Abstract
Circulating human red blood cells (RBCs) consist of mature erythrocytes and immature reticulocytes. Being anucleated, RBCs lack typical transcriptomes, but are known to contain small amounts of diverse long transcripts and microRNAs. However, the exact role and importance of these RNAs is lacking. Results To study this further, we explored the transcriptomes of RBCs and extracellular vesicles (EVs) of RBCs using next-generation sequencing. Furthermore, to understand the dynamics of the RBC transcriptome, we performed single-cell RNA sequencing on RBCs. An analysis of the single-cell transcriptomes revealed that approximately 10% of the cells contained detectable levels of mRNA and fell into three subpopulations based on their transcriptomes. Decrease in the mRNA quantity was observed across the populations. Qualitative changes included the differences in the globin transcripts and changes in the expression of ribosomal genes. A specific short splice form of a long non-coding RNA, Metastasis Associated Lung Adenocarcinoma Transcript 1 (MALAT1), was the most enriched marker in one subpopulation of RBCs, co-expressing with ribosomal structural transcripts. MALAT1 expression was confirmed by qPCR in CD71-enriched reticulocytes, which were also characterized with imaging flow cytometry at single cell level. Conclusions Analysis of the RBC transcriptome shows enrichment of pathways and functional categories required for the maturation of reticulocytes and erythrocyte functions. The RBC transcriptome was detected in their EVs, proposing vesiculation as a mechanism to remove the cellular contents from RBCs throughout their life cycle. Our experiments on single cell level revealed that lncRNA MALAT1 is the marker for one of the three RBC populations co-expressing with a group of ribosomal protein transcripts.
Dextran adsorption onto red blood cells revisited: single cell quantification by laser tweezers combined with microfluidics