DGCR8 Research Articles

MicroRNA and Rare Human Diseases

Background: The role of microRNAs (miRNAs) in the pathogenesis of rare genetic disorders has been gradually discovered. MiRNAs, a class of small non-coding RNAs, regulate gene expression by silencing target messenger RNAs (mRNAs). Their biogenesis involves transcription into primary miRNA (pri-miRNA), processing by the DROSHA–DGCR8 (DiGeorge syndrome critical region 8) complex, exportation to the cytoplasm, and further processing by DICER to generate mature miRNAs. These mature miRNAs are incorporated into the RNA-induced silencing complex (RISC), where they modulate gene expression. Methods/Results: The dysregulation of miRNAs is implicated in various Mendelian disorders and familial diseases, including DICER1 syndrome, neurodevelopmental disorders (NDDs), and conditions linked to mutations in miRNA-binding sites. We summarized a few mechanisms how miRNA processing and regulation abnormalities lead to rare genetic disorders. Examples of such genetic diseases include hearing loss associated with MIR96 mutations, eye disorders linked to MIR184 mutations, and skeletal dysplasia involving MIR140 mutations. Conclusions: Understanding these molecular mechanisms is crucial, as miRNA dysregulation is a key factor in the pathogenesis of these conditions, offering significant potential for the diagnosis and potential therapeutic intervention.

Regulatory role and molecular mechanism of METTL14 in vascular endothelial cell injury in preeclampsia.

Preeclampsia (PE) is a pregnancy-related disease characterized by vascular endothelial cell injury. This study aimed to investigate the role of methyltransferase-like protein 14 (METTL14) in vascular endothelial cell injury in PE. The PE cell model was established by treating human umbilical vein endothelial cells (HUVECs) with tumor necrosis factor-alpha (TNF-α) in vitro. METTL14 and forkhead box protein 1 (FOXP1) were silenced, and miR-34a-5p was overexpressed in HUVECs to evaluate their effects. HUVEC viability, apoptosis, and the levels of intercellular adhesion molecule 1, vascular cell adhesion molecule 1, and endothelin-1 were measured. The N6-methyladenosine (m6A) modification of pri-miR-34a-5p was quantified. The interactions between miR-34a-5p, DiGeorge syndrome critical region 8, and m6A enrichment in miR-34a-5p were analyzed. The relationship between miR-34a-5p and FOXP1 was also verified. The results showed that the expressions of METTL14, FOXP1, and miR-34a-5p were determined. METTL14 expression was elevated in the TNF-α-induced HUVEC injury model. Silencing METTL14 improved HUVEC viability, inhibited apoptosis, and reduced endothelial inflammation. METTL14 promoted miR-34a-5p expression through m6A modification. Overexpression of miR-34a-5p or silencing of FOXP1 reversed the protective effects of METTL14 silencing on cell injury in the PE model. In conclusion, METTL14 mediated m6A modification to promote miR-34a-5p expression, leading to the inhibition of FOXP1 expression, which aggravated endothelial cell damage in the PE cell model.

Advance of research on 22q11.2 deletion syndrome

22q11.2 deletion syndrome (22q11.2DS) is the most common chromosomal microdeletion disorder. Its phenotype is highly variable with incomplete penetrance. 22q11.2DS is a rare disease, and the research progress is relatively slow, which has restricted its treatment and intervention. In recent years, much progress has been made in the pathogenic mechanism and genome-wide association study of 22q11.2DS. In this review, the pathogenesis of 22q11.2DS was summarized. Thereafter, the molecular and pathological mechanisms of TBX1 and DGCR8 genes were clarified. Finally, factors affecting the penetrance of cardiac and immune system phenotypes were reviewed. This review may enhance the understanding of 22q11.2DS and has important clinical implications on the prenatal diagnosis, genetic counseling, treatment and intervention of this disease.

