A Novel Suppressive Long Noncoding RNA within the IGF1R Gene Locus Is Imprinted in Acute Myelocytic Leukemia

Background：In acute myeloid leukemia (AML) cells, the insulin-like growth factor type I receptor (IGF1R) is one of the most abundantly phosphorylated receptor tyrosine kinases, promoting cell growth and therapeutic resistance through the PI3K/Akt signaling pathway. However, little is known regarding the molecular mechanisms underlying IGF1R gene dysregulation in AML. Long non-coding RNAs have been implicated in a variety of important biological processes by epigenetic mechanisms.

Materials and methods：AML cell lines used in this study, KG-1, KG-1a were purchased from ATCC. The full length of IRAIN lncRNA was characterized by Marathon cDNA Amplification Kit. For qPCR, cDNA samples were amplified using CFX96^TM real-time system (BIO-RAD) by SYBR PrimeScript™ RT-PCR Kit (TaKaRa). Both reverse transcription-associated trap (RAT) and chromatin conformation capture (3C) assays were used to examine the chromosome architecture of IGF1R and DNA-RNA interaction. The protocol of AMLpatients bone marrow and peripheral blood cell samples was approved by the Human Medical Ethical Review Committee from Jilin University First Hospital and informed consent was obtained from each AML patient and normal donor.

Results：

1. we discovered a novel intragenic 5366bp long noncoding RNA (lncRNA) within the IGF1R locus, named IRAIN, which is transcribed in an antisense direction from an intronic promoter.

2. we demonstrated IRAIN was expressed exclusively from the paternal allele, with the maternal counterpart being silenced in normal donor and AML patients.

3. we confirmed that IRAIN interacts with IGF1R chromatin DNA and is involved in the formation of an intrachromosomal enhancer/promoter loop. Knockdown of IRAIN lncRNA with shRNA abolishes this intrachromosomal interaction.

4. We demonstrated IRAIN was downregulated both in AML cell lines and high risk AML patients. Furthermore，the expression of IRAIN was dramatical higher in APL patients achieved molecular remission.

Conclusion：Our results identify IRAIN as a new imprinted suppressive lncRNA involved in long range IGF1R DNA interactions in AML. As a putative tumor suppressor and a novel treatment target， further studies are needed to delineate the specific role of this newly identified lncRNA in the uregulation of the IGF pathway in AML.