Terminal Repeat Research Articles

Non-canonical imprinting, manifesting as post-fertilization placenta-specific parent-of-origin dependent methylation, is not conserved in humans.

Genomic imprinting is the parent-of-origin dependent monoallelic expression of genes often associated with regions of germline-derived DNA methylation that are maintained as differentially methylated regions (gDMRs) in somatic tissues. This form of epigenetic regulation is highly conserved in mammals and is thought to have co-evolved with placentation. Tissue-specific gDMRs have been identified in human placenta, suggesting that species-specific imprinting dependent on unorthodox epigenetic establishment or maintenance may be more widespread than previously anticipated. Non-canonical imprinting, reliant on differential allelic H3K27me3 enrichment, has been reported in mouse and rat pre-implantation embryos, often overlapping long terminal repeat (LTR)-derived promoters. These non-canonical imprints lose parental allele-specific H3K27me3 specificity, subsequently gaining DNA methylation on the same allele in extra-embryonic tissues resulting in placenta-specific, somatically acquired maternal DMRs. To determine if similar non-canonical imprinting is present in the human placenta, we interrogated allelic DNA methylation for a selected number of loci, including (i) the human orthologues of non-canonical imprinted regions in mouse and rat, (ii) promoters of human LTR-derived transcripts, and (iii) CpG islands with intermediate placenta-specific methylation that are unmethylated in gametes and pre-implantation embryos. We failed to identify any non-canonical imprints in the human placenta whole villi samples. Furthermore, the assayed genes were shown to be biallelically expressed in human pre-implantation embryos, indicating they are not imprinted at earlier time points. Together, our work reiterates the continued evolution of placenta-specific imprinting in mammals, which we suggest is linked to epigenetic differences during the maternal-to-embryo transition and species-specific integration of retrotransposable elements.

Reactivation of latent HIV-1 by the glucocorticoid receptor modulator AZD9567.

One key determinant of HIV-1 latency reversal is the activation of the viral long terminal repeat (LTR) by cellular transcription factors such as NF-κB and AP-1. Interestingly, the activity of these two transcription factors can be modulated by glucocorticoid receptors (GRs). Furthermore, the HIV-1 genome contains multiple binding sites for GRs. We therefore hypothesized that glucocorticoids and other GR modulators may influence HIV-1 latency and reactivation. To investigate how GR signaling affects latent HIV-1 reservoirs, we assembled a representative panel of GR modulators including natural steroidal agonists, selective and non-selective GR modulators, and clinically approved GR-modulating drugs. The effects of these compounds on HIV-1 reactivation were assessed using latently HIV-1-infected cell lines and primary cells, as well as reporter assays that monitored GR and LTR activities. We found that AZD9567 (Mizacorat), a non-steroidal partial GR agonist, reactivates latent HIV-1 in both lymphoid and myeloid cell lines and primary CD4+ T cells. Conversely, the GR antagonist mifepristone suppresses HIV-1 LTR-driven gene expression. Mechanistic analyses revealed that AZD9567-mediated reactivation partially depends on both GR and AP-1 binding sites in the LTR. In summary, we, here, identify the GR modulator AZD9567 as novel latency-reversing agent that activates LTR-driven gene expression, which may aid in advancing current shock-and-kill approaches in the treatment of HIV-1 infection.IMPORTANCELatently infected cells of people living with HIV are constantly exposed to fluctuating levels of glucocorticoid hormones such as cortisol. In addition, many HIV-infected individuals regularly take corticosteroids as anti-inflammatory drugs. Although corticosteroids are known to affect the activity of the viral long terminal repeat (LTR) promoter and influence ongoing HIV-1 replication, relatively little is known about the effect of corticosteroid hormones and other glucocorticoid receptor (GR) modulators on latent HIV-1. By systematically comparing natural and synthetic GR modulators, we, here, identify a first first-in-class, oral, partial GR agonist that reactivates latent HIV-1 from different cell types. This drug, AZD9567, was previously tested in clinical trials for rheumatoid arthritis. Mutational analyses shed light on the underlying mode of action and revealed transcription factor binding sites in the HIV-1 LTR that determine responsiveness to AZD9567.

