Search Results (75)

Repetitive DNA represents over 50% of the human genome and is an abundant component of circulating cell-free DNA (cfDNA). We previously showed that cfDNA levels and integrity can predict survival in elderly patients with cardiovascular disease. Here, we aimed to clarify whether a low-pass next-generation sequencing (NGS) approach can characterize the repeat content of cfDNA. Considering the bimodal distribution of cfDNA fragment lengths, we examined the occurrence of repetitive DNA subfamilies separately in dinucleosomal (>250 bp) and mononucleosomal (≤250 bp) cfDNA sequences from 24 patients admitted for heart failure. An increase in the relative abundance of Alu repetitive elements was observed in the longer fraction, while alpha satellites were enriched in the mononucleosomal fraction. The relative abundance of Alu, ALR, and L1HS DNA in the dinucleosomal fraction correlated with different prognostic biomarkers, and Alu DNA was negatively associated with the presence of chronic kidney disease comorbidity. These results, together with the observed inverse correlation between Alu DNA abundance and cfDNA integrity, suggest that the composition of plasma cfDNA could be determined by multiple mechanisms in different physio-pathological conditions. In conclusion, low-pass NGS is an inexpensive method to analyze the cfDNA repeat landscape and identify new cardiovascular disease biomarkers. Full article

Paraspeckles are nuclear membraneless structures composed of a long non-coding RNA, Nuclear-Enriched-Abundant-Transcript-1, and RNA-binding proteins, which associate with numerous mRNAs. It is therefore believed that their cellular function is to sequester in the nucleus their associated proteins and/or target mRNAs. However, little is known about the molecular determinant in mRNA targets that allows their association to paraspeckles, except that inverted repeats of Alu sequences (IRAlu) present in the 3′UTR of mRNAs may allow this association. While in a previous study we established the list of paraspeckle target RNAs in a rat pituitary cell line, we did not find, however, inverted repeated SINEs, the rat equivalent of primate IRAlus in 3′UTR of these RNAs. By developing a candidate gene strategy, we selected a paraspeckle target gene, namely calreticulin mRNA, and we searched for other potential RNA recruitment elements in its 3′UTR, since 3′UTRs usually contain the sequence recognition for nuclear localization. We found a 15-nucleotide sequence surrounded in 5′ by a C-rich sequence, which is present as a tandem repeat in the 3′UTR of this mRNA and which is involved in the nuclear retention by paraspeckles. As shown by mass spectrometry analysis, 6 proteins bound to the 15-nucleotide sequence are paraspeckle proteins and constitute, therefore, bridging proteins between paraspeckles and target mRNAs. Full article

Aberrant DNA methylation is a hallmark of colorectal cancer (CRC), contributing to tumor progression through the silencing of tumor suppressor genes and activation of oncogenes. Indicaxanthin (IND), a dietary betalain pigment from Opuntia ficus indica, has shown antiproliferative effects in CRC models, yet its epigenetic impact remains unexplored. In this study, we investigated the effects of IND on the methylome of Caco-2 cells using Reduced Representation Bisulfite Sequencing (RRBS). IND induced a global hypermethylation profile, particularly at gene promoters and CpG islands. Among the differentially methylated genes, 60% were protein-coding, and 10% encoded transcription factors, including PAX5 and TFAP4, both hypermethylated at active enhancers. Functional enrichment analysis revealed pathways beyond canonical intestinal functions, suggesting altered cell identity and plasticity. Transcription factor targets (SOX10, NFKB1, AHR, ARNT) were significantly enriched among the affected genes, several of which are involved in transdifferentiation processes. Methylation changes also indicated potential reprogramming toward epithelial cell types from pulmonary or neuroectodermal origin. Moreover, IND induced selective hypomethylation of Alu elements on chromosome 21 and hypermethylation of rDNA loci, hinting at suppressed ribosomal biogenesis. Overall, these findings highlight the epigenetic remodeling potential of IND and its possible role in modulating cell fate and metabolism in CRC cells. Full article