MiR-29a-3p orchestrates key signaling pathways for enhanced migration of human mesenchymal stem cells

BackgroundThe homing of human mesenchymal stem cells (hMSCs) is crucial for their therapeutic efficacy and is characterized by the orchestrated regulation of multiple signaling modules. However, the principal upstream regulators that synchronize these signaling pathways and their mechanisms during cellular migration remain largely unexplored.MethodsmiR-29a-3p was exogenously expressed in either wild-type or DiGeorge syndrome critical region 8 (DGCR8) knockdown hMSCs. Multiple pathway components were analyzed using Western blotting, immunohistochemistry, and real-time quantitative PCR. hMSC migration was assessed both in vitro and in vivo through wound healing, Transwell, contraction, and in vivo migration assays. Extensive bioinformatic analyses using gene set enrichment analysis and Ingenuity pathway analysis identified enriched pathways, upstream regulators, and downstream targets.ResultsThe global depletion of microRNAs (miRNAs) due to DGCR8 gene silencing, a critical component of miRNA biogenesis, significantly impaired hMSC migration. The bioinformatics analysis identified miR-29a-3p as a pivotal upstream regulator. Its overexpression in DGCR8-knockdown hMSCs markedly improved their migration capabilities. Our data demonstrate that miR-29a-3p enhances cell migration by directly inhibiting two key phosphatases: protein tyrosine phosphatase receptor type kappa (PTPRK) and phosphatase and tensin homolog (PTEN). The ectopic expression of miR-29a-3p stabilized the polarization of the Golgi apparatus and actin cytoskeleton during wound healing. It also altered actomyosin contractility and cellular traction forces by changing the distribution and phosphorylation of myosin light chain 2. Additionally, it regulated focal adhesions by modulating the levels of PTPRK and paxillin. In immunocompromised mice, the migration of hMSCs overexpressing miR-29a-3p toward a chemoattractant significantly increased.ConclusionsOur findings identify miR-29a-3p as a key upstream regulator that governs hMSC migration. Specifically, it was found to modulate principal signaling pathways, including polarization, actin cytoskeleton, contractility, and adhesion, both in vitro and in vivo, thereby reinforcing migration regulatory circuits.

Circulating DiGeorge Syndrome Critical Region (5) as a Tumor Suppressor Gene in Hepatocellular Carcinoma

Circulating DiGeorge Syndrome Critical Region (5) as a Tumor Suppressor Gene in Hepatocellular Carcinoma

Silencing METTL14 alleviates liver injury in non-alcoholic fatty liver disease by regulating mitochondrial homeostasis.

Mitochondrial dysfunction is an important pathogenic factor in non-alcoholic fatty liver disease (NAFLD). Methyltransferase-like 14 (METTL14) has been implicated in mitochondrial fission processes. This research aimed to investigate the mechanism of METTL14 in the mitochondrial function of NAFLD. We first established NAFLD mouse models and cell models, recording body and liver weights and examining pathological changes in liver tissues. Subsequently, serum levels of liver function indices (aspartate aminotransferase [AST], alanine aminotransferase [ALT], total cholesterol [TC], and triglycerides [TG]), inflammatory markers (tumor necrosis factor-alpha [TNF-α], interleukin [IL]-6, and IL-1β), and mitochondrial dysfunction indicators (fission 1 protein [Fis1], dynamin-related protein 1 [Drp1], mitofusin 2 [Mfn2], SID1 transmembrane family member 2 [SIDT2], and mitochondrial membrane potential [MMP]) in the liver and cells were evaluated. The N6-methyladenosine (m6A) modification level of primary microRNA (pri-miRNA) and m6A enrichment on pri-miR-34a were quantified. Co-immunoprecipitation and dual-luciferase reporter gene assays were utilized to validate gene interactions. Our findings revealed highly elevated METTL14 expression in NAFLD mouse and cell models. Silencing METTL14 reduced weight gain and mitigated adverse liver function indices, inflammation, hepatic steatosis, and structural damage in NAFLD mice. It also led to a decrease in Fis1/Drp1 levels and an increase in MMP/Mfn2 in the liver and cells. Moreover, METTL14 increased the m6A level, promoting the binding of DiGeorge syndrome critical region 8 (DGCR8) to pri-miR-34a, which enhanced miR-34a-5p expression. Databases and dual-luciferase reporter gene assays indicated that miR-34a-5p could suppress SIDT2 expression. The overexpression of miR-34a-5p or inhibition of SIDT2 expression negated the alleviative effects of METTL14 silencing on mitochondrial homeostasis imbalance. In conclusion, METTL14, through m6A modification, modulates the miR-34a-5p/SIDT2 axis, impairing mitochondrial homeostasis in NAFLD.