Expanding the Diversity of Actinobacterial Tectiviridae: A Novel Genus from Microbacterium

Six novel Microbacterium phages belonging to the Tectiviridae family were isolated using Microbacterium testaceum as a host. Phages MuffinTheCat, Badulia, DesireeRose, Bee17, SCoupsA, and LuzDeMundo were purified from environmental samples by students participating in the Science Education Alliance Phage Hunters Advancing Genomics and Evolutionary Science (SEA-PHAGES) program at Alliance University, New York. The phages have linear dsDNA genomes 15,438–15,636 bp with 112–120 bp inverted terminal repeats. Transmission electron microscopy (TEM) imaging analysis revealed that the six novel phages have six-sided icosahedral double-layered capsids with an internal lipid membrane that occasionally forms protruding nanotubules. Annotation analysis determined that the novel Microbacterium phages all have 32–34 protein-coding genes and no tRNAs. Like other Tectiviridae, the phage genomes are arranged into two segments and include three highly conserved family genes that encode a DNA polymerase, double jelly-roll major capsid protein, and packaging ATPase. Although the novel bacteriophages have 91.6 to 97.5% nucleotide sequence similarity to each other, they are at most 58% similar to previously characterized Tectiviridae genera. Consequently, these novel Microbacterium phages expand the diversity of the Tectiviridae family, and we propose they form the sixth genus, Zetatectivirus.

Graph pangenome reveals the regulation of malate content in blood-fleshed peach by NAC transcription factors

BackgroundFruit acidity and color are important quality attributes in peaches. Although there are some exceptions, blood-fleshed peaches typically have a sour taste. However, little is known about the genetic variations linking organic acid and color regulation in peaches.ResultsHere, we report a peach graph-based pangenome constructed from sixteen individual genome assemblies, capturing abundant structural variations and 82.3 Mb of sequences absent in the reference genome. Pangenome analysis reveals a long terminal repeat retrotransposon insertion in the promoter of the NAC transcription factor (TF) PpBL in blood-fleshed peaches, which enhances PpBL expression. Genome-wide association study identifies a significant association between PpBL and malate content. Silencing PpBL in peach fruit and ectopic overexpression of PpBL in tomatoes confirm that PpBL is a positive regulator of malate accumulation. Furthermore, we demonstrate that PpBL works synergistically with another NAC TF, PpNAC1, to activate the transcription of the aluminum-activated malate transporter PpALMT4, leading to increased malate content.ConclusionsThese findings, along with previous research showing that PpBL and PpNAC1 also regulate anthocyanin accumulation, explain the red coloration and sour taste in blood-fleshed peach fruits.

Chromosome-Level Genome Assembly of Five Emberiza Species Reveals the Genomic Characteristics and Intrinsic Drivers of Adaptive Radiation.

Emberiza buntings (Aves: Emberizidae) exhibit extensive diversity and rapid diversification within the Old World, particularly in the eastern Palearctic, making them valuable models for studying rapid radiation among sympatric species. Despite their ecological and morphological diversity, there remains a significant gap in understanding the genomic underpinnings driving their rapid speciation. To fill this gap, we assembled high-quality chromosome-level genomes of five representative Emberiza species (E. aureola, E. pusilla, E. rustica, E. rutila and E. spodocephala). Comparative genomic analysis revealed distinct migration-related evolutionary adaptations in their genomes, including variations in lipid metabolism, oxidative stress response, locomotor ability and circadian regulation. These changes may facilitate the rapid occupation of emerging ecological niches and provide opportunities for species diversification. Additionally, these five species exhibited abnormal abundances of long terminal repeat retrotransposons (LTRs), comprising over 20% of their genomes, with insertion times corresponding to their divergence (~2.5 million years ago). The presence of LTRs influenced genome size, chromosomal structure and single-gene expression, suggesting their role in promoting the rapid diversification of Emberiza species. These findings offer valuable insights into the adaptive radiation of Emberiza and establish a robust theoretical foundation for further exploration of the patterns and mechanisms underlying their diversification.

