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A novel antisense long noncoding RNA within the IGF1R gene locus is imprinted in hematopoietic malignancies.

Sun J, Li W, Sun Y, Yu D, Wen X, Wang H, Cui J, Wang G, Hoffman AR, Hu JF - Nucleic Acids Res. (2014)

Bottom Line: Using both reverse transcription-associated trap and chromatin conformation capture assays, we demonstrate that this lncRNA interacts with chromatin DNA and is involved in the formation of an intrachromosomal enhancer/promoter loop.In addition, IRAIN was downregulated both in leukemia cell lines and in blood obtained from high-risk AML patients.These data identify IRAIN as a new imprinted lncRNA that is involved in long-range DNA interactions.

View Article: PubMed Central - PubMed

Affiliation: Stem Cell and Cancer Center, First Affiliated Hospital, Jilin University, Changchun, Jilin 130061, PR China Stanford University Medical School, VA Palo Alto Health Care System, Palo Alto, CA 94304, USA.

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Characterization of IRAIN as an antisense lncNRA. (A) The diagram of the IRAIN/IGF1R locus. pIRAIN: IRAIN lncRNA promoter; pIR: IGF1R coding RNA promoter. Vertical arrows: the location of lncRNA PCR primers. (B and C) Mapping of the IRAIN lncRNA in K562 leukemia cells. gDNA: genomic DNA used as the control to test the efficiency of the PCR primers. M: 100 bp marker. (D) Northern blot of the IRAIN lncRNA in breast cancer tissues. Total RNA from three breast cancer tumors was separated on a 1.5% (w/v) denaturing agarose gel and was hybridized with the 32P-dCTP labeled IRAIN cDNA clone probe. GAPDH was used as the control. E: IRAIN lncRNA is an antisense lncRNA. Horizontal arrows: SSRT-PCR primers used to map the orientation of IRAIN lncRNA. The strand-specific cDNAs were synthesized using either the 5′- or the 3′-oligonucleotides at sites C, D and F. A pair of PCR primers located between two cDNA oligonucleotides was then used to determine the transcription orientation of the IRAIN lncRNA. M: 100 bp marker; input: total RNA collected before SSRT-PCR; RT: reverse transcriptase.
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Figure 1: Characterization of IRAIN as an antisense lncNRA. (A) The diagram of the IRAIN/IGF1R locus. pIRAIN: IRAIN lncRNA promoter; pIR: IGF1R coding RNA promoter. Vertical arrows: the location of lncRNA PCR primers. (B and C) Mapping of the IRAIN lncRNA in K562 leukemia cells. gDNA: genomic DNA used as the control to test the efficiency of the PCR primers. M: 100 bp marker. (D) Northern blot of the IRAIN lncRNA in breast cancer tissues. Total RNA from three breast cancer tumors was separated on a 1.5% (w/v) denaturing agarose gel and was hybridized with the 32P-dCTP labeled IRAIN cDNA clone probe. GAPDH was used as the control. E: IRAIN lncRNA is an antisense lncRNA. Horizontal arrows: SSRT-PCR primers used to map the orientation of IRAIN lncRNA. The strand-specific cDNAs were synthesized using either the 5′- or the 3′-oligonucleotides at sites C, D and F. A pair of PCR primers located between two cDNA oligonucleotides was then used to determine the transcription orientation of the IRAIN lncRNA. M: 100 bp marker; input: total RNA collected before SSRT-PCR; RT: reverse transcriptase.

Mentions: IGFIR is frequently overexpressed in both solid tumors and hematopoietic malignancies, participating in the regulation of cancer cell proliferation, survival, metabolism and metastasis. To address the mechanisms underlying the dysregulation of IGF1R in leukemia cells, we used a novel R3C method recently developed in our lab (Supplementary Figure S1) (22), and detected the presence of RNA molecules within the IGF1R promoter and intron 1, where the IGF1R mRNA is not transcribed. To characterize this novel RNA molecule, we designed a series of PCR primers covering the IGF1R promoter region and mapped its transcription in the IGF1R locus (Figure 1A). Using PCR, we detected the transcription of this noncoding RNA from the IGF1R promoter and intron 1 (Figure 1B, from C to H sites, lanes 3–8). No PCR products were detected further upstream (sites A, B; lanes 1–2) or the region near exon 2 (sites I, J; lanes 9–10).