Evaluation of expression levels of microRNA processing elements in patients with sudden sensorineural hearing loss

BackgroundMicroRNAs have a significant role in the function and development of the hearing system. Idiopathic sudden sensorineural hearing loss (SSNHL) is a complicated disorder with no long-established reason. Since microRNAs play imperative roles in every aspect of the neural system, their dysregulation may contribute to the onset of SSNHL. The current study aimed to assess the expression patterns of microRNA processing elements (DROSHA, DICER, and DGCR-8) as the vital factors in microRNA biology that can affect the expression levels of microRNA. This study assessed DROSHA, DICER, and DGCR-8 mRNA expression levels in the peripheral blood mononuclear cells (PBMC) of 50 patients with SSNHL and 50 matched controls. After the isolation of PBMC, total RNA was extracted, and the expression levels of DROSHA, DICER, and DGCR-8 genes were evaluated using quantitative real-time PCR.ResultsThe results illustrated significant up-regulation of DICER and DGCR-8 genes in SSNHL patients at the mRNA level. Furthermore, despite no significant change in DROSHA level, DICER and DGCR-8 were significantly correlated with SSNHL. However, there was no significant correlation between these gene expressions and the clinicopathological features of patients.ConclusionThis study verified for the first time that the DGCR_8 and DICER mRNA expression levels were significantly up-regulated in patients with SSNHL, proposing that microRNAs and their processing pathways play key roles in the progression and development of SSNHL.

The role of long noncoding RNA DGCR5 in cancers: Focus on molecular targets.

Long noncoding RNAs (lncRNAs) are major components of cellular transcripts that are emerging as important players in various biological pathways. Due to their specific expression and functional diversity in a variety of cancers, lncRNAs have promising applications in cancer diagnosis, prognosis, and therapy. Studies have shown that lncRNA DiGeorge syndrome critical region gene 5 (DGCR5) with high specificity and accuracy has the potential to become biomarkers in cancers. LncRNA DGCR5 can be noninvasively extracted from body fluids, tissues, and cells, and can be used as independent or auxiliary biomarkers to improve the accuracy of diagnosis or prognosis. Now, the underlying mechanisms of lncRNAs such as DGCR5 were explored as therapeutic targets, which have been investigated in clinical trials of several cancers. The DGCR5 lacks an appropriate animal model, which is necessary to gain greater knowledge of their functions. While some studies on the uses of DGCR5 have been carried out, the small sample size makes them unreliable. In this review, we presented a compilation of recent publications addressing the potential of lncRNA DGCR5 that could be considered as biomarkers or therapeutic targets, with the hopes of providing promised implications for future cancer therapy.

N6-methyladenosine-induced miR-182-5p promotes multiple myeloma tumorigenesis by regulating CAMK2N1.

Methyltransferase like 3 (METTL3) has been reported to promote tumorigenesis of multiple myeloma (MM), however, the molecular mechanism still needs further research. The N6-methyladenosine (m6A) level in tissues or cells was measured by m6A kit and dot blot assay. The mRNA and protein expression were detected by quantitative real-time PCR (RT-qPCR) and Western blot, respectively. The cell counting kit-8 and colony formation assay were used to detect the cell proliferation. Coimmunoprecipitation (Co-IP) experiment verified the binding of two proteins. The luciferase reporter experiment demonstrated the targeted binding of miR-182-5p and CaMKII inhibitor 1 (CAMK2N1). More importantly, tumor growth was measured in xenograft mice. Our data showed that the expression of METTL3 was significantly increased in MM patients' samples and MM cells. METTL3 overexpression promoted MM cells proliferation, and METTL3 knockdown inhibited MM cells proliferation. Mechanically, METTL3-dependent m6A participated in DiGeorge syndrome critical region 8 (DGCR8)-mediated maturation of pri-miR-182. Upregulation of miR-182-5p further enhanced the promoting proliferation effect of METTL3 overexpression on MM cells. Moreover, the luciferase reporter gene experiment proved that miR-182-5p targetedly regulated CAMK2N1 expression. Xenograft tumor in nude mice further verified that METTL3 promoted MM tumor growth through miR-182/CAMK2N1 signal axis. In summary, the METTL3/miR-182-5p/CAMK2N1 axis plays an important role in MM tumorigenesis, which may provide a new target for MM therapy.