Centrophilic retrotransposon integration via CENH3 chromatin in Arabidopsis

In organisms ranging from vertebrates to plants, major components of centromeres are rapidly evolving repeat sequences, such as tandem repeats (TRs) and transposable elements (TEs), which harbour centromere-specific histone H3 (CENH3)1,2. Complete centromere structures recently determined in human and Arabidopsis suggest frequent integration and purging of retrotransposons within the TR regions of centromeres3, 4–5. Despite the high impact of ‘centrophilic’ retrotransposons on the paradox of rapid centromere evolution, the mechanisms involved in centromere targeting remain poorly understood in any organism. Here we show that both Ty3 and Ty1 long terminal repeat retrotransposons rapidly turnover within the centromeric TRs of Arabidopsis species. We demonstrate that the Ty1/Copia element Tal1 (Transposon of Arabidopsis lyrata 1) integrates de novo into regions occupied by CENH3 in Arabidopsisthaliana, and that ectopic expansion of the CENH3 region results in spread of Tal1 integration regions. The integration spectra of chimeric TEs reveal the key structural variations responsible for contrasting chromatin-targeting specificities to centromeres versus gene-rich regions, which have recurrently converted during the evolution of these TEs. Our findings show the impact of centromeric chromatin on TE-mediated rapid centromere evolution, with relevance across eukaryotic genomes.

Correction: Detection and classification of long terminal repeat sequences in plant LTR-retrotransposons and their analysis using explainable machine learning

Correction: Detection and classification of long terminal repeat sequences in plant LTR-retrotransposons and their analysis using explainable machine learning

Detection and classification of long terminal repeat sequences in plant LTR-retrotransposons and their analysis using explainable machine learning

BackgroundLong terminal repeats (LTRs) represent important parts of LTR retrotransposons and retroviruses found in high copy numbers in a majority of eukaryotic genomes. LTRs contain regulatory sequences essential for the life cycle of the retrotransposon. Previous experimental and sequence studies have provided only limited information about LTR structure and composition, mostly from model systems. To enhance our understanding of these key sequence modules, we focused on the contrasts between LTRs of various retrotransposon families and other genomic regions. Furthermore, this approach can be utilized for the classification and prediction of LTRs.ResultsWe used machine learning methods suitable for DNA sequence classification and applied them to a large dataset of plant LTR retrotransposon sequences. We trained three machine learning models using (i) traditional model ensembles (Gradient Boosting), (ii) hybrid convolutional/long and short memory network models, and (iii) a DNA pre-trained transformer-based model using k-mer sequence representation. All three approaches were successful in classifying and isolating LTRs in this data, as well as providing valuable insights into LTR sequence composition. The best classification (expressed as F1 score) achieved for LTR detection was 0.85 using the hybrid network model. The most accurate classification task was superfamily classification (F1=0.89) while the least accurate was family classification (F1=0.74). The trained models were subjected to explainability analysis. Positional analysis identified a mixture of interesting features, many of which had a preferred absolute position within the LTR and/or were biologically relevant, such as a centrally positioned TATA-box regulatory sequence, and TG..CA nucleotide patterns around both LTR edges.ConclusionsOur results show that the models used here recognized biologically relevant motifs, such as core promoter elements in the LTR detection task, and a development and stress-related subclass of transcription factor binding sites in the family classification task. Explainability analysis also highlighted the importance of 5’- and 3’- edges in LTR identity and revealed need to analyze more than just dinucleotides at these ends. Our work shows the applicability of machine learning models to regulatory sequence analysis and classification, and demonstrates the important role of the identified motifs in LTR detection.

Tannic acid reactivates HIV-1 latency by mediating CBX4 degradation.

