High GC Content Research Articles

Comparison of three commercial DNA extraction kits and assemblers for AMR determinant detection in Pseudomonas aeruginosa and Enterobacter cloacae using long-read sequencing.

More than 50 repeat expansion disorders have been identified, with long-read sequencing marking a new milestone in the diagnosis of these disorders. Despite these major achievements, the comprehensive characterization of short tandem repeats in a pathological context remains challenging, primarily due to their inherent characteristics such as motif complexity, high GC content, and variable length. In this study, our aim was to thoroughly characterize repeat expansions in two neuromuscular diseases: myotonic dystrophy type 1 (DM1) and oculopharyngodistal myopathy (OPDM) using CRISPR/Cas9-targeted long-read sequencing (Oxford Nanopore Technologies, ONT). We conducted precise analyses of the DM1 and OPDM loci, determining repeat size, repeat length distribution, expansion architecture, and DNA methylation, using three different basecalling strategies (MinKnow software, Dorado, and Bonito). We demonstrated the importance of the basecalling strategy in repeat expansion characterization. We proposed guidelines to perform CRISPR-Cas9 targeted long-read sequencing (no longer supported by ONT), from library preparation to bioinformatical analyses. Finally, we showed, for the first time, somatic mosaicism, hypermethylation of LRP12 locus in OPDM symptomatic patients and changes in the repeat tract structure of these patients. We propose a strategy based on CRISPR/Cas9-enrichment long-read sequencing for repeat expansion diseases, which can be readily applicable not only in research but also in diagnostic settings.

Abstract Background and Aims Autosomal dominant tubulointerstitial kidney disease MUC1 (ADTKD-MUC1) is a rare genetic cause of renal impairment resulting from specific frameshift variants in the coding variable number tandem repeats (VNTR) of the MUC1 gene. ADTKD-MUC1 is characterized by progressive loss of kidney function leading to ESRD, histological evidence of tubular atrophy and interstitial fibrosis and an autosomal dominant inheritance pattern. Due to its repetitive nature and high GC content, there is great difficulty in detecting pathogenic variants in MUC1 VNTR using common methods such as exome or targeted sequencing. VNtyper is a new bioinformatics tool designed to genotype MUC1 VNTR using short-read sequencing data for reliable detection of pathogenic variants in this region. In this study, we used VNtyper for analysing exome data of large cohort of Irish renal patients in order to (i) validate its performance, (ii) explore its ability to reanalyse patients with chronic kidney disease (CKD) and (iii) examine the presence of pathogenic MUC1 VNTR variants in other renal disorders such as polycystic kidney disease (PKD). Method Patients were recruited via the Inherited Kidney Disease Clinic at Beaumont Hospital, Dublin, Ireland. Detection of pathogenic variants in MUC1 VNTR was performed by processing and analysing whole exome sequencing (WES) and targeted sequencing (TS) data of the patients using the VNtyper software. The performance of VNtyper was validated by testing samples from diagnosed ADTKD-MUC1 patients (n = 6) whose pathogenic variants were identified previously by the Snapshot method. We tested a cohort of 522 non-cystic CKD patients and 366 cystic kidney disease patients through a systematic scan by VNtyper. Results The VNtyper validated the results of the six ADTKD-MUC1 patients showed that they were all heterozygous carriers of a same pathogenic frameshift insertion in MUC1 VNTR. These findings are identical to the results of the Snapshot test conducted independently for these patients and are a complete replication of them. Among the 522 CKD patients reanalysed by VNtyper, four had heterozygous pathogenic frameshift insertions in the MUC1 VNTR. Clinically, the symptoms of these patients are consistent with those of ADTKD-MUC1. Among the 366 cystsic kidney disease patients screened by the VNtyper, four were found to carry heterozygous frameshift insertions in the MUC1 VNTR. Conclusion Until now, for many renal patients, their genomic testing for pathogenic variants in the MUC1 VNTR was difficult. The results of this study strengthen the reliability of the use of VNtyper and demonstrate its great potential for reanalysing short-read sequencing data for the diagnosis of ADTKD-MUC1 patients and for exploring the possible role of MUC1 in other kidney disorders.

