lncRNA DRAIR is downregulated in diabetic monocytes and modulates the inflammatory phenotype via epigenetic mechanisms
Marpadga A. Reddy, Vishnu Amaram, Sadhan Das, Vinay Singh Tanwar, Rituparna Ganguly, Mei Wang, Linda Lanting, Lingxiao Zhang, Maryam Abdollahi, Zhuo Chen, Xiwei Wu, Sridevi Devaraj, Rama Natarajan
Marpadga A. Reddy, Vishnu Amaram, Sadhan Das, Vinay Singh Tanwar, Rituparna Ganguly, Mei Wang, Linda Lanting, Lingxiao Zhang, Maryam Abdollahi, Zhuo Chen, Xiwei Wu, Sridevi Devaraj, Rama Natarajan
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Research Article Inflammation Metabolism

lncRNA DRAIR is downregulated in diabetic monocytes and modulates the inflammatory phenotype via epigenetic mechanisms

Abstract

Long noncoding RNAs (lncRNAs) are increasingly implicated in the pathology of diabetic complications. Here, we examined the role of lncRNAs in monocyte dysfunction and inflammation associated with human type 2 diabetes mellitus (T2D). RNA sequencing analysis of CD14+ monocytes from patients with T2D versus healthy controls revealed downregulation of antiinflammatory and antiproliferative genes, along with several lncRNAs, including a potentially novel divergent lncRNA diabetes regulated antiinflammatory RNA (DRAIR) and its nearby gene CPEB2. High glucose and palmitic acid downregulated DRAIR in cultured CD14+ monocytes, whereas antiinflammatory cytokines and monocyte-to-macrophage differentiation upregulated DRAIR via KLF4 transcription factor. DRAIR overexpression increased antiinflammatory and macrophage differentiation genes but inhibited proinflammatory genes. Conversely, DRAIR knockdown attenuated antiinflammatory genes, promoted inflammatory responses, and inhibited phagocytosis. DRAIR regulated target gene expression through interaction with chromatin, as well as inhibition of the repressive epigenetic mark H3K9me2 and its corresponding methyltransferase G9a. Mouse orthologous Drair and Cpeb2 were also downregulated in peritoneal macrophages from T2D db/db mice, and Drair knockdown in nondiabetic mice enhanced proinflammatory genes in macrophages. Thus, DRAIR modulates the inflammatory phenotype of monocytes/macrophages via epigenetic mechanisms, and its downregulation in T2D may promote chronic inflammation. Augmentation of endogenous lncRNAs like DRAIR could serve as novel antiinflammatory therapies for diabetic complications.

Authors

Marpadga A. Reddy, Vishnu Amaram, Sadhan Das, Vinay Singh Tanwar, Rituparna Ganguly, Mei Wang, Linda Lanting, Lingxiao Zhang, Maryam Abdollahi, Zhuo Chen, Xiwei Wu, Sridevi Devaraj, Rama Natarajan

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Figure 8

DRAIR interacts with G9a and alters levels of repressive histone modifications.

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DRAIR interacts with G9a and alters levels of repressive histone modifi...
(A) qPCR of RNA recovered from ChIRP with DRAIR probes or negative control luciferase (LUC) probes. Cell lysates from THP1 monocytes overexpressing DRAIR were subjected to ChIRP assays with biotinylated DRAIR probes and negative control biotinylated LUC probes. ChIRP complexes were captured on streptavidin beads, and RNA from an aliquot of beads was analyzed by qPCR with DRAIR primers. (B) The nucleic acid–protein complexes from ChIRP were fractionated on SDS-PAGE and subjected to mass spectrometry to identify proteins interacting with DRAIR probes. STRING analysis of indicated DRAIR-interacting proteins identified by ChIRP mass spectrometry (Supplemental Figure 7). Colors represent GO biological functions shown in Supplemental Figure 7. (C) Immunoblotting of proteins from RNA pulldown assays using DRAIR and DRAIR antisense (DRAIR-AS) probes with G9a antibody. (D) qPCR analysis of RNA recovered after RNA IP with indicated antibodies. (E–M) ChIP-qPCR analysis of DNA recovered from ChIP assays using lysates from THP1 cells overexpressing DRAIR and empty vector pcDNA3.1 (EV) with antibodies against H3K9me2 (E–H), G9a (I–L), and H3K27me3 (M) using indicated gene promoter primers. (N and O) Immunoblotting of THP1 nuclear extracts with indicated antibodies (N) and quantification of G9a in nuclear extracts (O). THP1 cells were treated with NG (5.5 mM glucose) and HG (25 mM glucose) for 72 hours. Palmitic acid (PA, 200 μM) was also added in the final 24 hours to NG- (PA) and HG-treated (HP) cells. *P < 0.05; **P < 0.01; ***P < 0.001 (n = 3) as determined by unpaired t test (E–L) and 2-way ANOVA with Sidak’s (M), and 1-way ANOVA with Dunnett’s (O) multiple-comparison testing.