HIV-1 can integrate viral DNA into host cell chromosomes and establish a long-term stable latent viral reservoir, a major obstacle in curing HIV-1 infection. The reactivation of latent proviruses with latency-reversing agents (LRAs) is a prerequisite for the eradication of viral reservoirs. Previous reports have shown that tannic acid (TA) exerts several biological functions, including antioxidant and antitumor activities. Here, we identified a novel function of TA as a reactivator of HIV-1 latency. TA showed similar features to the HIV-1 transactivator of transcription (Tat) and was able to reactivate a larger number of proviruses from various integration sites. TA also showed a strong synergistic effect with other LRAs acting on different signaling pathways. Further studies revealed that the polycomb repressive complex 1 component, chromobox protein homolog 4 (CBX4), is specifically degraded by TA through ubiquitination. CBX4 is associated with the tri-methylation at lysine 27 of histone H3 (H3K27me3) which was enriched on HIV-1 long terminal repeat regions. The TA-induced CBX4 degradation decreased the H3K27me3 enrichment and subsequently enhanced the transcriptional activity of the integrated proviruses. These results suggest that TA is an efficient LRA aiming to a new target for HIV-1 latency, which could be developed to eradicate latent proviruses.IMPORTANCEHIV-1 remains a global health challenge, with its ability to integrate into the host genome and evade the effects of drugs. To overcome this obstacle, the "shock and kill" strategy was proposed, targeting the reactivation of latent HIV-1 for subsequent eradication through antiretroviral medication and immune system reinforcement. Here, we found a new reactivator for HIV-1 latency, tannic acid (TA), which can reactivate HIV-1 latency widely and deeply. Moreover, we demonstrated that TA could promote the interaction between the polycomb repressive complex 1 component CBX4 and the E3 ubiquitin ligase cullin 4A (CUL4A), resulting in CBX4 degradation through the ubiquitin-proteasome system. These events reduce H3K27me3 enrichment in the HIV-1 long terminal repeat region, thereby promoting HIV-1 transcription and ultimately reactivating HIV-1 latent infection. Our work may facilitate the identification of new latency-reversing agents and provide more theoretical evidence for the molecular mechanism of HIV-1 latency.

Repeat competition and ecological shifts drive the evolution of the mobilome in Rhynchospora Vahl. (Cyperaceae), the holocentric beaksedges.

Genomic changes triggered by polyploidy, chromosomal rearrangements, and/ or environmental stress are among factors that affect the activity of mobile elements, particularly Long Terminal Repeats Retrotransposons (LTR-RTs) and DNA transposons. Because these elements can proliferate and move throughout host genomes, altering the genetic, epigenetic and nucleotypic landscape, they have been recognized as a relevant evolutionary force. Beaksedges (Rhynchospora) stand out for their wide cosmopolitan distribution, high diversity (~400 spp.) and holocentric chromosomes related to high karyotypic diversity and a centromere-specific satDNA Tyba. This makes the genus an interesting model to investigate the interactions between repetitive elements, phylogenetic relationships, and ecological variables. Here, we used comparative phylogenetic methods to investigate the forces driving the evolution of the entire set of mobile elements (mobilome) in the holocentric genus Rhynchospora. We statistically tested the impact of phylogenetic relationships, abundance of holocentromeric satDNA Tyba, diversity of repeatome composition, ecological variables, and chromosome number in mobile element diversification. Tyba abundance was found to be inversely correlated with LTR-RT content. Decrease of LTR abundance and diversity was also related to increase in chromosome number (likely due to fission events), and colonization of dry environments in the northern hemisphere. In contrast, we found constant LTR insertions throughout time in species with lower chromosome numbers in rainier environments in South America. A multivariate model showed that different traits drive LTR abundance, especially repeat diversity and Tyba abundance. Other mobile elements, such as non-LTR RTs and DNA transposons had insufficient abundance to be included in our models. Our findings suggest that LTR evolution is strongly impacted by the holocentric characteristics of Rhynchospora chromosomes, correlating with species diversification and biome shifts, and supporting a holokinetic drive model of evolution and a competitive scenario with Tyba. Altogether, our results present evidence of multi-trait influence on LTR-RT dynamics and provide a broader understanding of TE evolution in a macroevolutionary context.

Chromosome-level assemblies of the endemic Korean species Abeliophyllum distichum and Forsythia ovata