Dgcr8 functions in the secondary heart field for outflow tract and right ventricle development in mammals

The DGCR8 gene, encoding a critical miRNA processing protein, maps within the hemizygous region in patients with 22q11.2 deletion syndrome. Most patients have malformations of the cardiac outflow tract that is derived in part from the anterior second heart field (aSHF) mesoderm. To understand the function of Dgcr8 in the aSHF, we inactivated it in mice using Mef2c-AHF-Cre. Inactivation resulted in a fully penetrant persistent truncus arteriosus and a hypoplastic right ventricle leading to lethality by E14.5. To understand the molecular mechanism for this phenotype, we performed gene expression profiling of the aSHF and the cardiac outflow tract with right ventricle in conditional null versus normal mouse littermates at stage E9.5 prior to morphology changes. We identified dysregulation of mRNA gene expression, of which some are relevant to cardiogenesis. Many pri-miRNA genes were strongly increased in expression in mutant embryos along with reduced expression of mature miRNA genes. We further examined the individual, mature miRNAs that were decreased in expression along with pri-miRNAs that were accumulated that could be direct effects due to loss of Dgcr8. Among these genes, were miR-1a, miR-133a, miR-134, miR143 and miR145a, which have known functions in heart development. These early mRNA and miRNA changes may in part, explain the first steps that lead to the resulting phenotype in Dgcr8 aSHF conditional mutant embryos.

Results of karyotype analysis and 22q11.2 (DiGeorge Syndrome critical region) deletion investigation in fetal cardiac system anomalies

The results of karyotype analysis and 22q11.2 ( DiGeorge Syndrome critical region) deletion investigation in fetal cardiac system anomalies 
 Objective: We aimed to determine the presence of accompanying chromosomal anomalies and 22q11.2 deletion in patients with cardiac system anomalies.
 Material and Methods: In our retrospective and cross-sectional study, 99 fetuses born with cardiac anomalies were evaluated in terms of chromosomal anomalies. 36 fetuses were evaluated for 22q11.2 deletion. Fetuses who were diagnosed with prenatal congenital heart defect and underwent invasive prenatal diagnostic tests for fetal karyotyping between 01.01.2010 and 30.06.2017 at a tertiary cancer center were included in the study.
 Results: Of 99 cases, 48 (48.4%) had only cardiac anomalies and 51 (51.5%) had non-cardiac anomalies. Chromosomal anomalies were found in 37 (37.4%) of the cases. Autosomal trisomy 18(43.2%) and autosomal trisomy 21(32.4%) were the most common chromosomal anomalies. The study results support the strong association of chromosomal changes and cardiac malformation, especially in septal defects, atrioventricular septal defects, and conotruncal malformations. Deletion was detected in one(2.8%) of 36 cases evaluated for 22q11.2 deletion. The fetus with this deletion had isolated tetralogy of fallot and had no extracardiac anomaly.
 Conclusion: In cases with cardiac anomalies, isolated or accompanied by extracardiac anomalies, investigations should be made in terms of underlying chromosomal diseases in the perinatal evaluation. In addition, the investigation of 22q11.2 deletion in fetuses with conotruncal cardiac anomalies should be included in prenatal genetic examination.
 Keywords: DiGeorge Syndrome, Fetal Anomalies, karyotype, trisomy, 22q11.2 Deletion Syndrome

DiGeorge syndrome critical region gene 2 (DGCR2), a schizophrenia risk gene, regulates dendritic spine development through cell adhesion

BackgroundDendritic spines are the sites of excitatory synapses on pyramidal neurons, and their development is crucial for neural circuits and brain functions. The spine shape, size, or number alterations are associated with neurological disorders, including schizophrenia. DiGeorge syndrome critical region gene 2 (DGCR2) is one of the deleted genes within the 22q11.2 deletion syndrome (22q11DS), which is a high risk for developing schizophrenia. DGCR2 expression was reduced in schizophrenics. However, the pathophysiological mechanism of DGCR2 in schizophrenia or 22q11DS is still unclear.ResultsHere, we report that DGCR2 expression was increased during the neurodevelopmental period and enriched in the postsynaptic densities (PSDs). DGCR2-deficient hippocampal neurons formed fewer spines. In agreement, glutamatergic transmission and synaptic plasticity were decreased in the hippocampus of DGCR2-deficient mice. Further molecular studies showed that the extracellular domain (ECD) of DGCR2 is responsible for its transcellular interaction with cell adhesion molecule Neurexin1 (NRXN1) and spine development. Consequently, abnormal behaviors, like anxiety, were observed in DGCR2-deficient mice.ConclusionsThese observations indicate that DGCR2 is a novel cell adhesion molecule required for spine development and synaptic plasticity, and its deficiency induces abnormal behaviors in mice. This study provides a potential pathophysiological mechanism of DGCR2 in 22q11DS and related mental disorders.

