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750 Lncrna-Iκbα Promotes Erythroid Differentiation By Inhibiting NF-Κb Pathway As a Cerna Against Mir-24-3p in Polycythemia Vera

Program: Oral and Poster Abstracts
Session: 101. Red Cells and Erythropoiesis, Structure and Function, Metabolism, and Survival, Excluding Iron: Poster I
Hematology Disease Topics & Pathways:
Animal models, red blood cells, Biological Processes, Cell Lineage, erythropoiesis, Study Population, Clinically relevant
Saturday, December 5, 2020, 7:00 AM-3:30 PM

Zhigang Li1*, Xuemei Fu1*, Zhixiang Zuo, PhD2*, Fangfang Sun1*, Xiaojun Zhang1*, Jieping Chen3* and Yu Hou, PhD4

1Southwest Hospital, Third Military Medical University (Army Medical University), ChongQing, China
2State Key Laboratory of Oncology in South China, Collaborative Innovation Center for Cancer Medicine, Sun Yat-sen University Cancer Center, Guangzhou, China
3Southwest Hospital, Third Military Medical University (Army Medical University), Chongqing, China
4Department of Hematology, Southwest Hospital, Third Military Medical University (Army Medical University), CHONGQING, China

Myeloproliferative neoplasms (MPNs) are clonal hematopoietic stem/progenitor cell diseases characterized by the overproduction of one or more mature myeloid blood cell lineages, mainly including primary thrombocythemia (ET) and polycythemia vera (PV). The acquired kinase mutation Jak2V617F plays a central role in these disorders, but the mechanisms responsible for same genetic mutation leading to different clinical features are not fully understood. LncRNAs are critical regulators of diverse physiological and pathological cellular processes, including normal and malignant hematopoiesis. Although thousands of erythroid stage-specific lncRNAs have been identified, the biological functions of the vast majority of lncRNAs in PV remains to be elucidated.

In an attempt to identify lncRNAs associated with the occurrence and development of PV progression, we conducted three rounds of screening sequentially. Firstly, RNA-Seq datasets were generated form ET, PV patients and normal peoples. In total, we identified 32259 lncRNAs, including 31841 annotated and 418 novel lncRNAs. Secondly, 446 differentially expressed lncRNAs (DELs) were identified between normal peoples and patients (ET and PV). Meanwhile, 53 DELs were identified between ET and PV. Lastly, 26 DELs involved in PV were identified by intersecting these two DELs datasets with each other. KEGG pathway analysis showed that these lncRNAs were mainly concentrated in the NF-κB signal pathway. Subsequently, expression of 9 lncRNAs were confirmed by real-time PCR. From these 9 lncRNAs, 2 relative high-expression and most differentially expressed lncRNAs (foldchange > 4) were subjected to loss-of-function analysis in JAK2V617F positive HEL cells.

We focused on a novel lncRNA and named it lncRNA-IκBα, which inhibited hemin-induced erythroid differentiation in HEL cells. The expression of γ-globin and CD235a were significantly inhibited (~ reduced to 50%) in lncRNA-IκBα knockdown HEL cells compared to control cells. Likewise, flow cytometry revealed that the frequency of CD235+ cells and CD71+ cells was also lower in lncRNA-IκBα knockdown HEL cells than that in control cells (5% vs 29% , 1.64% vs 6.55%). Additionally, CCK-8 assays showed that the proliferation of lncRNA-IκBα knockdown cell was significantly slower than control cells . We assessed in vivo incorporation of Brdu to investigate the cell-cycle kinetics and found that 36% of lncRNA-IκBα knockdown cells and 44% of control cells were Brdu+, which indicated that knockdown of lncRNA-IκBα inhibit the proliferation. Analysis of lncRNA-miRNA-mRNA co-regulatory network showed that lncRNA-IκBα may target IκBα through competing binding with mir-24-3p. LncRNA-IκBα and IκBα have consistent expression patterns in MPN patients. In addition, lncRNA-IκBα has predicted binding site for mir-24-3p. Over-expression of lncRNA-IκBα in HEL cells can significantly up-regulate the expression of IκBα (~50%). Taken together, we suggested that lncRNA-IκBα regulates erythroid differentiation by regulating NF-κB signaling pathway through competitive binding endogenous miR-24-3p with IκBα. Furthermore, lncRNA-IκBα may affects the occurrence and development of PV by regulating erythroid differentiation.

Disclosures: No relevant conflicts of interest to declare.

*signifies non-member of ASH