Abeliophyllum distichum and Forsythia ovata are closely related species endemic to Korea and are highly valued as ornamental shrubs in the Oleaceae family. A combination of PacBio and Illumina sequencing with Hi-C scaffolding technologies was employed to develop chromosome-level genome assemblies of these species. The assembled genome sizes are 795.72 Mb for A. distichum and 1,108.53 Mb for F. ovata. The assemblies exhibit scaffold N50 lengths of 53.12 Mb and 68.97 Mb, with minimal gaps measuring 323.40 kb and 149.00 kb, and 97.71% and 98.82% BUSCO scores for Embryophyta single-copy orthologs, respectively, indicating high contiguity and completeness. The genomes contain 485.24 Mb and 691.68 Mb of repetitive sequences, 4,926 and 7,175 full-length long terminal repeat retrotransposons, and 49,414 and 57,587 protein-coding genes, respectively. The 14 pseudochromosomes encompass 93.80% of the A. distichum genome and 89.11% of the F. ovata genome, thereby demonstrating one-to-one chromosome-level collinearity. These high-quality genome assemblies serve as invaluable resources for genetic and breeding studies, facilitating a deeper understanding of the evolutionary history of these distinctive species.

High-Resolution Genome Assembly and Population Genetic Study of the Endangered Maple Acer pentaphyllum (Sapindaceae): Implications for Conservation Strategies

Abstract Acer pentaphyllum Diels (Sapindaceae), a highly threatened maple endemic to the dry-hot valleys of the Yalong River in western Sichuan, China, represents a valuable resource for horticulture and conservation. This study presents the first chromosomal-scale genome assembly of A. pentaphyllum (~626 Mb, 2n = 26), constructed using PacBio HiFi and Hi-C sequencing technologies. Comparative genomic analyses revealed significant recent genomic changes through rapid amplification of transposable elements, particularly long terminal repeat retrotransposons, coinciding with the dramatic climate change during recent uplift of the Hengduan Mountains. Genes involved in photosynthesis, plant hormone signal transduction, and plant-pathogen interaction showed expansion and/or positive selection, potentially reflecting adaptation to the species’ unique dry-hot valley habitat. Population genomic analysis of 227 individuals from 28 populations revealed low genetic diversity (1.04 ± 0.97 × 10-3) compared to other woody species. Phylogeographic patterns suggest an unexpected upstream colonization along the Yalong River, while Quaternary climate fluctuations drove its continuous lineage diversification and population contraction. Assessment of genetic diversity, inbreeding, and genetic load across populations revealed concerning levels of inbreeding and accumulation of deleterious mutations in small, isolated populations, particularly those at range edges (TKX, CDG, TES). Based on these results, we propose conservation strategies, including the identification of management units and recommendations for genetic rescue. These findings not only facilitate the conservation of A. pentaphyllum but also serve as a valuable resource for future horticultural development and as a model for similar studies on other endangered plant species adapted to extreme environments.

BLV-CoCoMo Dual qPCR Assay Targeting LTR Region for Quantifying Bovine Leukemia Virus: Comparison with Multiplex Real-Time qPCR Assay Targeting pol Region.

The proviral load (PVL) of the bovine leukemia virus (BLV) is a useful index for estimating disease progression and transmission risk. Real-time quantitative PCR techniques are widely used for PVL quantification. We previously developed a dual-target detection method, the "Liquid Dual-CoCoMo assay", that uses the coordination of common motif (CoCoMo) degenerate primers. This method can detect two genes simultaneously using a FAM-labeled minor groove binder (MGB) probe for the BLV long terminal repeat (LTR) region and a VIC-labeled MGB probe for the BoLA-DRA gene. In this study, we evaluated the diagnostic and analytical performance of the Dual-CoCoMo assay targeting the LTR region by comparing its performance against the commercially available Takara multiplex assay targeting the pol region. The diagnostic sensitivity and specificity of the Liquid Dual-CoCoMo assay based on the diagnostic results of the ELISA or original Single-CoCoMo qPCR were higher than those of the Takara multiplex assay. Furthermore, using a BLV molecular clone, the analytical sensitivity of our assay was higher than that of the Takara multiplex assay. Our results provide the first evidence that the diagnostic and analytical performances of the Liquid Dual-CoCoMo assay are better than those of commercially available multiplex assays that target the pol region.