A novel antisense long noncoding RNA within the IGF1R gene locus is imprinted in hematopoietic malignancies.

Sun J, Li W, Sun Y, Yu D, Wen X, Wang H, Cui J, Wang G, Hoffman AR, Hu JF - Nucleic Acids Res. (2014)

Characterization of IRAIN as an antisense lncNRA. (A) The diagram of the IRAIN/IGF1R locus. pIRAIN: IRAIN lncRNA promoter; pIR: IGF1R coding RNA promoter. Vertical arrows: the location of lncRNA PCR primers. (B and C) Mapping of the IRAIN lncRNA in K562 leukemia cells. gDNA: genomic DNA used as the control to test the efficiency of the PCR primers. M: 100 bp marker. (D) Northern blot of the IRAIN lncRNA in breast cancer tissues. Total RNA from three breast cancer tumors was separated on a 1.5% (w/v) denaturing agarose gel and was hybridized with the 32P-dCTP labeled IRAIN cDNA clone probe. GAPDH was used as the control. E: IRAIN lncRNA is an antisense lncRNA. Horizontal arrows: SSRT-PCR primers used to map the orientation of IRAIN lncRNA. The strand-specific cDNAs were synthesized using either the 5′- or the 3′-oligonucleotides at sites C, D and F. A pair of PCR primers located between two cDNA oligonucleotides was then used to determine the transcription orientation of the IRAIN lncRNA. M: 100 bp marker; input: total RNA collected before SSRT-PCR; RT: reverse transcriptase.
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Figure 1: Characterization of IRAIN as an antisense lncNRA. (A) The diagram of the IRAIN/IGF1R locus. pIRAIN: IRAIN lncRNA promoter; pIR: IGF1R coding RNA promoter. Vertical arrows: the location of lncRNA PCR primers. (B and C) Mapping of the IRAIN lncRNA in K562 leukemia cells. gDNA: genomic DNA used as the control to test the efficiency of the PCR primers. M: 100 bp marker. (D) Northern blot of the IRAIN lncRNA in breast cancer tissues. Total RNA from three breast cancer tumors was separated on a 1.5% (w/v) denaturing agarose gel and was hybridized with the 32P-dCTP labeled IRAIN cDNA clone probe. GAPDH was used as the control. E: IRAIN lncRNA is an antisense lncRNA. Horizontal arrows: SSRT-PCR primers used to map the orientation of IRAIN lncRNA. The strand-specific cDNAs were synthesized using either the 5′- or the 3′-oligonucleotides at sites C, D and F. A pair of PCR primers located between two cDNA oligonucleotides was then used to determine the transcription orientation of the IRAIN lncRNA. M: 100 bp marker; input: total RNA collected before SSRT-PCR; RT: reverse transcriptase.
Mentions: IGFIR is frequently overexpressed in both solid tumors and hematopoietic malignancies, participating in the regulation of cancer cell proliferation, survival, metabolism and metastasis. To address the mechanisms underlying the dysregulation of IGF1R in leukemia cells, we used a novel R3C method recently developed in our lab (Supplementary Figure S1) (22), and detected the presence of RNA molecules within the IGF1R promoter and intron 1, where the IGF1R mRNA is not transcribed. To characterize this novel RNA molecule, we designed a series of PCR primers covering the IGF1R promoter region and mapped its transcription in the IGF1R locus (Figure 1A). Using PCR, we detected the transcription of this noncoding RNA from the IGF1R promoter and intron 1 (Figure 1B, from C to H sites, lanes 3–8). No PCR products were detected further upstream (sites A, B; lanes 1–2) or the region near exon 2 (sites I, J; lanes 9–10).

Bottom Line: Using both reverse transcription-associated trap and chromatin conformation capture assays, we demonstrate that this lncRNA interacts with chromatin DNA and is involved in the formation of an intrachromosomal enhancer/promoter loop.In addition, IRAIN was downregulated both in leukemia cell lines and in blood obtained from high-risk AML patients.These data identify IRAIN as a new imprinted lncRNA that is involved in long-range DNA interactions.

View Article: PubMed Central - PubMed

Affiliation: Stem Cell and Cancer Center, First Affiliated Hospital, Jilin University, Changchun, Jilin 130061, PR China Stanford University Medical School, VA Palo Alto Health Care System, Palo Alto, CA 94304, USA.

Show MeSH
Related in: MedlinePlus