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Nucleotide substitution in the intron 2 of HLA-DPB1*04:02:01:01 results in a novel allele, HLA-DPB1*04:02:01:44., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-A*24:617 differs from HLA-A*24:02:01:01 by one nucleotide in exon 4., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-A*02:1146 differs from HLA-A*02:01:01:01 by one nucleotide in exon 1., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

The coding sequence of HLA-B*40:540N differs from HLA-B*40:02:01:01 by a non-synonymous nucleotide substitution in exon 3., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-A*24:630 differs from HLA-A*24:20:01:01 by one nucleotide substitution in codon 131 in exon 3., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-C*17:78 differs from HLA-C*17:03:01:03 by one nucleotide change C>T in exon 3 (GCG>GTG)., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-C*07:02:151 differs from HLA-C*07:02:01:01 by one nucleotide substitution in codon 166 in exon 3., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-DRB1*08:126 differs from HLA-DRB1*08:04:01:01 by one nucleotide substitution in codon 152 in exon 3., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

The novel HLA-DPB1*14:01:15 allele differs from DPB1*14:01:01:01 by change of C > T in exon 3., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

The novel HLA-C*15:279 allele differs from HLA-C*15:02:01:01 by five nucleotide substitutions in exons 4 and 5., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Increasing evidence suggests that long noncoding RNAs (lncRNAs) play important regulatory roles in biological processes and are dysregulated in numerous tumors. The lncRNA GRASLND functions as an oncogene in many cancers, but its role in skin cutaneous melanoma (SKCM) requires further investigation., Methods: SiRNA transfection, wound - healing and transwell assays were performed to evaluate the effect of GRASLND on cellular function., Results: The present study demonstrated that GRASLND expression is increased in SKCM tissues and cell lines. The high expression of GRASLND was correlated with poor prognosis and immunotherapy outcomes. Knockdown of GRASLND significantly inhibited cell migration and invasion. In addition, we found that miR-218-5p directly binds to its binding site on GRASLND, and GRASLND and miR-218-5p demonstrate mutual inhibition. Furthermore, the miR-218-5p inhibitor partially eliminated the knockdown of GRASLND and inhibited its expression. We also demonstrated that GRASLND acts as a miR-218-5p sponge that positively regulates STAM2 expression in SKCM cells., Conclusion: In summary, these data suggest that GRASLND functions by regulating miR-218-5p/STAM2 expression, suggesting an important role for the lncRNA‒miRNA-mRNA functional network and a new potential therapeutic target for SKCM., (© 2024. The Author(s).)

The HLA-C*01:274 allele differs from HLA-C*01:02:01:01 by one nucleotide substitution in codon 240 in exon 4., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-DRB4*01:182 differs from HLA-DRB4*01:03:01:01 by one nucleotide substitution in codon 172 in exon 3., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-C*02 246 has one nucleotide change from HLA-C*02:02:02:01 at nucleotide 523 changing Arginine to Cysteine at residue 151., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-A*11:01:127 differs from HLA-A*11:01:01:01 by one nucleotide in exon 3., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-C*04:520, a novel HLA-C allele, differs from HLA-C*04:01:01 by one mismatch in exon 5., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-B*15:679 and HLA-C*15:02:01:61 differ from HLA-B*15:12:01 and HLA-C*15:02:01:01 by a single nucleotides., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-B*07:02:01:110 differs from the HLA-B*07:02:01:01 allele by two nucleotide substitutions in the 3'UTR., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Nucleotide substitution in codon 238 of HLA-C*12:02:02:01 results in a novel allele, HLA-C*12:02:53., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-C*03:620 differs from the HLA-C*03:04:01:02 allele by one nucleotide substitution in the exon 3., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-DPB1*05:01:20 differs from HLA-DPB1*05:01:01:01 by one nucleotide in exon 3., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-C*14:155 differs from HLA-C*14:02:01:01 by one nucleotide in exon 1., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Compared with HLA-DRB1*09:01:02:05, the alleles HLA-DRB1*09:57 and HLA-DRB1*09:58 each show one nucleotide change, respectively., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-C*03:652 differs from the HLA-C*03:04:01:01 by one nucleotide in exon 3., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

The novel HLA-A*02:829 allele is likely generated from the recombination of both HLA-A*02:05:01:01 and HLA-A*32:01:01:01 alleles., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