Chromosome‐scale genomes of Toona fargesii provide insights into competency of root sprouting

AbstractToona fargesii A. Chev., a versatile tree in the Toona genus of the Meliaceae family, is renowned for its exquisite timber and medicinal properties, offering promising benefits. Due to natural regeneration obstacles and long‐term excessive exploitation, it has been threatened in China. Intriguingly, root sprouting, which may diminish the genetic diversity and hinder population development, dominates the reproductive pattern of T. fargesii in the wild. However, the lack of complete genome information has hampered basic studies on the regeneration, classification, evolution and conservation of this species. Here, we report the genome of T. fargesii, which was sequenced using the PacBio platform and assembled into a high‐quality genome with a total size of 535.24 Mb. Of this, 97.93% of the assembled contigs were anchored onto 28 pseudochromosomes, achieving a chromosome‐level genome. The long terminal repeat assembly index score was 21.34, and the consensus quality value was 39.90%, indicating the accuracy and completeness of the genome. Comparative genome analysis suggested that a recent whole genome duplication event occurred between 22.1 and 50.1 Mya in the Toona genus, with the divergence time between T. fargesii and its relative T. sinensis estimated at approximately ~16.7 Mya. Additionally, 13 TfARR genes, which play integral roles in root sprouting by mediating cytokinin signaling, underwent rapid gene expansion and showed significant enrichment in the plant hormone signal transduction pathway. Furthermore, transcriptomic analysis demonstrated that differentially expressed genes between root sprouts and nonroot sprouts were significantly enriched in the zeatin biosynthesis pathway, indicating that cytokinin regulation is involved in root sprouting development. Collectively, the findings provide valuable genomic resources for the Toona genus and genetic insights into the mechanisms of root sprouting in T. fargesii.

Genome sequences of five Klebsiella bacteriophages that belong to the genus Jiaodavirus.

We describe the genomes of five lytic Klebsiella pneumoniae myophages, therapeutic candidates, that belong to the family Straboviridae and genus Jiaodavirus. The genomes ranged from 165,574 to 169,768 bp, with ca. 40% GC content, contained 289-300 coding sequences, had 15-16 tRNA genes, and no terminal repeats.

Degradation and stable maintenance of adeno-associated virus inverted terminal repeats in E. coli.

Current plasmid propagation in E. coli compromises large inverted repeats, such as inverted terminal repeats (ITRs) of adeno-associated virus (AAV). Direct long-read sequencing analyses upon varying strains and culture conditions revealed ITR instability caused by a slipped misalignment mechanism, although other mechanism probably contribute. ITRs stabilized in absence of SbcC, which is part of the SbcCD nuclease complex, a human Mre11-Rad50 homolog, or at elevated growth temperatures (e.g. 42°C), with a combination being optimal. Resulting full ITR transgene plasmids improved rAAV yield and purity in HEK-293 productions. The findings advance plasmid biology, cloneable sequencesand therapeutic AAV manufacturing.

Deep Sequencing Analysis of Virome Components, Viral Gene Expression and Antiviral RNAi Responses in Myzus persicae Aphids.

The green peach aphid (Myzus persicae) is a generalist pest damaging crops and transmitting viral pathogens. Using Illumina sequencing of small (s)RNAs and poly(A)-enriched long RNAs, we analyzed aphid virome components, viral gene expression and antiviral RNA interference (RNAi) responses. Myzus persicae densovirus (family Parvoviridae), a single-stranded (ss)DNA virus persisting in the aphid population, produced 22 nucleotide sRNAs from both strands of the entire genome, including 5'- and 3'-inverted terminal repeats. These sRNAs likely represent Dicer-dependent small interfering (si)RNAs, whose double-stranded RNA precursors are produced by readthrough transcription beyond poly(A) signals of the converging leftward and rightward transcription units, mapped here with Illumina reads. Additionally, the densovirus produced 26-28 nucleotide sRNAs, comprising those enriched in 5'-terminal uridine and mostly derived from readthrough transcripts and those enriched in adenosine at position 10 from their 5'-end and mostly derived from viral mRNAs. These sRNAs likely represent PIWI-interacting RNAs generated by a ping-pong mechanism. A novel ssRNA virus, reconstructed from sRNAs and classified into the family Flaviviridae, co-persisted with the densovirus and produced 22 nucleotide siRNAs from the entire genome. Aphids fed on plants versus artificial diets exhibited distinct RNAi responses affecting densovirus transcription and flavivirus subgenomic RNA production. In aphids vectoring turnip yellows virus (family Solemoviridae), a complete virus genome was reconstituted from 21, 22 and 24 nucleotide viral siRNAs likely acquired with plant phloem sap. Collectively, deep-sequencing analysis allowed for the identification and de novo reconstruction of M. persicae virome components and uncovered RNAi mechanisms regulating viral gene expression and replication.