METTL14 mediates m6a modification on osteogenic proliferation and differentiation of bone marrow mesenchymal stem cells by regulating the processing of pri-miR-873.

N6-methyl-adenosine (m6a) is involved in the occurrence and development of various diseases such as autogenic immune disease and tumors. Methyltransferases regulate primary (pri)-microRNA (miRNA/miR) processing by mediating m6a modifications, consequently affecting pathological processes including immune-related diseases by regulating both innate and adaptive immune cells. However, the roles of m6a on the biological functions of bone marrow mesenchymal stem cells (BMSCs) remain to be elucidated. The relative expression levels of methyltransferase-like 14 (METTL14) and other methyltransferases, demethylases, and miR-873 in bone samples from patients with osteoporosis and from normal individuals were measured by reverse transcription-quantitative PCR. Cell Counting Kit-8 assay was used to examine the proliferation of BMSCs. Co-immunoprecipitation (Co-IP) was used to investigate the binding of METTL14 to DiGeorge syndrome critical region 8 (DGCR8). RNA immunoprecipitation (RIP) was used to examine the binding of METTL14 to pri-miR-873. METTL14 and m6a modifications were highly detected in patients with osteoporosis compared with the controls. Co-IP results indicated that silencing of METTL14 reduced METTL14 and m6a modification levels in BMSCs. Downregulation of METTL14 significantly promoted the proliferation of BMSCs. RIP results suggested that METTL14/m6a methylation modification promoted the processing of pri-miR-873 by binding to DGCR8 in BMSCs. Furthermore, overexpression of miR-873 inhibited the proliferation of BMSCs. The results also showed that miR-873 mimics significantly inhibited the proliferation in small interfering (si)-METTL14 transfected BMSCs; however, miR-873 inhibitors markedly promoted the proliferation of si-METTL14 transfected BMSCs. METTL14 and m6a modifications were upregulated in osteoporosis samples. METTL14 promoted the processing of pri-miR-873 into mature miR-873 by regulating m6a modification. Furthermore, overexpression of miR-873 significantly inhibited the proliferation of BMSCs. Therefore, the METTL14/m6a/miR-873 axis may be a potential target for the treatment of osteoporosis.

ADAR1 Non-Editing Function in Macrophage Activation and Abdominal Aortic Aneurysm.

Macrophage activation plays a critical role in abdominal aortic aneurysm (AAA) development. However, molecular mechanisms controlling macrophage activation and vascular inflammation in AAA remain largely unknown. The objective of the study was to identify novel mechanisms underlying adenosine deaminase acting on RNA (ADAR1) function in macrophage activation and AAA formation. Aortic transplantation was conducted to determine the importance of nonvascular ADAR1 in AAA development/dissection. Ang II (Angiotensin II) infusion of ApoE-/- mouse model combined with macrophage-specific knockout of ADAR1 was used to study ADAR1 macrophage-specific role in AAA formation/dissection. The relevance of macrophage ADAR1 to human AAA was examined using human aneurysm specimens. Moreover, a novel humanized AAA model was established to test the role of human macrophages in aneurysm formation in human arteries. Allograft transplantation of wild-type abdominal aortas to ADAR1+/- recipient mice significantly attenuated AAA formation, suggesting that nonvascular ADAR1 is essential for AAA development. ADAR1 deficiency in hematopoietic cells decreased the prevalence and severity of AAA while inhibited macrophage infiltration and aorta wall inflammation. ADAR1 deletion blocked the classic macrophage activation, diminished NF-κB (nuclear factor kappa B) signaling, and enhanced the expression of a number of anti-inflammatory microRNAs. Mechanistically, ADAR1 interacted with Drosha to promote its degradation, which attenuated Drosha-DGCR8 (DiGeorge syndrome critical region 8) interaction, and consequently inhibited pri- to pre-microRNA processing of microRNAs targeting IKKβ, resulting in an increased IKKβ (inhibitor of nuclear factor kappa-B) expression and enhanced NF-κB signaling. Significantly, ADAR1 was induced in macrophages and interacted with Drosha in human AAA lesions. Reconstitution of ADAR1-deficient, but not the wild type, human monocytes to immunodeficient mice blocked the aneurysm formation in transplanted human arteries. Macrophage ADAR1 promotes aneurysm formation in both mouse and human arteries through a novel mechanism, that is, Drosha protein degradation, which inhibits the processing of microRNAs targeting NF-kB signaling and thus elicits macrophage-mediated vascular inflammation in AAA.

