The Genomic Analysis of Erythrocyte microRNA Expression in Sickle Cell Diseases

Shao-Yin Chen,Jen-Tsan Chi,Yulei Wang,Marilyn J Telen,Baohong Zhang

doi:10.1371/journal.pone.0002360

Abstract

BackgroundSince mature erythrocytes are terminally differentiated cells without nuclei and organelles, it is commonly thought that they do not contain nucleic acids. In this study, we have re-examined this issue by analyzing the transcriptome of a purified population of human mature erythrocytes from individuals with normal hemoglobin (HbAA) and homozygous sickle cell disease (HbSS).Methods and FindingsUsing a combination of microarray analysis, real-time RT-PCR and Northern blots, we found that mature erythrocytes, while lacking ribosomal and large-sized RNAs, contain abundant and diverse microRNAs. MicroRNA expression of erythrocytes was different from that of reticulocytes and leukocytes, and contributed the majority of the microRNA expression in whole blood. When we used microRNA microarrays to analyze erythrocytes from HbAA and HbSS individuals, we noted a dramatic difference in their microRNA expression pattern. We found that miR-320 played an important role for the down-regulation of its target gene, CD71 during reticulocyte terminal differentiation. Further investigation revealed that poor expression of miR-320 in HbSS cells was associated with their defective downregulation CD71 during terminal differentiation.ConclusionsIn summary, we have discovered significant microRNA expression in human mature erythrocytes, which is dramatically altered in HbSS erythrocytes and their defect in terminal differentiation. Thus, the global analysis of microRNA expression in circulating erythrocytes can provide mechanistic insights into the disease phenotypes of erythrocyte diseases.

Highlights

Erythrocytes constitute more than 90% of the cell population in the peripheral blood and are responsible for efficient gas exchange in the human body
In summary, we have discovered significant microRNA expression in human mature erythrocytes, which is dramatically altered in HbSS erythrocytes and their defect in terminal differentiation
Purification of Human Mature Erythrocytes To test for the possibility that mature erythrocytes contain previously undetected RNAs, we developed a protocol to obtain a pure population of mature erythrocytes by removing other blood cells through a series of purification procedures (Fig 1A)

Summary

Introduction

Erythrocytes constitute more than 90% of the cell population in the peripheral blood and are responsible for efficient gas exchange in the human body. Since hemoglobin makes up the majority of erythrocyte cellular proteins, these cells are frequently thought to serve merely as inert and passive containers of hemoglobins. Their more dynamic nature was suggested by the recent finding that erythrocytes can mount cellular signaling and trigger functional responses during physiological and pathological stresses [3,4]. This suggests that erythrocytes have a more sophisticated intracellular environment than previously appreciated. Since mature erythrocytes are terminally differentiated cells without nuclei and organelles, it is commonly thought that they do not contain nucleic acids. We have re-examined this issue by analyzing the transcriptome of a purified population of human mature erythrocytes from individuals with normal hemoglobin (HbAA) and homozygous sickle cell disease (HbSS)

Methods

Results

Conclusion