Abstract
BackgroundCompared to proteins, glycans, and lipids, much less is known about RNAs on the cell surface. We develop a series of technologies to test for any nuclear-encoded RNAs that are stably attached to the cell surface and exposed to the extracellular space, hereafter called membrane-associated extracellular RNAs (maxRNAs).ResultsWe develop a technique called Surface-seq to selectively sequence maxRNAs and validate two Surface-seq identified maxRNAs by RNA fluorescence in situ hybridization. To test for cell-type specificity of maxRNA, we use antisense oligos to hybridize to single-stranded transcripts exposed on the surface of human peripheral blood mononuclear cells (PBMCs). Combining this strategy with imaging flow cytometry, single-cell RNA sequencing, and maxRNA sequencing, we identify monocytes as the major type of maxRNA+ PBMCs and prioritize 11 candidate maxRNAs for functional tests. Extracellular application of antisense oligos of FNDC3B and CTSS transcripts inhibits monocyte adhesion to vascular endothelial cells.ConclusionsCollectively, these data highlight maxRNAs as functional components of the cell surface, suggesting an expanded role for RNA in cell-cell and cell-environment interactions.
Highlights
The cell surface is the crucial interface between the interior and exterior of the cell.Bioactive molecules, including proteins, glycans, lipids, and their chemically modified variations, are essential for the cell surface functions, e.g., extracellular signal sensing, extracellular matrix anchoring, and antigen presentation
Surface-seq: characterization of membrane-associated extracellular RNAs (maxRNAs) by sequencing To inspect the possible cell surface-presentation of nuclear-encoded RNA, we developed a technology called Surface-seq
Surface-seq is based on a nanotechnology that extracts the plasma membrane from cells and tightly assembles the membrane around polymeric cores to form membrane-coated nanoparticles (MCNP) [10,11,12]
Summary
The cell surface is the crucial interface between the interior and exterior of the cell. Bioactive molecules, including proteins, glycans, lipids, and their chemically modified variations, are essential for the cell surface functions, e.g., extracellular signal sensing, extracellular matrix anchoring, and antigen presentation. The contribution of nucleic acids RNAs to the cell surface functions is largely unknown. The nuclear genome encoded RNAs (ngRNA) are not expected to be present on the surface of eukaryotic cells with intact cell membranes [1]. Glycans, and lipids, much less is known about RNAs on the cell surface. We develop a series of technologies to test for any nuclearencoded RNAs that are stably attached to the cell surface and exposed to the extracellular space, hereafter called membrane-associated extracellular RNAs (maxRNAs)
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