The relationship between prognosis of patients with traumatic brain injury and microRNA biogenesis proteins.

This study aims to investigate whether the expression levels of proteins involved in microRNA (miRNA) biogenesis vary in early- and late-stage traumatic brain injury (TBI) patients and to evaluate its effect on prognosis. Dicer, Drosha, DiGeorge Syndrome Critical Region eight (DGCR8), Exportin5 (XPO5), and Argonaute2 (AGO2) levels were measured in the blood samples of severe TBI patients collected 4-6 h and 72 h after the trauma and compared with the control group. Prognostic follow-up of the patients was performed using the Glasgow Coma Scale score. There were no statistically significant changes in the expression of the miRNA biogenesis proteins Dicer, Drosha, DGCR8, XPO5, and AGO2 in patients with severe TBI. However, the expression of Dicer increased in the patients who improved from the severe TBI grade to the mild TBI grade, and the expression of AGO2 decreased in most of these patients. The Dicer expression profile was found to increase in patients discharged from the intensive care unit in a short time. MicroRNAs and their biogenesis proteins may guide prognostic and therapeutic decisions for patients with TBI in the future.

Activation of the HNRNPA2B1/miR-93-5p/FRMD6 axis facilitates prostate cancer progression in an m6A-dependent manner.

It is becoming increasingly clear that N6-methyladenosine (m6A) plays a key role in post-transcriptional modification of eukaryotic RNAs in cancer. The regulatory mechanism of m6A modifications in prostate cancer is still not completely elucidated. Heterogeneous nuclear ribonucleoprotein A2/B1 (HNRNPA2B1), an m6A reader, has been revealed to function as an oncogenic RNA-binding protein. However, its contribution to prostate cancer progression remains poorly understood. Here, we found that HNRNPA2B1 was highly overexpressed and correlated with a poor prognosis in prostate cancer. In vitro and in vivo functional experiments demonstrated that HNRNPA2B1 knockout impaired proliferation and metastasis of prostate cancer. Mechanistic studies indicated that HNRNPA2B1 interacted with primary miRNA-93 and promoted its processing by recruiting DiGeorge syndrome critical region gene 8 (DGCR8), a key subunit of the Microprocessor complex, in an METTL3-dependent mechanism, while HNRNPA2B1 knockout significantly restored miR-93-5p levels. HNRNPA2B1/miR-93-5p downregulated FERM domain-containing protein 6 (FRMD6), a cancer suppressor, and enhanced proliferation and metastasis in prostate cancer. In conclusion, our findings identified a novel oncogenic axis, HNRNPA2B1/miR-93-5p/FRMD6, that stimulates prostate cancer progression via an m6A-dependent manner.

The Potential Role of Dysregulated miRNAs in Adolescent Idiopathic Scoliosis and 22q11.2 Deletion Syndrome.

Background: Adolescent idiopathic scoliosis (AIS), affecting 2-4% of adolescents, is a multifactorial spinal disease. Interactions between genetic and environmental factors can influence disease onset through epigenetic mechanisms, including DNA methylation, histone modifications and miRNA expression. Recent evidence reported that, among all clinical features in individuals with 22q11.2 deletion syndrome (DS), scoliosis can occur with a higher incidence than in the general population. Methods: A PubMed and Ovid Medline search was performed for idiopathic scoliosis in the setting of 22q11.2DS and miRNA according to PRISMA guidelines. Results: Four papers, accounting for 2841 individuals, reported clinical data about scoliosis in individuals with 22q11.2DS, showing that approximately 35.1% of the individuals with 22q11.2DS developed scoliosis. Conclusions: 22q11.2DS could be used as a model for the study of AIS. The DGCR8 gene seems to be essential for microRNA biogenesis, which is why we propose that a possible common pathological mechanism between scoliosis and 22q11.2DS could be the dysregulation of microRNA expression. In the current study, we identified two miRNAs that were altered in both 22q11.2DS and AIS, miR-93 and miR-1306, thus, corroborating the hypothesis that the two diseases share common molecular alterations.