The mitochondrial isoform of LIG3 is proposed to catalyze both circularization of newly replicated mitochondrial DNA (mtDNA) and rejoining of free mtDNA strands in base excision and single-strand break repair. Inactivation of LIG3 has been reported to cause embryonic lethality in mice due to loss of mtDNA. Here, we applied genome editing to inactivate LIG3 in HEK 293 cells and observed only a moderate decrease of mtDNA copy numbers. BrdU incorporation experiments confirmed ongoing synthesis of intact supercoiled mtDNA. Using ultra-deep long-read sequencing of isolated mtDNA, we detected increased frequencies of single-strand and double-strand breaks clustering at sites with high GC-content, as well as hallmarks of accelerated degradation of linear mtDNA. This is likely due to the missing repair of intrinsic oxidative single-strand breaks, since the frequency of detected single-strand breaks was dependent on oxygen tension and on expression levels of enzymes involved in ROS (reactive oxygen species) defense. Exogenous oxidative challenge, that resulted in transient mtDNA damage in wild-type cells, caused dramatic mtDNA loss in LIG3−/− cell lines. Thus, our data provide evidence for the pivotal role of LIG3 in preventing mtDNA loss after oxidative damage and corroborate the hypothesis that oxidative strand break-induced mtDNA degradation is highly relevant for mtDNA turnover in vivo.

Aims: To develop cox1 based DNA barcodes for Heteropneustes fossilis and Mystus bleekeri, from Upper Lake, Bhopal, to support accurate species identification and molecular taxonomy. Study Design: The study involved the collection and morphological identification of samples followed by DNA isolation, PCR amplification, Sanger sequencing, and barcode generation. Place and Duration of Study: Laboratory of molecular Biology and Genomics, Department of Bioscience, Barkatullah University, Bhopal (M.P.), India, between March 2024 to July 2025. Methodology: The genomic DNA was extracted using Phenol:Chloroform:Isoamyl (25:24:1) method. The quality of extracted DNA was determined on a 1% agarose gel. The cox1 gene fragments were amplified using universal primer (FishF1 and FishR1). Sequenced through Sanger sequencing and analysed using BLAST, MEGA11, and BOLD systems for species identification, phylogenetic assessment and barcode development. Results: In this study, two samples were analysed to generate DNA barcodes. A 615 bases long fragment of the cox1 gene was sequenced, resulting sequences (658 bp for H. fossilis and 609 bp for M. bleekeri) were submitted to GenBank (accession number PP754237 and PX116877). Sequence composition analysis revealed a moderate AT-bias (A+T = 56.35%) with the highest GC content at the first codon position. BLAST analysis confirmed 99-100% similarity with reference sequences, validating species level identification. Phylogenetic analyses using Neighbor-Joining (NJ) trees highlighted contrasting patterns: H. fossilis sequences clustered into a single cohesive clade with low divergence across South Asia, while M. bleekeri grouped within a diverse genus level dataset showing clear interspecific separation but some overlap with congeners. Conclusion: These findings confirm the reliability of cox1 as a DNA barcode for both species. Overall, this study contributes validates barcodes for two freshwater catfishes of Upper Lake and emphasizes the value of DNA barcoding in fish taxonomy, biodiversity.

