Department of Bioengineering, University of California San Diego, San Diego, CA, 92093, USA.
Department of NanoEngineering, University of California San Diego, San Diego, CA, 92093, USA.
Genome Biol. 2020 Sep 10;21(1):225. doi: 10.1186/s13059-020-02145-6.
Compared 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).
We 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.
Collectively, these data highlight maxRNAs as functional components of the cell surface, suggesting an expanded role for RNA in cell-cell and cell-environment interactions.
相较于蛋白质、聚糖和脂类，人们对细胞表面上的 RNA 知之甚少。我们开发了一系列技术来检测任何稳定附着在细胞表面并暴露于细胞外空间的核编码 RNA，以下称为膜结合细胞外 RNA（maxRNA）。
我们开发了一种称为 Surface-seq 的技术，用于选择性地对 maxRNA 进行测序，并通过 RNA 荧光原位杂交验证了两种由 Surface-seq 鉴定的 maxRNA。为了测试 maxRNA 的细胞类型特异性，我们使用反义寡核苷酸与人类外周血单核细胞（PBMC）表面暴露的单链转录本杂交。将这种策略与成像流式细胞术、单细胞 RNA 测序和 maxRNA 测序相结合，我们鉴定出单核细胞是 maxRNA+PBMC 的主要类型，并为功能测试确定了 11 种候选 maxRNA。FNDC3B 和 CTSS 转录本的反义寡核苷酸的细胞外应用抑制了单核细胞与血管内皮细胞的黏附。
总的来说，这些数据突出了 maxRNA 作为细胞表面功能成分的作用，表明 RNA 在细胞-细胞和细胞-环境相互作用中发挥了扩展作用。
