Abstract
MicroRNAs (miRNAs) are emerging as effective therapeutic agents. When testing whether miR-145-5p could alleviate kidney injury, we unexpectedly found that extracellular vesicles loaded with miR-145-5p induced proteinuria and podocyte foot process effacement in normal control mice. To explore the mechanism of miR-145-5p’s toxicity to podocytes, we hypothesized that miR-145-5p could enter podocytes and inhibit genes essential for podocytes. We demonstrated that systemically administered miRNA can enter podocytes. Next, we predicted 611 podocyte essential genes based on single-cell RNA sequencing (RNA-seq) and found that 32 of them are predicted to be targeted by miR-145-5p. Functional annotation of the 32 podocyte essential genes revealed small GTPase-mediated signal transduction as the top pathway. We experimentally validated that miR-145-5p targeted Arhgap24 and Srgap1, the essential regulators of the Rho family of small GTPases, increased the activity of Rac1 and Cdc42, and reduced RhoA activity, accompanied by cellular injury, in podocytes. These results explain how miR-145-5p has deleterious effect on podocytes. Most importantly, our study provides a novel approach to investigate how a miRNA affects a given cell type, allowing not only identification of the molecular mechanism underlying an observed side effect of a miRNA drug but also prediction of miRNA drug toxicity on various cell types.
Highlights
MicroRNAs are posttranscriptional modulators of gene expression, and their aberrant expression is involved in the development of many diseases
We investigated whether systemically administered miR-145-5p could mitigate kidney injury in mouse models, and, unexpectedly, we found that treatment of normal control mice with extracellular vesicles (EVs) loaded with miR-145-5p resulted in albuminuria and podocyte foot process effacement
We identified candidate podocyte essential genes based on our mouse podocyte single-cell RNA sequencing (RNA-seq) data and experimentally validated their essentiality for podocytes, demonstrating the feasibility of the approach.[19]
Summary
MicroRNAs (miRNAs) are posttranscriptional modulators of gene expression, and their aberrant expression is involved in the development of many diseases. Global downregulation of miRNAs is frequently observed in tumors,[1] affecting many cellular processes, including cell cycle,[2] proliferation,[3] differentiation,[4] and apoptosis,[5] thereby facilitating tumorigenesis. Let-7 was found to diminish the mass of murine lung cancer,[7] and miR-26a adeno-assosiated virus suppressed tumorigenesis in mice.[8] In addition, other miRNAs have been tested for treatment of diseases (e.g., miR-34a,9 miR-16,10 miR-29,11 and miR-143/miR-14512,13). Therapeutic miRNAs are often loaded in liposomes,[14] minicells,[10] extracellular vehicles,[15] or other vehicles[16] and systemically administered. Administered vehicles and miRNAs can reach many normal tissues, including kidney. It is desirable to treat kidney diseases using miRNA-based drugs
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