Department of Radiobiology and Molecular Genetics, "Vinča" Institute of Nuclear Sciences, National Institute of the Republic of Serbia, University of Belgrade, Belgrade, Serbia.
Department of Molecular Biology and Endocrinology, "Vinča" Institute of Nuclear Sciences, National Institute of the Republic of Serbia, University of Belgrade, Belgrade, Serbia.
Sci Rep. 2022 Oct 22;12(1):17746. doi: 10.1038/s41598-022-22749-1.
Rare copy number variants (CNVs) are among the most common genomic disorders underlying CAKUT. miRNAs located in rare CNVs represent well-founded functional variants for human CAKUT research. The study aimed to identify and functionally interpret miRNAs most frequently affected by rare CNVs in CAKUT and to estimate the overall burden of rare CNVs on miRNA genes in CAKUT. The additional aim of this study was to experimentally confirm the effect of a rare CNV in CAKUT on candidate miRNA's expression and the subsequent change in mRNA levels of selected target genes. A database of CAKUT-associated rare CNV regions, created by literature mining, was used for mapping of the miRNA precursors. miRNAs and miRNA families, most frequently affected by rare CAKUT-associated CNVs, have been subjected to bioinformatic analysis. CNV burden analysis was performed to identify chromosomes with over/underrepresentation of miRNA genes in rare CNVs associated with CAKUT. A functional study was performed on HEK293 MIR484 KO and HEK293 WT cell lines, followed by the analysis of relative miRNA and mRNA target gene levels. 80% of CAKUT patients with underlying rare CNV had at least one miRNA gene overlapping the identified CNV. Network analysis of the most frequently affected miRNAs has revealed the dominant regulation of the two miRNAs, hsa-miR-484 and hsa-miR-185-5p. Additionally, miR-548 family members have shown substantial enrichment in rare CNVs in CAKUT. An over/underrepresentation of miRNA genes in rare CNVs associated with CAKUT was observed in multiple chromosomes, such as chr16, chr20, and chr21. A significant 0.37 fold downregulation of hsa-miR-484, followed by a notable upregulation of MDM2 and APAF1 and downregulation of NOTCH3 was detected in HEK293 MIR484 KO compared to HEK293 WT cell lines, supporting the study hypothesis. miRNA genes are frequently affected by rare CNVs in CAKUT patients. Understanding the potential of CNV-affected miRNAs to participate in CAKUT as genetic drivers represent a crucial implication for the development of novel therapeutic approaches.
罕见的拷贝数变异（CNVs）是导致 CAKUT 的最常见基因组疾病之一。位于罕见 CNVs 中的 miRNA 代表了人类 CAKUT 研究中合理的功能变异。本研究旨在鉴定和功能解释 CAKUT 中受罕见 CNVs 影响最频繁的 miRNA，并估计罕见 CNVs 对 CAKUT 中 miRNA 基因的总体负担。本研究的另一个目的是通过实验证实 CAKUT 中罕见 CNV 对候选 miRNA 表达的影响，以及随后对选定靶基因 mRNA 水平的变化。通过文献挖掘创建了一个与 CAKUT 相关的罕见 CNV 区域数据库，用于 miRNA 前体的映射。对受罕见 CAKUT 相关 CNV 影响最频繁的 miRNA 和 miRNA 家族进行了生物信息学分析。进行了 CNV 负担分析，以鉴定在与 CAKUT 相关的罕见 CNVs 中 miRNA 基因过度/低表达的染色体。在 HEK293 MIR484 KO 和 HEK293 WT 细胞系上进行了功能研究，随后分析了相对 miRNA 和 mRNA 靶基因水平。80%的具有潜在罕见 CNV 的 CAKUT 患者至少有一个 miRNA 基因与鉴定的 CNV 重叠。对受影响最频繁的 miRNA 进行的网络分析表明，两个 miRNA，hsa-miR-484 和 hsa-miR-185-5p，具有主导调节作用。此外，miR-548 家族成员在 CAKUT 中的罕见 CNVs 中表现出显著富集。在与 CAKUT 相关的罕见 CNVs 中观察到 miRNA 基因在多个染色体上的过度/低表达，例如 chr16、chr20 和 chr21。与 HEK293 WT 细胞系相比，在 HEK293 MIR484 KO 中检测到 hsa-miR-484 的显著下调 0.37 倍，随后 MDM2 和 APAF1 的显著上调以及 NOTCH3 的下调，这支持了研究假设。CAKUT 患者的 miRNA 基因经常受到罕见 CNVs 的影响。理解受 CNV 影响的 miRNA 作为遗传驱动因素参与 CAKUT 的潜力，代表了开发新治疗方法的关键意义。
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