TY - JOUR T1 - Differential stability of miR-9-5p and miR-9-3p in the brain is determined by their unique <em>cis</em>- and <em>trans</em>-acting elements JF - eneuro JO - eNeuro DO - 10.1523/ENEURO.0094-20.2020 SP - ENEURO.0094-20.2020 AU - C. Kim AU - A. Asimes AU - M. Zhang AU - B Son AU - J. A. Kirk AU - T. R. Pak Y1 - 2020/05/06 UR - http://www.eneuro.org/content/early/2020/05/06/ENEURO.0094-20.2020.abstract N2 - microRNAs (miR) are fundamental regulators of protein coding genes. In the central nervous system, miR-9 is highly enriched and critical for neuronal development and function. Mature miRs are derived from a duplex precursor, and the -5p strand (“guide”) is preferentially incorporated into an RNA-induced silencing complex to exert its regulatory functions, while the complementary -3p strand (“passenger”) is thought to be rapidly degraded. By contrast, both strands of the miR-9 duplex have unique functions critical for neuronal physiology, yet their respective degradation rates and mechanisms governing degradation are not well understood. Therefore, we determined the degradation kinetics of miR-9-5p and miR-9-3p and investigated the cis and trans elements that affected their stability in the brain. Using a combination of homogeneous neuronal/astrocyte cell models and heterogeneous brain tissue lysate, we demonstrate the novel finding that miR-9-3p was more stable than the miR-9-5p guide strand in all models tested. Moreover, the degradation kinetics of both miR-9-5p and miR-9-3p were brain-region specific, suggesting that each brain region was differentially enriched for specific degradation factors. We also determined that the 3’ nucleotides harbor important cis elements required to not only maintain stability, but also to recruit potential protein degradation factors. We used mass spectrometry to assess the miR-9 interacting proteins and found that the -5p and -3p strands were associated with functionally distinct proteins. Overall, these studies revealed unique miR-9-5p and miR-9-3p degradation kinetics in the brain and propose critical nucleotide sequences and protein partners that could contribute to this differential stability.Significance Statement miR-9-5p and miR-9-3p are two single-stranded microRNAs (miR) derived from the same RNA duplex that are critical for normal neuronal function. Here, we report the differential degradation kinetics of these neuronally enriched miRs, as well as identify unique regulatory cis and trans elements that could contribute to the distinct miR-9-5p and miR-9-3p stability in neurons. These findings contribute to the current understanding of how neuronal miRs are degraded and could have functional implications for their respective mRNA targets. ER -