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RNA Biol. 2017 Jan 2;14(1):58-72. doi: 10.1080/15476286.2016.1249092. Epub 2016 Oct 28.

Two distinct extracellular RNA signatures released by a single cell type identified by microarray and next-generation sequencing.

Author information

1
a Krefting Research Center, Department of Internal Medicine and Clinical Nutrition , University of Gothenburg , Gothenburg , Sweden.
2
b Neurogenomics, TGen , Phoenix , AZ , USA.
3
c Department of Life Sciences , Pohang University of Science and Technology , Pohang , Gyeongbuk , Republic of Korea.
4
d Department of Biopathology, and Medical Biotechnologies , Section of Biology and Genetics, Università di Palermo , Palermo , Italy.

Abstract

Cells secrete extracellular RNA (exRNA) to their surrounding environment and exRNA has been found in many body fluids such as blood, breast milk and cerebrospinal fluid. However, there are conflicting results regarding the nature of exRNA. Here, we have separated 2 distinct exRNA profiles released by mast cells, here termed high-density (HD) and low-density (LD) exRNA. The exRNA in both fractions was characterized by microarray and next-generation sequencing. Both exRNA fractions contained mRNA and miRNA, and the mRNAs in the LD exRNA correlated closely with the cellular mRNA, whereas the HD mRNA did not. Furthermore, the HD exRNA was enriched in lincRNA, antisense RNA, vault RNA, snoRNA, and snRNA with little or no evidence of full-length 18S and 28S rRNA. The LD exRNA was enriched in mitochondrial rRNA, mitochondrial tRNA, tRNA, piRNA, Y RNA, and full-length 18S and 28S rRNA. The proteomes of the HD and LD exRNA-containing fractions were determined with LC-MS/MS and analyzed with Gene Ontology term finder, which showed that both proteomes were associated with the term extracellular vesicles and electron microscopy suggests that at least a part of the exRNA is associated with exosome-like extracellular vesicles. Additionally, the proteins in the HD fractions tended to be associated with the nucleus and ribosomes, whereas the LD fraction proteome tended to be associated with the mitochondrion. We show that the 2 exRNA signatures released by a single cell type can be separated by floatation on a density gradient. These results show that cells can release multiple types of exRNA with substantial differences in RNA species content. This is important for any future studies determining the nature and function of exRNA released from different cells under different conditions.

KEYWORDS:

Exosomes; RNA; extracellular RNA; extracellular vesicles; microarray; next-generation sequencing; proteomics; transcriptomics

PMID:
27791479
PMCID:
PMC5270547
DOI:
10.1080/15476286.2016.1249092
[Indexed for MEDLINE]
Free PMC Article

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