Interferon response and epigenetic modulation by SMARCA4 mutations drive ovarian tumor immunogenicity.

Cell-intrinsic mechanisms of immunogenicity in ovarian cancer (OC) are not well understood. Damaging mutations in the SWI/SNF chromatin remodeling complex, such as SMARCA4 (BRG1), are associated with improved response to immune checkpoint blockade; however, the mechanism by which this occurs is unclear. We found that SMARCA4 loss in OC models resulted in increased cancer cell-intrinsic immunogenicity, characterized by up-regulation of long-terminal RNA repeats, increased expression of interferon-stimulated genes, and up-regulation of antigen presentation machinery. Notably, this response was dependent on STING, MAVS, and IRF3 signaling but was independent of the type I interferon receptor. Mouse ovarian and melanoma tumors with SMARCA4 loss demonstrated increased infiltration and activation of cytotoxic T cells, NK cells, and myeloid cells in the tumor microenvironment. These results were recapitulated in BRG1 inhibitor-treated SMARCA4-proficient tumor models, suggesting that modulation of chromatin remodeling through targeting SMARCA4 may serve as a strategy to overcome cancer immune evasion.

Expression of an Efficient Selection Marker Out of a Duplicated Site in the ITRs of a Modified Vaccinia Virus Ankara (MVA).

Background/Objectives: Poxviruses are large DNA viruses that replicate in the host cytoplasm without a nuclear phase. As vaccine vectors, they can package and express large recombinant cassettes from different positions of their genomic core region. We present a comparison between wildtype modified vaccinia Ankara (MVA) and isolate CR19, which has significantly expanded inverted terminal repeats (ITRs). With this expansion, a site in wildtype MVA, called deletion site (DS) IV, has been duplicated at both ends of the genome and now occupies an almost central position in the newly formed ITRs. Methods: We inserted various reporter genes into this site and found that the ITRs can be used for transgene expression. However, ITRs are genomic structures that can rapidly adapt to selective pressure through transient duplication and contraction. To test the potential utility of insertions into viral telomers, we inserted a factor from the cellular innate immune system that interferes with viral replication as an example of a difficult transgene. Results: A site almost in the centre of the ITRs can be used for transgene expression, and both sides are mirrored into identical copies. The example of a challenging transgene, tetherin, proved to be surprisingly efficient in selecting candidate vectors against the large background of parental viruses. Conclusions: Insertion of transgenes into ITRs automatically doubles the gene doses. The functionalisation of viruses with tetherin may accelerate the identification and generation of recombinant vectors for personalised medicine and pandemic preparedness.

A chromosome-level genome assembly of the Peruvian Algarrobo (Neltuma pallida) provides insights on its adaptation to its unique ecological niche.

The dry forests of northern Peru are dominated by the legumous tree Neltuma pallida which is adapted to hot arid and semiarid conditions in the tropics. Despite having been successfully introduced in multiple other areas around the world, N. pallida is currently threatened in its native area, where it is invaluable for the dry forest ecosystem and human subsistence. A major tool for enhancing ecosystem conservation and understanding the adaptive properties of N. pallida to dry forest ecosystems is the construction of a reference genome sequence. Here, we report on a high-quality reference genome for N. pallida. The final genome assembly size is 403.7 Mb, consisting of 14 pseudochromosomes and 63 scaffolds with an N50 size of 26.2 Mb and a 34.3% GC content. Use of Benchmarking Universal Single Copy Orthologs revealed 99.2% complete orthologs. Long terminal repeat elements dominated the repetitive sequence content which was 51.2%. Genes were annotated using N. pallida transcripts, plant protein sequences, and ab initio predictions resulting in 22,409 protein-coding genes encoding 24,607 gene models. Comparative genomic analysis showed evidence of rapidly evolving gene families related to disease resistance, transcription factors, and signaling pathways. The chromosome-scale N. pallida reference genome will be a useful resource for understanding plant evolution in extreme and highly variable environments.