Transposable elements (TEs) make up around half of the human genome. Their transcription is repressed in most somatic cells to maintain genome integrity and function. The repression is chiefly maintained by a combination of epigenetic modifications such as DNA methylation and histone modifications. However, recent research suggests that liver steatosis is associated with extensive changes to the hepatocyte epigenome. Furthermore, studies in mice have reported diet- and drug-induced changes to TE transcript levels in liver. The confirmation of these effects in human liver has not previously been undertaken. Here, we examined TE transcription in liver tissue from three patient cohorts with histologically confirmed liver steatosis caused by alcohol consumption or metabolic dysfunction. The quantitation of the number of transcripts with TE-homology in RNA-Seq data from a cohort of 90 bariatric surgery patients with metabolic dysfunction-associated steatotic liver disease (MASLD) revealed a trend for the reduction in TEs of all classes due to increasing steatosis, but no effect of fibrosis. This pattern was also present in a separate cohort of MASLD and HCC patients, as RT-qPCR also showed a reduction in Alu element transcripts in advanced steatosis, but again, no effect of fibrosis. Contrastingly, in a cohort of alcohol-related liver disease patients, the reduction in LINE-1 transcripts was associated with either increased steatosis or increased fibrosis. Moreover, the examination of LINE-1 DNA methylation levels in the MASLD and HCC cohort indicated that DNA methylation was also negatively associated with LINE-1 transcription in MASLD. This study suggests that TE transcript levels in human liver are slightly reduced by steatosis, that DNA methylation is an influential epigenetic regulator of LINE-1 retrotransposon transcription in steatosis, and that Alu transcript levels in background liver could be a new biomarker for HCC in cirrhotic and non-cirrhotic MASLD. Full article

Breast cancer (BC) and prostate cancer (PCa) are major health problems for women and men worldwide. Although therapeutic approaches have increased, the complexity associated with their heterogeneity and progression requires better ways to monitor them over time. Cell-free DNA integrity (cfDI) represents a viable alternative to needle biopsy and has the potential to be representative of cancer at all stages. In addition to the advantages of liquid biopsy in terms of cost and reduced invasiveness, cfDI can be used to detect repetitive DNA elements (e.g., ALU and LINE1), which could circumvent the problem of mutational heterogeneity in BC and PCa. In this review, we summarise the latest findings on cfDI studies in BC and PCa. The results show that cfDI has the potential to improve early detection, metastasis, and recurrence of BC, while limited studies prevent its clinical value in PCa from being fully defined. However, it is expected that further studies in the near future will help to introduce the use of cfDI as another biomarker for the clinical monitoring of BC and PCa patients. Full article

Circular RNAs (circRNAs) are a class of unique transcripts characterized by a covalently closed loop structure, which differentiates them from conventional linear RNAs. The formation of circRNAs occurs co-transcriptionally and post-transcriptionally through a distinct type of splicing known as back-splicing, which involves the formation of a head-to-tail splice junction between a 5′ splice donor and an upstream 3′ splice acceptor. This process, along with exon skipping, intron retention, cryptic splice site utilization, and lariat-driven intron processing, results in the generation of three main types of circRNAs (exonic, intronic, and exonic–intronic) and their isoforms. The intricate biogenesis of circRNAs is regulated by the interplay of cis-regulatory elements and trans-acting factors, with intronic Alu repeats and RNA-binding proteins playing pivotal roles, at least in the formation of exonic circRNAs. Various hypotheses regarding pathways of circRNA turnover are forwarded, including endonucleolytic cleavage and exonuclease-mediated degradation; however, similarly to the inconclusive nature of circRNA biogenesis, the process of their degradation and the factors involved remain largely unclear. There is a knowledge gap regarding whether these processes are guided by universal pathways or whether each category of circRNAs requires special tools and particular mechanisms for their life cycles. Understanding these factors is pivotal for fully comprehending the biological significance of circRNAs. This review provides an overview of the various pathways involved in the biogenesis and degradation of different types of circRNAs and explores key factors that have beneficial or adverse effects on the formation and stability of these unique transcripts in higher eukaryotes. Full article

Background/Objectives: The highly polymorphic Major Histocompatibility Complex (MHC) genomic region, located on the short arm of chromosome 6, is implicated genetically in Parkinson’s disease (PD), a progressive neurodegenerative disorder with motor and non-motor symptoms. Previously, we reported significant associations between SINE-VNTR-Alu (SVA) expression quantitative trait loci (eQTLs) and Human Leucocyte Antigen (HLA) class I genotypes in PD. In this study, we aimed to evaluate SVA associations and their regulatory effects on transposable element (TE) transcription in the MHC class I region. Methods: Transcriptome data from the peripheral blood cells of 1530 individuals in the Parkinson’s Progression Markers Initiative (PPMI) cohort were reanalyzed for TE and gene expression using publicly available bioinformatics tools, including Salmon and Matrix-eQTL. Results: Four structurally polymorphic SVAs regulated the transcription of 18 distinct clusters of 235 TE loci, comprising LINEs (33%), SINEs (19%), LTRs (35%), and ancient transposon DNA elements (12%) located near HLA genes. The transcribed TEs were predominantly short, with an average length of 445 nucleotides. The regulatory effects of these SVAs varied significantly in terms of TE types, numbers, and transcriptional activation or repression. The SVA-regulated TE RNAs in blood cells appear to function as enhancer-like elements, differentially influencing the expression of HLA class I genes, non-HLA genes, and noncoding RNAs. Conclusions: These findings highlight the roles of SVAs and their associated TEs in the complex regulatory networks governing coding and noncoding gene expression in the MHC class I region, with potential implications for immune function and disease susceptibility. Full article