Detection of cancer at early stage can significantly improve the five-year survival rate of patients. Bisulfite-based methylation detection can cause DNA damage, especially in high GC content regions which was associated with the development of cancers. Loss of aberrant methylated CpG sites in cfDNA will lead to the undetectability of certain circulating tumor DNA (ctDNA), consequently may affect the cancer detection. Our study uses enzymatic method to detect whole genome abnormal methylation regions in esophageal squamous cell carcinoma (ESCC). We also provide a pretrained neural network, hybrid of BERT and CNN (BCNN), to identify ctDNA robustly. Maximum posterior probability is utilized to estimate the fraction of ESCC-derived ctDNA in plasma for predicting the risk of ESCC cancer. Our results analysis indicated that enzyme-based whole-genome methylation sequencing retained more longer cfDNA and detected more CpG sites than bisulfite-based method in both gDNA and cfDNA. Enrichment analysis of differentially methylated regions (DMR) showed that top five pathways were associated with ESCC. Compared to traditional models, our BCNN demonstrates the best performance in identifying ctDNA (AUC = 0.970). By estimating the fraction of plasma ctDNA, our BCNN exhibits high accuracy in ESCC detection even at ultralow sequencing depths (AUC = 0.946). Specifically, in the validation cohort, when the specificity is 93.75%, 7 out of 8 early-stage ESCC (TNM Stage I) were identified as positive in our preliminary results. In conclusion, our preliminary results reinforce the idea of employing BCNN as a novel strategy for ESCC early detection in clinical practice. However, to be applied in clinical, further validation with a larger sample size is necessary.Supplementary InformationThe online version contains supplementary material available at 10.1038/s41598-025-18278-2.

IntroductionPsoriasis is a chronic autoimmune skin disorder with a complex genetic basis. However, the codon usage patterns and nucleotide features of psoriasis-related genes remain unexplored, despite their potential to influence gene expression and disease progression.MethodsWe analyzed 79 psoriasis-associated genes to investigate codon usage bias (CUB) and nucleotide composition. Metrics included GC content, effective number of codons (ENC), and relative synonymous codon usage (RSCU). Evolutionary influences were assessed using correspondence analysis, parity rule 2 (PR2) plots, and neutrality plots.ResultsFunctional enrichment analysis identified pathway involvement. Comparative genomic analysis evaluated differences in coding sequence and UTR lengths and GC content relative to the genome-wide background. Psoriasis-related genes showed high GC content (mean = 53.3 ± 9.3%) with a strong preference for GC-ending codons, especially at the third codon position (GC3 = 60.6 ± 16.1%). RSCU analysis revealed frequent use of GCC (alanine), CTG (leucine), and GTG (valine). While the mean ENC (46.2 ± 9.9) suggested moderate codon bias, several genes displayed strong bias (ENC < 30). Selection pressure accounted for 71% of codon usage variation, with mutation pressure contributing 29%. Functional enrichment showed significant involvement in IL-17 (FDR = 3.4×10−3), JAK-STAT (FDR = 3.4×10−3), and TNF (FDR = 8.0×10−³) signaling pathways. These genes also tended to have shorter coding sequences and 5′UTRs and higher GC content compared to genome-wide averages.ConclusionIn conclusion, this study reveals that psoriasis-related genes are under strong selective pressure, enriched in key inflammatory pathways, and exhibit codon and nucleotide features that may optimize expression in inflamed tissues. These insights have translational relevance for designing codon-optimized mRNAs, gene therapies, and diagnostic tools tailored to autoimmune diseases like psoriasis.

