microRNA Changes in Diabetic Cardiac Mitochondria: What are they doing there?
Authors: Pinti Mark V. , Hathaway Quincy A. , Kunovac Amina , Durr Andrya J. , Cook Chris C , Roberts Harold G. , Salman Muhammad , Hollander John M. 
Published: June 2021

The mitochondrion has been found to harbor non‐coding RNA (ncRNA) that appear to be dynamically regulated in health and disease. Due to the tremendous energy demand of the heart and the well‐characterized cardiac dysfunction observed in type 2 diabetes mellitus (T2DM), we investigated whether mitochondrial microRNA (miR) were differentially expressed in human and mouse diabetic heart tissue. In isolated mitochondria from human right atrial appendages (n = 7 non‐diabetic and n = 7 T2DM), small ncRNA sequencing was performed. Libraries were constructed using the NEXTflex Small RNA‐Seq Kit v3 and sequenced on the HiSeq 2500 in Rapid Run mode using 2 lanes of single‐end (SE) 40 base pair reads. After identifying miRNA expression changes in human cardiac mitochondria between non‐diabetic and T2DM patients, we sought to determine the role of these nuclear‐encoded miRNA in the mitochondria. To accomplish this, we were interested in the binding of mitochondria‐localized RNA‐induced silencing complex (RISC) components to miRNA in cardiac mitochondria of a diabetic mouse model. In FVB (n = 5 pooled samples) and FVB‐db/db (n=5 pooled samples) mouse models, cross‐linked immunoprecipitation (CLIP) sequencing was performed for both argonaute 2 (Ago2) and fragile X mental retardation syndrome‐related protein 1 (Fxr1). Libraries were constructed using the TruSeq Small RNA Library Prep Kit and sequenced on the HiSeq 2500 in Rapid Run mode using 2 lanes of single‐end (SE) 40 base pair reads. Files were aligned to the reference genomes GRCh38 (human) and GRCm38 (mouse) release 94 through Bowtie running “best” and “strata.” Overall, we identified differential expression of 236 miR species in non‐diabetic and T2DM human cardiac mitochondria. Those increased greater than 3‐fold in T2DM cardiac mitochondria included miR‐3687‐1, miR‐3687‐2; and some of those decreased greater than 3‐fold included miR‐23A, miR‐23B, miR‐30A, miR‐30B, miR‐30C1, miR‐30C2, miR‐30D, miR‐30E, miR‐101‐1, miR‐191, miR‐340, miR‐361, and miR‐374A. Interestingly, we found overlap of some miRNA species found in human cardiac mitochondria with those found bound to Ago2 and FXR1 in diabetic mouse cardiac mitochondria. These included miRNA‐let7i (p=0.049), miRNA‐26a‐1 (p=0.00045), miRNA‐26a‐2 (p=0.00045), miRNA‐27a (p=0.0023), miRNA‐27b (p=0.0014), miRNA‐30a (p=3.13E‐05), and miRNA‐181a‐1 (p=0.0046). In conclusion, the association of RISC components with miRNA species found to have significant expression changes in the T2DM human heart suggests that not only is the FVB‐db/db a clinically relevant model of T2DM, but also that miRNA play a role in translational repression in the T2DM human heart.Support or Funding InformationThis work was supported by: R01 HL‐128485 (JMH), AHA‐17PRE33660333 (QAH), WVU Genomics Core Facility, and the Community Foundation for the Ohio Valley Whipkey Trust.This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.