Background/Objective: Large genomic rearrangements of PALB2 gene, particularly deletions and duplications, have been linked to hereditary breast–ovarian cancer. Our research specifically focuses on delineating the intronic breakpoints associated with rearrangements of PALB2 exon 11, which is crucial for understanding the mechanisms underlying these genomic changes in patients with hereditary breast and ovarian syndrome. Methods: By using next-generation sequencing, we identified one duplication and three deletions of PALB2 exon 11, confirmed by Multiplex Ligation-Dependent Probe Amplification analysis. To assess the impact on transcription and potential splicing issues, reverse-transcription PCR was performed on patients’ RNA. For the detailed characterization of intronic breakpoints, the primer walking approach and long-range PCR were implemented, followed by Sanger sequencing. Results: Our analysis revealed a tandem duplication of 5134 base pairs (bp) mediated by AluY repeats located in introns 10 and 11, respectively. Moreover, identical deletions were identified in three unrelated patients, encompassing an approximate 8050 bp region mediated by AluSx elements. Both genomic alterations resulted in a truncated PALB2 protein due to the introduction of a premature stop codon. Conclusions: This study underscores the remarkable instability of intronic regions flanking exon 11 of PALB2 and identifies a previously unreported hotspot involving Alu repeats with very high sequence homology in introns 10 and 11 of the gene. Our findings suggest avenues for further research, such as investigating the prevalence of similar genomic rearrangements in larger cohorts and exploring functional studies to understand how these alterations contribute to hereditary breast cancer pathogenesis. Full article

Papillary thyroid cancer (PTC) is one of the fastest-growing cancers worldwide, lacking established causal factors or validated early diagnostics. Human endogenous retroviruses (HERVs), comprising 8% of human genomes, have potential as PTC biomarkers due to their comparably high baseline expression in healthy thyroid tissues, indicating homeostatic roles. However, HERV regions are often overlooked in genome-wide association studies because of their highly repetitive nature, low sequence coverage, and decreased sequencing quality. Using targeted whole-genome sequence analysis in conjunction with high sequencing depth to overcome methodological limitations, we identified associations of specific HERV variants with PTC. Analyzing WGS data from 138 patients with PTC generated through The Cancer Genome Atlas project and 2015 control samples from the 1000 Genomes Project, we examined the mutational variation in HERVs within a 20 kb radius of known cancer predisposition genes (CPGs) differentially expressed in PTC. We discovered 15 common and 13 rare germline HERV variants near or within 20 CPGs that distinguish patients with PTC from healthy controls. We identified intragenic–intronic HERV variants within RYR2, LRP1B, FN1, MET, TCRVB, UNC5D, TRPM3, CNTN5, CD70, RYR1, RUNX1, CRLF2, and PCDH1X, and three variants downstream of SERPINA1 and RUNX1T1. Sanger sequencing analyses of 20 thyroid and 5 non-thyroid cancer cell lines confirmed associations with PTC, particularly for MSTA HERV-L variant rs200077102 within the FN1 gene and HERV-L MLT1A LTR variant rs78588384 within the CNTN5 gene. Variant rs78588384, in particular, was shown in our analyses to be located within a POL2 binding site regulating an alternative transcript of CNTN5. In addition, we identified 16 variants that modified the poly(A) region in Alu elements, potentially altering the potential to retrotranspose. In conclusion, this study serves as a proof-of-concept for targeted variant analysis of HERV regions and establishes a basis for further exploration of HERVs in thyroid cancer development. Full article

