Identification of abundant and evolutionarily conserved opioid receptor circular RNAs in the nervous system modulated by morphine

TY - JOUR

T1 - Identification of abundant and evolutionarily conserved opioid receptor circular RNAs in the nervous system modulated by morphine

AU - Irie, Takeshi

AU - Shum, Rebecca

AU - Deni, Ioanna

AU - Hunkele, Amanda

AU - Le Rouzic, Valerie

AU - Xu, Jin

AU - Wilson, Roger

AU - Fischer, Gregory W.

AU - Pasternak, Gavril W.

AU - Pan, Ying Xian

N1 - Funding Information: This work was supported, in part, by funds from the Department of Anesthesiology (to T.I.), the National Institutes of Health National Institute on Drug Abuse [Grants DA06241, DA07242, DA042888, DA040858 and DA046714], the Mayday Foundation, the Peter F. McManus Charitable Trust, and a core grant from the National Institutes of Health National Cancer Institute [Grant CA008748] to the Memorial Sloan-Kettering Cancer Center. Publisher Copyright: © 2019 by The American Society for Pharmacology and Experimental Therapeutics.

PY - 2019

Y1 - 2019

N2 - Circular RNAs (circRNAs) are a distinct category of single-stranded, covalently closed RNAs formed by backsplicing. The functions of circRNAs are incompletely known and are under active investigation. Here, we report that in addition to traditional linear mRNAs (linRNA), mouse, rat, and human opioid receptor genes generate exonic circRNA isoforms. Using standard molecular biologic methods, Oprm1 circRNAs (circOprm1) were detected in RNAs of rodent and human brains and spinal cords, as well as human neuroblastoma cells, suggesting evolutionary conservation. Sequencing confirmed backsplicing using canonical splice sites. Oprm1 circRNAs were sense-stranded circRNAs resistant to RNase R digestion. The relative abundance of Oprm1 circRNA to linRNA determined by quantitative reverse transcription polymerase chain reaction varied among mouse brain regions, with circRNA isoforms predominating in rostral structures and less abundant in brain stem. Chronic morphine exposure in mice increased brain circOprm1.e2.3 and circOprm1.e2.e3.e4(302) levels by 1.5- to 1.6-fold relative to linRNA. Sequence analysis predicted numerous microRNA binding sites within Oprm1 circRNA sequences, suggesting a potential role in microRNA sequestration through sponging. In addition, we observed that other opioid receptor genes including d, k, and nociceptin receptor genes produced similar circRNAs. In conclusion, all members of the opioid receptor gene family express circRNAs, with Oprm1 circRNA levels exceeding those of linear forms in some regions.

AB - Circular RNAs (circRNAs) are a distinct category of single-stranded, covalently closed RNAs formed by backsplicing. The functions of circRNAs are incompletely known and are under active investigation. Here, we report that in addition to traditional linear mRNAs (linRNA), mouse, rat, and human opioid receptor genes generate exonic circRNA isoforms. Using standard molecular biologic methods, Oprm1 circRNAs (circOprm1) were detected in RNAs of rodent and human brains and spinal cords, as well as human neuroblastoma cells, suggesting evolutionary conservation. Sequencing confirmed backsplicing using canonical splice sites. Oprm1 circRNAs were sense-stranded circRNAs resistant to RNase R digestion. The relative abundance of Oprm1 circRNA to linRNA determined by quantitative reverse transcription polymerase chain reaction varied among mouse brain regions, with circRNA isoforms predominating in rostral structures and less abundant in brain stem. Chronic morphine exposure in mice increased brain circOprm1.e2.3 and circOprm1.e2.e3.e4(302) levels by 1.5- to 1.6-fold relative to linRNA. Sequence analysis predicted numerous microRNA binding sites within Oprm1 circRNA sequences, suggesting a potential role in microRNA sequestration through sponging. In addition, we observed that other opioid receptor genes including d, k, and nociceptin receptor genes produced similar circRNAs. In conclusion, all members of the opioid receptor gene family express circRNAs, with Oprm1 circRNA levels exceeding those of linear forms in some regions.

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U2 - https://doi.org/10.1124/mol.118.113977

DO - https://doi.org/10.1124/mol.118.113977

M3 - Article

C2 - 31243060

SN - 0026-895X

VL - 96

SP - 247

EP - 258

JO - Molecular Pharmacology

JF - Molecular Pharmacology

IS - 2

ER -