The HLA-A*24:627 allele differs from HLA-A*24:02:01:01 by one nucleotide substitution in codon 172 in exon 2., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-DRB4*01:01:12 differs from HLA-DRB4*01:01:01:01 by one nucleotide substitution in codon 175 in exon 3., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-B*15:659 differs from HLA-B*15:02:01:01 by one nucleotide in exon 2., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

The novel allele HLA-DPB1*1467:01 differs from HLA-DPB1*09:01:01:01 by one non-synonymous nucleotide substitution in exon 2., (© 2024 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-DQB1*06:465 differs from HLA-DQB1*06:04:01:01 by a non-synonymous nucleotide substitution in codon 38., (© 2023 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

One nucleotide substitution in codon 152 of HLA-DQB1*06:03:01:01 results in a novel allele, HLA-DQB1*06:474., (© 2023 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-DRB3*02:193 differs from DRB3*02:02:01:11 by one nucleotide substitution in exon 1, and intronic changes., (© 2023 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-DPB1*1461:01 differs from DPB1*02:01:02:01 in exon 2, codon 51 CTG > CGG a Leucine to Arginine replacement., (© 2023 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

HLA-DPB1*1463:01N differs from HLA-DPB1*02:01:02:04 by one nucleotide substitution in codon 128 in exon 3., (© 2023 The Authors. HLA: Immune Response Genetics published by John Wiley & Sons Ltd.)

Golden Gate Assembly is an efficient and rapid cloning method but requires dedicated vectors. Here, we modified Golden Gate to expand its compatibility to a broader range of destination vectors while maintaining its strengths. Our Expanded Golden Gate (ExGG) assembly adds to the insert(s) type IIS restriction sites that generate protruding ends compatible with traditional type IIP sites on the recipient vector. The ligated product cannot be cleaved again, owing to a single-base change near the junction. This allows the reaction to proceed in a single tube without an intermediate purification step. ExGG can be used to introduce multiple fragments into a vector simultaneously, including shorter fragments (<100 bp) and fragments with shared sequences, which can be difficult to assemble with other fast cloning strategies. Thus, ExGG extends the convenience of Golden Gate to a much larger space of pre-existing vectors designed for conventional cloning., Competing Interests: The authors have no conflict of interest to declare., (© 2023 The Authors.)

We present RBPNet, a novel deep learning method, which predicts CLIP-seq crosslink count distribution from RNA sequence at single-nucleotide resolution. By training on up to a million regions, RBPNet achieves high generalization on eCLIP, iCLIP and miCLIP assays, outperforming state-of-the-art classifiers. RBPNet performs bias correction by modeling the raw signal as a mixture of the protein-specific and background signal. Through model interrogation via Integrated Gradients, RBPNet identifies predictive sub-sequences that correspond to known and novel binding motifs and enables variant-impact scoring via in silico mutagenesis. Together, RBPNet improves imputation of protein-RNA interactions, as well as mechanistic interpretation of predictions., (© 2023. BioMed Central Ltd., part of Springer Nature.)

One nucleotide substitution in codon 140 of HLA-DRB1*04:05:01:01 results in a novel allele, HLA-DRB1*04:358., (© 2023 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Fundoscopy is the standard method for diagnosis and follow-up of intraocular retinoblastoma, but it is sometimes insufficient to discern whether tumors are inactivated following treatments. In this work, we hypothesized that the amount of conserved nuclear DNA sequences in the cell-free DNA (cfDNA) fraction of the aqueous humor (AH) might complement fundoscopy for retinoblastoma follow-up. To address our hypothesis, we developed highly sensitive droplet digital polymerase chain reaction (ddPCR) methods to quantify highly conserved DNA sequences of nucleus-encoded genes (GAPDH and B4GALNT1) and of a mitochondrial gene, MT-ATP6. We obtained AH samples during intravitreal treatments. We analyzed 42 AH samples from 25 patients with intraocular retinoblastoma and 11 AH from controls (non-cancer patients). According to clinical criteria, we grouped patients as having progression-free or progressive retinoblastoma. cfDNA concentration in the AH was similar in both retinoblastoma groups. Copy counts for nucleus-derived sequences of GAPDH and B4GALNT1 were significantly higher in the AH from patients with progressive disease, compared to the AH from progression-free patients and control non-cancer patients. The presence of mitochondrial DNA in the AH explained that both retinoblastoma groups had similar cfDNA concentration in AH. The optimal cut-off point for discriminating between progressive and progression-free retinoblastomas was 108 GAPDH copies per reaction. Among patients having serial AH samples analyzed during their intravitreal chemotherapy, GAPDH copies were high and decreased below the cut-off point in those patients responding to chemotherapy. In contrast, one non-responder patient remained with values above the cut-off during follow-up, until enucleation. We conclude that the measurement of conserved nuclear gene sequences in AH allows follow-up of intraocular retinoblastoma during intravitreal treatment. The method is applicable to all patients and could be relevant for those in which fundoscopy evaluation is inconclusive., (© 2022 The Authors. The Journal of Pathology: Clinical Research published by The Pathological Society of Great Britain and Ireland and John Wiley & Sons Ltd.)