Orf virus (ORFV), the prototype species of the Parapoxvirus genus within the Poxviridae family (subfamily Chordopoxvirinae), is a global pathogen infecting sheep, goats, and other ruminants, with zoonotic potential for humans. In this study, an outbreak of ORFV infection occurred in a sheep flock in Changling County, Jilin province, China, causing papules, pustules, and crusting lesions on the lips and eyelids. Typical parapoxvirus particles were observed using electron microscopy, and a wild ORFV strain was isolated, characterized, and designated as ORFV-CL24. To clarify the epidemiological and genomic characteristics of ORFV in the region, we completed its whole-genome sequencing (GenBank accession number: PV126639). Genome analysis revealed that ORFV-CL24 shares a conserved structure with other isolates available in GenBank, which possess a complete genomic sequence of 138,500 bp of dsDNA harboring 131 putative open reading frames (ORFs) flanked by inverted terminal repeat (ITR) regions of 3,264 bp at both termini. Additionally, the genome exhibited high GC-content (63.3%), indicating its key role in DNA stability. Phylogenetic analysis placed the wild strain within a subclade with the attenuated ORFV strain D1701, implying a putative common ancestor or epidemiological linkages. Further analysis of B2L (ORFV 011) and E3L (ORFV 020) genes further revealed genetic diversity and evolutionary patterns. Notably, despite phylogenetic relatedness, specific mutations in ORF020 further distinguished ORFV-CL24 from D1701, reflecting stepwise mutation accumulation during host adaptation. In conclusion, our results provided valuable genetic insights into ORFV-CL24, which contributed to a better understanding of its evolution, biological properties, and endemic trends in China.

The Proline-Glutamate/Proline-Proline-Glutamate (PE/PPE) gene family comprises approximately 10% of the Mycobacterium tuberculosis (Mtb) genome and is characterized by GC-rich, highly repetitive sequences. As a result, these genes are usually excluded from short-read-based whole-genome sequencing analyses, leaving their sequence diversity and evolutionary dynamics poorly characterized. Recently, a genome masking approach demonstrated that roughly 54% of PE/PPE sequences are recoverable from short-read data, providing an opportunity to examine the evolution of this gene family at a population level. Here, we analyzed 51,229 Mtb genomes to characterize sequence diversity and selection pressures across the PE/PPE gene family. Overall, we observed that PE/PPE genes are under relaxed purifying selection compared to other gene categories, as evidenced by higher ratios of nonsynonymous to synonymous polymorphisms (pNpS) and greater mutation burdens. We identified 12 PE/PPE genes with signatures of positive selection and 7 with selective pressure associated with antibiotic resistance. Among these genes, PPE51 exhibited selection favoring loss-of-function mutations, which occurred only in Mtb strains that were already multidrug-resistant (MDR). This pattern suggests either compensatory evolution or adaptation related to resistance against second-line or newly introduced drugs. Additionally, we identified T-cell epitopes in six PE/PPE genes that were subject to diversifying selection, suggesting immune-driven adaptation. Collectively, this work provides a baseline characterization of genetic diversity in PE/PPE genes and highlights specific genes that may be involved in adaptation to host immunity and antibiotic pressure and represent candidates for further investigation.IMPORTANCETuberculosis remains a significant global health challenge, partly due to Mycobacterium tuberculosis (Mtb)'s remarkable evolutionary adaptation to antibiotics and human immune responses. Around 10% of its genome comprises PE/PPE genes, whose functions and evolutionary dynamics are poorly understood due to their repetitive sequences and high GC content. In this study, we analyzed 51,229 global Mtb genomes using an advanced genome-masking method, revealing numerous PE/PPE genes under positive selection, potentially facilitating antibiotic resistance and immune evasion. Notably, PPE51 often loses its function in strains resistant to multiple antibiotics, suggesting a role in bacterial survival during drug treatment. Additionally, we identified mutation-prone regions within six PE/PPE genes, highlighting potential targets for future vaccine development. Collectively, our findings underscore the crucial role of PE/PPE genes in Mtb evolution and drug resistance, providing valuable insights to inform novel therapeutic and vaccine strategies.

RNA editing sites in coding sequences of ABO blood group alleles.