Background/Objectives: Alu element insertion in the exon 4 of the RP1 gene was newly identified through whole genome sequencing (WGS). This was not detected in previous next-generation sequencing (NGS) analysis. We report three cases of Korean retinitis pigmentosa (RP) patients with compound heterozygous variants including Alu element insertion in the RP1 gene, indicating that Alu element insertion could be a cause of RP; Methods: Among patients diagnosed with RP having variants in the RP1 gene in the Asan Medical Center, WGS was additionally performed for genetically unsolved cases in previous NGS analysis to detect any presence of Alu element insertion. For cases detected to have Alu element insertion in the exon 4 of the RP1 gene, genetic and clinical characteristics were analyzed; Results: Among 16 patients with RP, 3 patients were detected to have Alu element insertion in the RP1 gene. Alu element insertion in the RP1 gene was also detected using WGS. It was revealed to be a pathogenic variant. Therefore, RP1 gene mutation was the confirmed genetic cause of RP for these three cases and genetic counseling was enabled for them; Conclusions: Alu element insertion in the RP1 gene could be a genetic cause of autosomal recessive RP patients with compound heterozygous variants. Through WGS, the identification of this pathogenic variant was possible. Confirmation is needed to check the presence of Alu element insertion in patients with compound heterozygous variants in the RP1 gene. Full article

Background: It is widely accepted that the 3D chromatin organization in human cell nuclei is not random and recent investigations point towards an interactive relation of epigenetic functioning and chromatin (re-)organization. Although chromatin organization seems to be the result of self-organization of the entirety of all molecules available in the cell nucleus, a general question remains open as to what extent chromatin organization might additionally be predetermined by the DNA sequence and, if so, if there are characteristic differences that distinguish typical regions involved in dysfunction-related aberrations from normal ones, since typical DNA breakpoint regions involved in disease-related chromosome aberrations are not randomly distributed along the DNA sequence. Methods: Highly conserved k-mer patterns in intronic and intergenic regions have been reported in eukaryotic genomes. In this article, we search and analyze regions deviating from average spectra (ReDFAS) of k-mer word frequencies in the human genome. This includes all assembled regions, e.g., telomeric, centromeric, genic as well as intergenic regions. Results: A positive correlation between k-mer spectra and 3D contact frequencies, obtained exemplarily from given Hi-C datasets, has been found indicating a relation of ReDFAS to chromatin organization and interactions. We also searched and found correlations of known functional annotations, e.g., genes correlating with ReDFAS. Selected regions known to contain typical breakpoints on chromosomes 9 and 5 that are involved in cancer-related chromosomal aberrations appear to be enriched in ReDFAS. Since transposable elements like ALUs are often assigned as major players in 3D genome organization, we also studied their impact on our examples but could not find a correlation between ALU regions and breakpoints comparable to ReDFAS. Conclusions: Our findings might show that ReDFAS are associated with instable regions of the genome and regions with many chromatin contacts which is in line with current research indicating that chromatin loop anchor points lead to genomic instability. Full article

SINE-VNTR-Alu (SVA) retrotransposons can regulate expression quantitative trait loci (eQTL) of coding and noncoding genes including transposable elements (TEs) distributed throughout the human genome. Previously, we reported that expressed SVAs and human leucocyte antigen (HLA) class II genotypes on chromosome 6 were associated significantly with Parkinson’s disease (PD). Here, our aim was to follow-up our previous study and evaluate the SVA associations and their regulatory effects on the transcription of TEs within the HLA class II genomic region. We reanalyzed the transcriptome data of peripheral blood cells from the Parkinson’s Progression Markers Initiative (PPMI) for 1530 subjects for TE and gene RNAs with publicly available computing packages. Four structurally polymorphic SVAs regulate the transcription of 20 distinct clusters of 235 TE loci represented by LINES (37%), SINES (28%), LTR/ERVs (23%), and ancient transposon DNA elements (12%) that are located in close proximity to HLA genes. The transcribed TEs were mostly short length, with an average size of 389 nucleotides. The numbers, types and profiles of positive and negative regulation of TE transcription varied markedly between the four regulatory SVAs. The expressed SVA and TE RNAs in blood cells appear to be enhancer-like elements that are coordinated differentially in the regulation of HLA class II genes. Future work on the mechanisms underlying their regulation and potential impact is essential for elucidating their roles in normal cellular processes and disease pathogenesis. Full article