Immobile four-way junctions (4WJs) are core structural motifs employed in the design of programmed DNA assemblies. Understanding the impact of sequence on their equilibrium structure and flexibility is important to informing the design of complex DNA architectures. While core junction sequence is known to impact the preferences for the two possible isomeric states that junctions reside in, previous investigations have not quantified these preferences based on molecular-level interactions. Here, we use all-atom molecular dynamics simulations to investigate base-pair level structure and dynamics of four-way junctions, using the canonical Seeman J1 junction as a reference. Comparison of J1 with equivalent single-crossover topologies and isolated nicked duplexes reveal conformational impact of the double-crossover motif. We additionally contrast J1 with a second junction core sequence termed J24, with equal thermodynamic preference for each isomeric configuration. Analyses of the base-pair degrees of freedom for each system, free energy calculations, and reduced-coordinate sampling of the 4WJ isomers reveal the significant impact base sequence has on local structure, isomer bias, and global junction dynamics., (© The Author(s) 2021. Published by Oxford University Press on behalf of Nucleic Acids Research.)

A number of inherited ataxias is known in humans, with more than 250 loci implicated, most of which are included in human ataxia screening panels. Anecdotally, cases of ataxia in the Norwegian elkhound black have been known for the last 40 years. Affected puppies from three litters were clinically and neurologically examined, and postmortem samples were collected for morphological studies, including ultrastructural analyses. The puppies displayed vestibulocerebellar neurological signs and had degenerative histopathological alterations in cerebellum and brain stem. Three affected dogs, each from different litters, as well as both parents and one healthy littermate from each litter, were whole genome sequenced. Through variant calling we discovered a disease-associated 1 bp deletion in HACE1 (CFA12), resulting in a frameshift at codon 333 and a premature stop codon at codon 366. The perfect association combined with the predicted significant molecular effect, strongly suggest that we have found the causative mutation for Norwegian elkhound black ataxia. We have identified a novel candidate gene for ataxia where dogs can serve as a spontaneous model for improved understanding of ataxia, also in human., Competing Interests: The authors have declared that no competing interests exist.

Multiple sequence alignment (MSA) is fundamental to many biological applications. But most classical MSA algorithms are difficult to handle large-scale multiple sequences, especially long sequences. Therefore, some recent aligners adopt an efficient divide-and-conquer strategy to divide long sequences into several short sub-sequences. Selecting the common segments (i.e. anchors) for division of sequences is very critical as it directly affects the accuracy and time cost. So, we proposed a novel algorithm, FMAlign, to improve the performance of multiple nucleotide sequence alignment. We use FM-index to extract long common segments at a low cost rather than using a space-consuming hash table. Moreover, after finding the longer optimal common segments, the sequences are divided by the longer common segments. FMAlign has been tested on virus and bacteria genome and human mitochondrial genome datasets, and compared with existing MSA methods such as MAFFT, HAlign and FAME. The experiments show that our method outperforms the existing methods in terms of running time, and has a high accuracy on long sequence sets. All the results demonstrate that our method is applicable to the large-scale nucleotide sequences in terms of sequence length and sequence number. The source code and related data are accessible in https://github.com/iliuh/FMAlign., (© The Author(s) 2021. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

We have used a variety of theoretical and experimental techniques to study the role of four basic amino acids-Arginine, Lysine, Ornithine and L-2,4-Diaminobutyric acid-on the structure, flexibility and sequence-dependent stability of DNA. We found that the presence of organic ions stabilizes the duplexes and significantly reduces the difference in stability between AT- and GC-rich duplexes with respect to the control conditions. This suggests that these amino acids, ingredients of the primordial soup during abiogenesis, could have helped to equalize the stability of AT- and GC-rich DNA oligomers, facilitating a general non-catalysed self-replication of DNA. Experiments and simulations demonstrate that organic ions have an effect that goes beyond the general electrostatic screening, involving specific interactions along the grooves of the double helix. We conclude that organic ions, largely ignored in the DNA world, should be reconsidered as crucial structural elements far from mimics of small inorganic cations., Competing Interests: The authors have declared that no competing interests exist.