BackgroundADTKD-MUC1 is caused by frameshift mutations in MUC1 gene that produce a frameshifted protein (MUC1fs) toxic to kidney cells. The gene’s variable number of tandem repeats (VNTR), with high GC content, makes it largely inaccessible to standard sequencing. As a result, both the reference sequence and natural variation in this region remain poorly defined, complicating mutation detection and data interpretation. Standard methods also fail to pinpoint the exact VNTR unit affected, limiting insight into mutation mechanisms and genotype–phenotype correlations.MethodsWe employed Single Molecule, Real-Time (SMRT) sequencing and characterized the genomic sequence of MUC1 in 300 individuals including 279 individuals from 143 families suspected of having ADTKD-MUC1. We compared these results to those obtained using the CLIA-approved mass spectrometry-based probe extension (PE) assay, which specifically detect the most prevalent 59dupC mutation. We correlated the structural features of the MUC1 VNTR with the rate of kidney function decline in affected individuals.ResultsWe identified MUC1 consensus sequences for 205 unique VNTR alleles, with 9 distinct types of frameshift mutations present on 52 distinct mutated VNTR alleles. MUC1 frameshift mutations were identified in 71 of 143 families (50%) with suspected ADTKD, comprising 135 genetically affected individuals (48%). The SMRT assay exhibited complete concordance and revealed that the PE assay is capable of detecting frameshift mutations in approximately 85% of affected families. The constellation of VNTR structures supports a genotype–progression model, in which fast progressors exhibit a significantly lower number of repeat units on the wild-type allele and a higher number of repeats on the mutation-bearing allele, including an increased number of frameshifted repeat units.ConclusionsSMRT sequencing outperforms current diagnostic methods for ADTKD-MUC1 and reveals the prognostic value of VNTR structures. Although their contribution to disease progression is modest (~6% variance explained), it remains biologically and clinically meaningful.

Marginal seas are increasingly impacted by anthropogenic activities, leading to widespread eutrophication, yet the responses of marine microbial communities remain poorly understood. We compared sediments from the highly eutrophic Yangtze River Estuary (YRE) and the oligotrophic East China Sea (ECS) to examine how eutrophication alters microbial abundance, community structure, assembly processes, functional profiles, and life-history strategies. Our results showed that YRE sediments harbored significantly higher microbial abundance (1.3 × 108-1.1 × 109 cells g−1 vs. 8.0 × 107-7.1 × 108 cells g−1), Chao1 richness (9,782–18,129 vs. 9,366–14,903), and Shannon diversity (6.19–7.47 vs. 6.05–7.07). Functional profiling revealed an enrichment of nitrogen- and carbon-cycling genes, human pathogens, and antibiotic-resistance genes in YRE. Life-history traits in YRE microbial communities showed higher average 16S rRNA gene copy numbers (median 2.75 vs. 2.56), greater codon usage bias (0.0181 vs. 0.0178), higher maximum predicted growth rates (0.1054 vs. 0.0951 h−1), larger genome sizes (5.59 vs. 5.46 Mb), higher GC content (56.43 vs. 55.83%) and increased transposase abundance (3.46 vs. 1.71%), collectively indicating a shift from K-strategists to r-strategists in the eutrophic environment. Neutral and null model analyses, and statistical analyses revealed that human activities, especially those altering water quality and chemistry, drive significant shifts in microbial community structure, function, and assembly processes, which in turn reshape microbial life-history strategies in estuarine benthic ecosystems.

The emergence of beta-lactamase producing multidrug-resistant (MDR) gram-negative bacteria presents a significant challenge to effective treatment of infections. This study focuses on the isolation, amplification, and molecular characterization of β-lactamase genes from clinical strains of Escherichia coli and Klebsiella pneumoniae. Seven new partial gene sequences, including novel variants of blaOXA and blaNDM, were identified after screening 108 clinical samples and submitted to NCBI GenBank. In silico analysis revealed considerable diversity and distribution of these resistance genes among different strains of bacteria. Gene structure predictions using GENSCAN showed that blaOXA genes typically contain single exons with moderate GC content, whereas blaNDM genes feature longer exons with higher GC content. Multiple sequence alignment showed that NDM and OXA β-lactamases were highly similar, with only slight differences in a few amino acids. The study also analyzed the physico-chemical properties, functional domains, and phosphorylation patterns of the β-lactamase proteins. Secondary structure prediction indicated a dominance of beta sheets, contributing to protein stability, while tertiary modeling provided insights into their 3D structure. Overall, these findings provide critical insights into the genetic diversity and potential mechanisms of β-lactamase-mediated resistance, offering valuable information for the development of novel therapeutic strategies and surveillance programs.