Retrotransposons are invasive genetic elements, which replicate by copying and pasting themselves throughout the genome in a process called retrotransposition. The most abundant retrotransposons by number in the human genome are Alu and LINE-1 elements, which comprise approximately 40% of the human genome. The ability of retrotransposons to expand and colonize eukaryotic genomes has rendered them evolutionarily successful and is responsible for creating genetic alterations leading to significant impacts on their hosts. Previous research suggested that hypomethylation of Alu and LINE-1 elements is associated with global hypomethylation and genomic instability in several types of cancer and diseases, such as neurodegenerative diseases, obesity, osteoporosis, and diabetes mellitus (DM). With the advancement of sequencing technologies and computational tools, the study of the retrotransposon’s association with physiology and diseases is becoming a hot topic among researchers. Quantifying Alu and LINE-1 methylation is thought to serve as a surrogate measurement of global DNA methylation level. Although Alu and LINE-1 hypomethylation appears to serve as a cellular senescence biomarker promoting genomic instability, there is sparse information available regarding their potential functional and biological significance in DM. This review article summarizes the current knowledge on the involvement of the main epigenetic alterations in the methylation status of Alu and LINE-1 retrotransposons and their potential role as epigenetic markers of global DNA methylation in the pathogenesis of DM. Full article

Objectives: We aimed to evaluate the DNA methylation levels in perimenopausal and postmenopausal women, measured through Long Interspersed Element-1 (LINE-1) and Alu, and the sleep parameters in relation to the presence of hot flashes (HFs). Methods: This cross-sectional study included 30 peri- or postmenopausal women aged between 45 and 55. The menopausal status was determined according to STRAW + 10 criteria and all participants had a low cardiovascular disease (CVD) risk profile determined by Framingham risk score. The sample was divided into two groups based on the presence or absence of HFs documented in their medical history during their initial visit: Group 1 (n = 15) with HFs present and Group 2 (n = 15) with HFs absent. The patients had polysomnography test and HFs were recorded both by sternal skin conductance and self-report overnight. Genomic DNA was extracted from the women’s blood and methylation status was analyzed by fluorescence-based real-time quantitative PCR. The quantified value of DNA methylation of a target gene was normalized by β-actin. The primary outcome was the variation in methylation levels of LINE-1 and Alu and sleep parameters according to the presence of HFs. Results: LINE-1 and Alu methylation levels were higher in Group 1 (HFs present), although statistically non-significant. LINE-1 methylation levels were negatively correlated with age. Sleep efficiency was statistically significantly lower for women in Group 1 (HFs present) (74.66% ± 11.16% vs. 82.63% ± 7.31%; p = 0.03). The ratio of duration of awakening to total sleep time was statistically significantly higher in Group 1 (HFs present) (22.38% ± 9.99% vs. 15.07% ± 6.93, p = 0.03). Objectively recorded hot flashes were significantly higher in Group 1 (4.00 ± 3.21 vs. 1.47 ± 1.46, p = 0.03). None of the cases in Group 2 self-reported HF despite objectively recorded HFs during the polysomnography. The rate of hot flash associated with awakening was 41.4% in the whole sample. Conclusions: Women with a history of hot flashes exhibited lower sleep efficiency and higher awakening rates. Although a history of experiencing hot flashes was associated with higher LINE-1 and Alu methylation levels, no statistical significance was found. Further studies are needed to clarify this association. This study was funded by the Scientific Research Projects Coordination Unit of Istanbul University-Cerrahpasa. Project number: TTU-2021-35629. Full article

Background: DNA methylation may be a link between HIV, aging, and the increased risk of lung comorbidities. We investigated whether bronchoalveolar lavage (BAL) cells of people living with HIV (PLWH) demonstrate epigenetic disruptions and advanced epigenetic aging. Methods: BAL cell DNA methylation from 25 PLWH and 16 HIV-uninfected individuals were tested for differential methylation of Alu and LINE-1 sites, markers of aging. We used a weighted gene correlation network analysis to identify HIV- and age-associated co-methylation networks. We tested the effect of HIV on DNA methylation using a robust linear model (false discovery rate < 0.10). Results: The BAL cells of PLWH were marked by global hypomethylation in both Alu and LINE-1 elements. Six co-methylated CpG networks were identified that were significantly associated with age; of these, the red module was significantly differentially methylated in PLWH and enriched pathways (e.g., Ras signaling and T-cell receptors). We identified 6428 CpG sites associated with HIV. Conclusions: We have shown here for the first time that alterations in the DNA methylation of BAL cells in the lung with HIV show a pattern of advanced aging. This study strongly supports that HIV may contribute to an increased the risk of lung comorbidities through the epigenetics of aging. Full article