Construction of an immune-related ceRNA network to screen for potential diagnostic markers for autism spectrum disorder.

Purpose: The diagnosis of autism spectrum disorder (ASD) is reliant on evaluation of patients' behavior. We screened the potential diagnostic and therapeutic targets of ASD through bioinformatics analysis. Methods: Four ASD-related datasets were downloaded from the Gene Expression Omnibus database. The "limma" package was employed to analyze differentially expressed messenger (m)RNAs, long non-coding (lnc)RNAs, and micro (mi)RNAs between ASD patients and healthy volunteers (HVs). We constructed a competing endogenous-RNA (ceRNA) network. Enrichment analyses of key genes were undertaken using the Gene Ontology database and Kyoto Encyclopedia of Genes and Genomes database. The ImmucellAI database was used to analyze differences in immune-cell infiltration (ICI) in ASD and HV samples. Synthetic analyses of the ceRNA network and ICI was done to obtain a diagnostic model using LASSO regression analysis. Analyses of receiver operating characteristic (ROC) curves were done for model verification. Results: The ceRNA network comprised 49 lncRNAs, 30 miRNAs, and 236 mRNAs. mRNAs were associated with 41 cellular components, 208 biological processes, 39 molecular functions, and 35 regulatory signaling pathways. Significant differences in the abundance of 10 immune-cell species between ASD patients and HVs were noted. Using the ceRNA network and ICI results, we constructed a diagnostic model comprising five immune cell-associated genes: adenosine triphosphate-binding cassette transporter A1 (ABCA1), DiGeorge syndrome critical region 2 (DGCR2), glucose-fructose oxidoreductase structural domain gene 1 (GFOD1), glutaredoxin (GLRX), and SEC16 homolog A (SEC16A). The diagnostic performance of our model was revealed by an area under the ROC curve of 0.923. Model verification was done using the validation dataset and serum samples of patients. Conclusion: ABCA1, DGCR2, GFOD1, GLRX, and SEC16A could be diagnostic biomarkers and therapeutic targets for ASD.

Partial Disturbance of Microprocessor Function in Human Stem Cells Carrying a Heterozygous Mutation in the DGCR8 Gene.

Maturation of microRNAs (miRNAs) begins by the "Microprocessor" complex, containing the Drosha endonuclease and its partner protein, "DiGeorge Syndrome Critical Region 8" (DGCR8). Although the main function of the two proteins is to coordinate the first step of precursor miRNAs formation, several studies revealed their miRNA-independent functions in other RNA-related pathways (e.g., in snoRNA decay) or, for the DGCR8, the role in tissue development. To investigate the specific roles of DGCR8 in various cellular pathways, we previously established a human embryonic stem-cell (hESC) line carrying a monoallelic DGCR8 mutation by using the CRISPR-Cas9 system. In this study, we genetically characterized single-cell originated progenies of the cell line and showed that DGCR8 heterozygous mutation results in only a modest effect on the mRNA level but a significant decrease at the protein level. Self-renewal and trilineage differentiation capacity of these hESCs were not affected by the mutation. However, partial disturbance of the Microprocessor function could be revealed in pri-miRNA processing along the human chromosome 19 miRNA cluster in several clones. With all these studies, we can demonstrate that the mutant hESC line is a good model to study not only miRNA-related but also other "noncanonical" functions of the DGCR8 protein.

The developmentally timed decay of an essential microRNA family is seed-sequence dependent.

SUMMARYMicroRNA (miRNA) abundance is tightly controlled by regulation of biogenesis and decay. Here, we show that the mir-35 miRNA family undergoes selective decay at the transition from embryonic to larval development in C. elegans. The seed sequence of the miRNA is necessary and largely sufficient for this regulation. Sequences outside the seed (3′ end) regulate mir-35 abundance in the embryo but are not necessary for sharp decay at the transition to larval development. Enzymatic modifications of the miRNA 3′ end are neither prevalent nor correlated with changes in decay, suggesting that miRNA 3′ end display is not a core feature of this mechanism and further supporting a seed-driven decay model. Our findings demonstrate that seed-sequence-specific decay can selectively and coherently regulate all redundant members of a miRNA seed family, a class of mechanism that has great biological and therapeutic potential for dynamic regulation of a miRNA family’s target repertoire.