Paramecium, a group of ciliates with a long evolutionary history, plays essential roles in freshwater ecosystems and has been model for genetic, cellular, and evolutionary studies for over a century. Despite the valuable contributions of genomic resources such as ParameciumDB, genomic data are still mostly limited to species in and near the P. aurelia group. This study addresses this gap by HiFi sequencing, assembling, and annotating the macronuclear genomes of five rare Paramecium species: P. calkinsi, P. duboscqui, P. nephridiatum, P. putrinum, and P. woodruffi. These genomes enable a comprehensive exploration of genomic diversity, genome evolution, and phylogenomic relationships within the genus Paramecium. The genome sizes range from 47.78 to 113.16 Mb, reflecting unexpected variation in genomic content, and genic features differ from those of other reported Paramecium genomes, such as larger intron sizes and higher GC content. Nonetheless, the de novo assemblies indicate that macronuclear genomes of all Paramecium are highly streamlined, with ~77% being protein-coding gene regions. Based on gene-duplication depths, synonymous mutations in paralogs, and phylogenomic relationships, we discovered that the five species experienced at least three whole-genome duplication (WGD) events, independent of those previously found in the P. aurelia complex. Using all available WGD data for Paramecium, we further explore the paralog dynamics after WGD events by modeling. This study contributed to a more comprehensive and deeper understanding of genome architecture and evolution in Paramecium.

One of the characteristics of actinomycetes, especially streptomycetes, is the high GC content in their genome, which often leads to the failure of heterologous expression in E. coli, and thus hinders in vitro enzyme activity experiments. Therefore, we have developed a precisely regulated and efficient Streptomyces expression system, pTZYp, that relied on the strong promoter stnY and the cmt operon. As tested in three model Streptomyces strains (S. albus J1074, S. coelicolor M1152 and S. lividans TK24), the reporter protein sfGFP was not detected without the addition of the inducer cumate, whereas sfGFP was significantly produced when a certain amount of inducer cumate was added to the medium, demonstrating that the pTZYp expression system can achieve the goal of precise regulation and efficient expression. After optimization of the expression conditions, the maximum sfGFP production was obtained when the inducer was added to the final concentration of 100μM and cultivated for about 24h. pTZYp has also been used to express other six non-model proteins in Streptomyces, and all of them have been successfully expressed. The pTZYp expression system demonstrated robustness, high efficiency (relying on the stnY promotor), precise regulation (relying on cmt operon and moderate production of regulatory protein CymR) and low experimental cost (relying on the lower cost of the inducer cumate), which may be an efficient and widely applicable heterologous expression tool for genes with high GC content in actinomycetes.

Fully integrated centrifugal microfluidic platform for rapid HLA-B∗58:01 allele identification using duplex RPA assay.

Methods for fast and inexpensive gene synthesis from oligonucleotide pools enable rapid iteration of genetic designs. Here, we describe iggypop (indexed Golden Gate gene assembly from PCR-amplified oligonucleotide pools), a simple computational-experimental pipeline that allows for low-cost design and synthesis of hundreds of genes from oligonucleotide pools using Golden Gate assembly methods. We used iggypop to synthesize a series of single-transcript autonomously bioluminescent reporters (STARBURSTs) that link the five genes of a fungal bioluminescence pathway via ribosomal skipping LP4/2A sequences into a 9.5 kb transcript that function in planta. We also synthesized RUBY reporters (a reporter gene system producing red betalain pigment) recoded to match dicot codon usage, as RUBY was codon optimized for rice codon usage and has a high GC content. Surprisingly, the recoded RUBYs substantially reduced betalain production in transient Nicotiana benthamiana assays. Based on this observation, we synthesized six GC-boosted STARBURSTs, which produced robust luminescence in both transient assays and transgenic Arabidopsis plants. Thus, iggypop enabled the rapid synthesis of multiple genetic designs to deliver a bright single transcript autobioluminescent reporter. Iggypop should enable the facile synthesis and optimization of new genetic parts and complex polycistronic pathways.

