MicroRNA‐574 regulates FAM210A expression and influences pathological cardiac remodeling

Abstract Aberrant expression of mitochondrial proteins impairs cardiac function and causes heart disease. The mechanism of regulation of mitochondria encoded protein expression during cardiac disease, however, remains underexplored. Here, we show that multiple pathogenic cardiac stressors induce the expression of miR‐574 guide and passenger strands (miR‐574‐5p/3p) in both humans and mice. miR‐574 knockout mice exhibit severe cardiac disorder under different pathogenic cardiac stresses while miR‐574‐5p/3p mimics that are delivered systematically using nanoparticles reduce cardiac pathogenesis under disease insults. Transcriptomic analysis of miR‐574‐null hearts uncovers family with sequence similarity 210 member A (FAM210A) as a common target mRNA of miR‐574‐5p and miR‐574‐3p. The interactome capture analysis suggests that FAM210A interacts with mitochondrial translation elongation factor EF‐Tu. Manipulating miR‐574‐5p/3p or FAM210A expression changes the protein expression of mitochondrial‐encoded electron transport chain (ETC) genes but not nuclear‐encoded mitochondrial ETC genes in both human AC16 cardiomyocyte cells and miR‐574‐null murine hearts. Together, we discovered that miR‐574 regulates FAM210A expression and modulates mitochondrial‐encoded protein expression, which may influence cardiac remodeling in heart failure.


D E F
. Expression of miR-574-5p/3p in murine organs and cardiac cells.
A Northern blot detection of miR-574-5p/3p expression and RT-qPCR measurement of Fam210a mRNA expression in various murine organs (n = 3). B miR-574-5p and miR-574-3p expression in adult CM and CF cells isolated from murine hearts of WT mice at baseline. Postn and Myh6 are used as cardiac fibroblast and cardiomyocyte marker, respectively. N = 3 per group. C miR-574-5p and miR-574-3p expression in adult CM and CF cells isolated from murine hearts of WT mice undergoing TAC surgery after 3 days. N = 3 per group. D, E Expression of miR-574-5p and miR-574-3p in the heart from mice undergoing TAC surgery at different time points. N = 3 per group. F ISO induces miR-574-5p and miR-574-3p expression in isolated murine ACMs. N = 3 per group.
Data information: Data were presented as mean AE SEM. P values were calculated by one-way ANOVA with Tukey's multiple comparisons test (C-E) and two-way ANOVA with Tukey's multiple comparisons test (F). Source data are available online for this figure. A B C E D Figure EV2. Phenotypic characterization of isolated CMs and hearts of WT and miR-574 À/À mice.
A Phalloidin staining of primary ACMs from WT and miR-574 À/À mice in response to isoproterenol (ISO) treatment (10 lM for 24 h). Data are obtained from 3 individual experiments (n> 100 CM cells/group). Scale bar: 50 lm. B Trypan blue staining of primary ACMs from WT and miR-574 À/À mice under ISO (10 lM for 48 h) versus vehicle treatment. N = 5 per group. Scale bar: 100 lm. C, D DHE staining of frozen sections of hearts from WT and miR-574 À/À mice under Veh. and ISO treatment or Sham and TAC operation (4 weeks post-surgery). N = 5 per group for (C) and n = 8 per group for (D). Scale bar: 20 lm. E TUNEL assay for heart tissue sections from WT and miR-574 À/À mice under Sham versus TAC surgery. N = 5 per group. Scale bar: 5 lm. ▸ Figure EV3. Phenotypic characterization of the therapeutic model using miR-574-5p/3p mimic injection in mice subject to TAC surgery.
A RT-qPCR of miR-574-5p and miR-574-3p after injections of the miRNA mimics for 2 or 4 days. N = 4 per group. B Creatine kinase test and alanine transaminase (ALT) assay in kidneys and livers from miRNA mimic injected mice. N = 4 per group. TAC: transverse aortic constriction; Sham is control mock surgery for TAC surgery (no constriction). C RT-qPCR of hypertrophy and fibrosis marker genes in the hearts of therapeutic mouse models. N = 5 per group. D DHE staining of frozen sections of hearts from WT mice under treatment with miRNA mimics in TAC-induced HF mouse models. Sham operation was used as a control for TAC. N = 4 per group. Scale bar: 20 lm. E ATP level in heart lysates from WT mice under treatment with miRNA mimics in TAC-induced HF mouse models (6 weeks post-surgery). N = 4 per group. F TUNEL assay of murine hearts in the therapeutic models. N = 4 per group. Scale bar: 5 lm. Green color: a-actinin immunostaining signal. Red color: TUNEL signal.
Blue color: DAPI signal. G miR-574-5p and miR-574-3p decreased ISO-activated mouse CM hypertrophy. Isolated mouse primary neonatal CMs were stained by FITC phalloidin. The cells were transfected with negative control miRNA, miR-574-3p, and miR-574-5p mimics, followed by ISO (10 lm Figure EV5. miR-574-FAM210A axis modulates mitochondrial protein expression and mitochondrial activity in murine hearts. A, B IF analysis of protein expression of ETC complex protein components in WT and miR-574 KO hearts 3 days after TAC surgery. n = 5 hearts per group with> 100 CMs measured per group. Scale bar: 10 lm. The dashed line in the violin plot shows medium value for the group and the dotted lines represent two quartile lines in each group. C Measurement of mitochondrial copy number in WT and miR-574 KO hearts 3 days after TAC surgery (n = 4). D Measurement of mitochondrial electron transport chain complex enzymatic activities in heart lysates of WT and miR-574 KO mice at 4 weeks after TAC or Sham surgery (n = 5 per group).
Data information: Data were presented as mean AE SEM. P values were calculated by two-way ANOVA with Tukey's multiple comparisons test (B-D). Source data are available online for this figure.
A C B D Figure EV5.