disease in greenhouse inoculation tests.