We report on a 12-year-old boy with congenital thrombotic thrombocytopenic purpura, on who had an erroneous diagnosis as chronic immune thrombocytopenia. The patient presented with complaints of jaundice and skin rash. Laboratory analysis showed nonimmune hemolytic anemia and severe thrombocytopenia. Peripheral blood smear showed 8% schistocytes, polychromasia, and anisocytosis. The ADAMTS13 antigen and activity were suspected to be lower than 5% with any antibodies against the enzyme. The DNA sequence analyses resulted in compound heterozygosity consisting of c.291&#95;391del in exon 3 and c.4143dupA in exon 29. Schistocyte (fragmented erythrocytes) on the peripheral blood smear is a light that illuminates the diagnosis. Early recognition of the disease can prevent inappropriate treatments and morbidities due to organ damage., Competing Interests: The authors declare no conflict of interest., (Copyright © 2020 Wolters Kluwer Health, Inc. All rights reserved.)

Systemic lupus erythematosus (SLE) is a devastating autoimmune disorder characterized by failure of self-tolerance with resultant production of autoreactive antibodies. The etiology of this syndrome is complex, involving perturbations in immune cell signaling and development. The NZBWF1 mouse spontaneously develops a lupus-like syndrome and has been widely used as a model of SLE for over 60 years. The NZBWF1 model represents the F1 generation of a cross between New Zealand Black (NZB) and New Zealand White (NZW) mice. In order to better understand the factors that contribute to the development of autoimmunity, single cell RNA sequencing was conducted using the bone marrow from female NZBWF1 mice prior to the development of overt disease. The results were contrasted with single cell RNA sequencing results from the two parental strains. The expected findings of B cell abundance and upregulation, and evidence of interferon signaling were validated in this model. In addition, several novel areas of inquiry were identified. Most notably, the data showed a marked upregulation of the ferritin light chain across all cell types in the NZBWF1 mice compared to parental controls. This data can serve as a gene expression atlas of all hematopoietic cells in the NZBWF1 bone marrow prior to the development of autoimmunity., (© 2021. The Author(s).)

More than 900 variants have been described in the GLA gene. Some intronic variants and copy number variants in GLA can cause Fabry disease but will not be detected by classical Sanger sequence. We aimed to design and validate a method for sequencing the GLA gene using long-read Oxford Nanopore sequencing technology. Twelve Fabry patients were blindly analyzed, both by conventional Sanger sequence and by long-read sequencing of a 13 kb PCR amplicon. We used minimap2 to align the long-read data and Nanopolish and Sniffles to call variants. All the variants detected by Sanger (including a deep intronic variant) were also detected by long-read sequencing. One patient had a deletion that was not detected by Sanger sequencing but was detected by the new technology. Our long-read sequencing-based method was able to detect missense variants and an exonic deletion, with the added advantage of intronic analysis. It can be used as an efficient and cost-effective tool for screening and diagnosing Fabry disease., (© 2021. The Author(s).)

Efforts to elucidate protein-DNA interactions at the molecular level rely in part on accurate predictions of DNA-binding residues in protein sequences. While there are over a dozen computational predictors of the DNA-binding residues, they are DNA-type agnostic and significantly cross-predict residues that interact with other ligands as DNA binding. We leverage a custom-designed machine learning architecture to introduce DNAgenie, first-of-its-kind predictor of residues that interact with A-DNA, B-DNA and single-stranded DNA. DNAgenie uses a comprehensive physiochemical profile extracted from an input protein sequence and implements a two-step refinement process to provide accurate predictions and to minimize the cross-predictions. Comparative tests on an independent test dataset demonstrate that DNAgenie outperforms the current methods that we adapt to predict residue-level interactions with the three DNA types. Further analysis finds that the use of the second (refinement) step leads to a substantial reduction in the cross predictions. Empirical tests show that DNAgenie's outputs that are converted to coarse-grained protein-level predictions compare favorably against recent tools that predict which DNA-binding proteins interact with double-stranded versus single-stranded DNAs. Moreover, predictions from the sequences of the whole human proteome reveal that the results produced by DNAgenie substantially overlap with the known DNA-binding proteins while also including promising leads for several hundred previously unknown putative DNA binders. These results suggest that DNAgenie is a valuable tool for the sequence-based characterization of protein functions. The DNAgenie's webserver is available at http://biomine.cs.vcu.edu/servers/DNAgenie/., (© The Author(s) 2021. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