Orphan genes (OGs) lack homologs in related species and have been associated with adaptive evolution. However, it is poorly characterized in Brassica napus (rapeseed). This study aims to identify and characterize OGs in rapeseed to evaluate their association with stress adaptation and lineage-specific traits. Through comprehensive comparative genomics analysis, all rapeseed genes were categorized into four distinct evolutionary classes. Furthermore, bioinformatics analyses were carried out to evaluate the structural, evolutionary, and expression dynamics, which were further validated by qRT-PCR analysis of different tissues and in cold stress. In total, 4 B. napus OGs (BnaOGs), 2859 Brassica-specific genes (BSGs), 9650 Cruciferae-specific genes (CSGs), and 94,720 evolutionarily conserved genes (ECGs) were identified. BnaOGs and BSGs indicated shorter sequences, higher GC content, fewer transcription factors, and limited functional annotation compared to ECGs. Similarly, transcriptomic analysis determined the tissue-specific and stress-responsive expression patterns in BnaOGs and BSGs. qRT-PCR validation revealed four BnaOGs and five BSGs from different tissue-specific and cold-responsive expression modules in rapeseed. Overall, this study identified OGs associated with lineage-specific adaptation in rapeseed, potentially related to cold tolerance and phenotypic diversity. The identified expression patterns and structural divergence provide novel insights for breeding stress-resilient varieties.

Nuclear export of mRNAs in the form of messenger ribonucleoprotein particles (mRNPs) is an obligatory step for eukaryotic gene expression. The DEAD-box ATPase DDX39B (also known as UAP56) is a multifunctional regulator of nuclear mRNPs. How DDX39B mediates mRNP assembly and export in a controlled manner remains elusive. Here, we identify a novel complex TREX-2.1 localized in the nucleus that facilitates the release of DDX39B from the mRNP. TREX-2.1 is composed of three subunits, LENG8, PCID2, and DSS1, and shares the latter two subunits with the nuclear pore complex-associated TREX-2 complex. Cryo-EM structures of TREX-2.1/DDX39B and TREX-2/DDX39B identify a conserved trigger loop in the LENG8 and GANP subunit of the respective TREX-2.1 and TREX-2 complex that is critical for DDX39B regulation. RNA sequencing from LENG8 knockdown cells shows that LENG8 influences the nucleocytoplasmic ratio of a subset of mRNAs with high GC content. Together, our findings lead to a mechanistic understanding of the functional cycle of DDX39B and its regulation by TREX-2 and TREX-2.1 in mRNP processing.

Meiotic recombination is essential for chromosomal segregation and facilitates the exchange between homologs, which leads to the transmission of new combinations of linked alleles to the progeny. The eukaryotic meiotic machinery is generally highly conserved, but the frequency of crossover occurrence can vary dramatically across species and populations, between individuals, and across sexes. The chicken and the guinea fowl exhibit interspecific variation in the distribution of crossovers along their largest chromosomes. In many organisms, an association has been observed between the preferred crossover location and certain sequence parameters, such as high GC content, CpG islands, or gene promoters. Here, we compared the distribution of these genomic parameters with the recombination landscape, represented by MLH1 focus frequencies, in the two birds. We found an association between GC content density and recombination in the chicken, but the remaining parameters showed weak or no association with recombination, especially in the guinea fowl. We conclude that despite the different broad-scale crossover distribution, the investigated genomic parameters remained remarkably similar in these two species. We suggest that the density of these genomic features is more likely related to microscale variations in recombination rates, such as those determined by open chromatin configurations.