Cell fates are mainly determined by transcription factor, transcription factors are regulated by post-transcription regulation20. microRNA (miRNA) is a class of small non-coding RNA of about 22 nt, which is widely involved in the post-transcriptional regulation of genes21. So far, it has been reported that miRNA affected myeloid cell differentiation includes miR-23322, miR-146a23, miR-2124, miR-19625 and miR-142-3p26. It is well known miR-144 and miR-451 are essential for erythrocyte proliferation and differentiation27,28. More and more evidence show that miRNA is involved in the hematopoiesis, but so far, it remains to be discovered that miRNA regulates both erythropoiesis and myeloid cell differentiation. Here, we discovered that a novel miRNA-7145-cuedc2 axis controls hematopoiesis through JAK1/STAT3 signaling pathway (Supplemental Figure. 6).
Hematopoiesis in zebrafish embryos mainly occurs in two waves, which is primitive wave and definitive wave1,29. The primitive wave mainly produces erythrocytes and limited myeloid cells10. Macrophages and granulocytes were specified from common myeloid progenitor cells in the primitive wave. Around 11 hours post fertilization, the myeloid progenitor cells were derived from the anterior lateral mesoderm, while erythrocytes were main derived from the posterior lateral mesoderm30. Around 30 hours post fertilization, the transient definitive wave arises in the posterior blood island (PBI), which can produce erythromyeloid progenitors (EMPs) with the ability to differentiate into erythrocytes and myeloid cells. Subsequently, the embryo enters the definitive wave, and the ventral wall of the dorsal aorta produces definitive HSCs, which then migrate to the posterior region of the tail, called the caudal hematopoietic tissue30. The expression pattern of miRNA-144 and miRNA-451 are similar to gata1a in primitive wave, which are expressed in ICM at 24 hpf28. However, miRNA-7145 is expressed in all tissues at 24 hpf, but not in yolk sac. It is possible that miRNA-7145 may have other functions in addition to hematopoietic functions during early embryonic development.
Interestingly, in definitive wave, expression pattern of miRNA-7145 is similar to gata1a at 48 hpf. This suggests that miRNA-7145 may be a key determinant of HSCs lineage specification.
Knock-down of miRNA-451 could increases the number of immature erythroid in zebrafish28. However, Knock-down of miR-7145 enhanced myeloid progenitor cell differentiation while against to erythropoiesis commitment. This result is in contrast to the over-expression of miRNA-7145. This result suggests that the phenotype caused by miRNA-7145 knock-down or over-expression is not due to off-target. To test this, we also did co-injection of miRNA mimics and MO, and the blood phenotype disappeared as expected.
A target gene of miRNA-451, gata2, was identified by fluorescence reporting experiments28. Similarly, we also identified cuedc2, a target gene of miRNA-7145, through bioinformatics analysis and fluorescence reporting experiments. Over-expression of the cuedc2 can partially rescue the phenotype caused by over-expression of the miRNA-7145. Cuedc2, a significant inflammation-related factor31, was highly expressed in peritoneal macrophages, and the expression level was dramatically increased when monocytes differentiate to macrophages32. However, it is unclear whether cuedc2 is involved in hematopoiesis. Here we showed that miRNA-7145 controls hematopoiesis through regulate cuedc2 expression. Moreover we found that cuedc2 enhanced myeloid progenitor cell differentiation while against to erythropoiesis commitment. The role of the EPO-JAK-STAT pathway in erythropoiesis is highly conserved in vertebrates. In zebrafish, EPO is required for erythropoiesis in both primitive and definitive wave33. Once EPOR binds to EPO, it recruits JAK, a tyrosine kinase that phosphorylates EPOR. In zebrafish, over-expression of JAK/STAT3 signaling pathway promotes erythropoiesis15–18. Here, we found that zebrafish cuedc2 can inhibits human JAK1/STAT3 signaling pathway.
In conclusion, we find that miRNA-7145-cuedc2 axis controls hematopoiesis through JAK1/STAT3 signaling pathway. This study not only provides new insight for erythropoiesis and myeloid cell differentiation, but also provides a possible evolutionary conserved mechanism for the regulation of HSCs differentiation.