Circular RNAs (circRNAs) are widely expressed in highly diverged eukaryotes. Although circRNAs have been known for many years, their function remains unclear. Interaction with RNA-binding protein (RBP) to influence post-transcriptional regulation is considered to be an important pathway for circRNA function, such as acting as an oncogenic RBP sponge to inhibit cancer. In this study, we design a deep learning framework, CRPBsites, to predict the binding sites of RBPs on circRNAs. In this model, the sequences of variable-length binding sites are transformed into embedding vectors by word2vec model. Bidirectional LSTM is used to encode the embedding vectors of binding sites, and then they are fed into another LSTM decoder for decoding and classification tasks. To train and test the model, we construct four datasets that contain sequences of variable-length binding sites on circRNAs, and each set corresponds to an RBP, which is overexpressed in bladder cancer tissues. Experimental results on four datasets and comparison with other existing models show that CRPBsites has superior performance. Afterwards, we found that there were highly similar binding motifs in the four binding site datasets. Finally, we applied well-trained CRPBsites to identify the binding sites of IGF2BP1 on circCDYL, and the results proved the effectiveness of this method. In conclusion, CRPBsites is an effective prediction model for circRNA-RBP interaction site identification. We hope that CRPBsites can provide valuable guidance for experimental studies on the influence of circRNA on post-transcriptional regulation., (© The Author(s) 2021. Published by Oxford University Press. All rights reserved. For Permissions, please email: journals.permissions@oup.com.)

The application of aptamers in biomedicine is emerging as an essential technology in the field of cancer research. As small single-stranded DNA or RNA ligands with high specificity and low immunogenicity for their targets, aptamers provide many advantages in cancer therapeutics over protein-based molecules, such as antibodies. Vimentin is an intermediate filament protein that is overexpressed in endothelial cells of cancerous tissue. High expression levels of vimentin have been associated with increased capacity for migration and invasion of the tumor cells. We have selected and identified thioated aptamers with high specificity for vimentin using human ovarian cancer tissues. Tentative binding motifs were chosen for two vimentin aptamers based on predicted secondary structures. Each of these shorter, tentative binding motifs was synthesized, purified, and characterized via cell binding assays. Two vimentin binding motifs with high fidelity binding were selected and further characterized via cell and tissue binding assays, as well as flow cytometric analysis. The equilibrium binding constants of these small thioated aptamer constructs were also determined. Future applications for the vimentin binding aptamer motifs include conjugation of the aptamers to synthetic dyes for use in targeted imaging and therapy, and ultimately more detailed and precise monitoring of treatment response and tumor progression in ovarian pathology.

Transposable elements (TEs) are mobile genetic elements that make up a large fraction of mammalian genomes. While select TEs have been co-opted in host genomes to have function, the majority of these elements are epigenetically silenced by DNA methylation in somatic cells. However, some TEs in mice, including the Intracisternal A-particle (IAP) subfamily of retrotransposons, have been shown to display interindividual variation in DNA methylation. Recent work has revealed that IAP sequence differences and strain-specific KRAB zinc finger proteins (KZFPs) may influence the methylation state of these IAPs. However, the mechanisms underlying the establishment and maintenance of interindividual variability in DNA methylation still remain unclear. Here, we report that sequence content and genomic context influence the likelihood that IAPs become variably methylated. IAPs that differ from consensus IAP sequences have altered KZFP recruitment that can lead to decreased KAP1 recruitment when in proximity of constitutively expressed genes. These variably methylated loci have a high CpG density, similar to CpG islands, and can be bound by ZF-CxxC proteins, providing a potential mechanism to maintain this permissive chromatin environment and protect from DNA methylation. These observations indicate that variably methylated IAPs escape silencing through both attenuation of KZFP binding and recognition by ZF-CxxC proteins to maintain a hypomethylated state., Competing Interests: KC, AL, CT, ML, MB, JP, DS No competing interests declared, (© 2021, Costello et al.)