Host Genome Research Articles

Towards development of the 4c-based method detecting interactions of plasmid dna with host genome

Chromosome conformation capture techniques have revolutionized our understanding of chromatin architecture and dynamics at the genome-wide scale. In recent years, these methods have been applied to a diverse array of species, revealing fundamental principles of chromosomal organization. However, structural organization of the extrachromosomal entities, like viral genomes or plasmids, and their interactions with the host genome, remain relatively underexplored. In this work, we introduce an enhanced 4C-protocol tailored for probing plasmid DNA interactions. We design specific plasmid vector and optimize protocol to allow high detection rate of contacts between the plasmid and host DNA.

3D chromatin maps of a brown alga reveal U/V sex chromosome spatial organization.

Nuclear three dimensional (3D) folding of chromatin structure has been linked to gene expression regulation and correct developmental programs, but little is known about the 3D architecture of sex chromosomes within the nucleus, and how that impacts their role in sex determination. Here, we determine the sex-specific 3D organization of the model brown alga Ectocarpus chromosomes at 2 kb resolution, by mapping long-range chromosomal interactions using Hi-C coupled with Oxford Nanopore long reads. We report that Ectocarpus interphase chromatin exhibits a non-Rabl conformation, with strong contacts among telomeres and among centromeres, which feature centromere-specific LTR retrotransposons. The Ectocarpus chromosomes do not contain large local interactive domains that resemble TADs described in animals, but their 3D genome organization is largely shaped by post-translational modifications of histone proteins. We show that the sex determining region (SDR)within the U and V chromosomes are insulated and span the centromeres andwe link sex-specific chromatin dynamics and gene expression levels to the 3D chromatin structure of the U and V chromosomes. Finally, we uncover the unique conformation of a large genomic region on chromosome 6 harboring an endogenous viral element, providing insights regarding the impact of a latent giant dsDNA virus on the host genome's 3D chromosomal folding.

Variable orthogonality of serine integrase interactions within the ϕC31 family

Serine integrases are phage- (or mobile element-) encoded enzymes that catalyse site-specific recombination reactions between a short DNA sequence on the phage genome (attP) and a corresponding host genome sequence (attB), thereby integrating the phage DNA into the host genome. Each integrase has its unique pair of attP and attB sites, a feature that allows them to be used as orthogonal tools for genome modification applications. In the presence of a second protein, the Recombination Directionality Factor (RDF), integrase catalyses the reverse excisive reaction, generating new recombination sites, attR and attL. In addition to promoting attR x attL reaction, the RDF inhibits attP x attB recombination. This feature makes the directionality of integrase reactions programmable, allowing them to be useful for building synthetic biology devices. In this report, we describe the degree of orthogonality of both integrative and excisive reactions for three related integrases (ϕC31, ϕBT1, and TG1) and their RDFs. Among these, TG1 integrase is the most active, showing near complete recombination in both attP x attB and attR x attL reactions, and the most directional in the presence of its RDF. Our findings show that there is varying orthogonality among these three integrases – RDF pairs. ϕC31 integrase was the least selective, with all three RDFs activating it for attR x attL recombination. Similarly, ϕC31 RDF was the least effective among the three RDFs in promoting the excisive activities of the integrases, including its cognate ϕC31 integrase. ϕBT1 and TG1 RDFs were noticeably more effective than ϕC31 RDF at inhibiting attP x attB recombination by their respective integrases, making them more suitable for building reversible genetic switches. AlphaFold-Multimer predicts very similar structural interactions between each cognate integrase – RDF pair. The binding surface on the RDF is much more conserved than the binding surface on the integrase, an indication that specificity is determined more by the integrase than the RDF. Overall, the observed weak integrase/RDF orthogonality across the three enzymes emphasizes the need for identifying and characterizing more integrase – RDF pairs. Additionally, the ability of a particular integrase’s preferred reaction direction to be controlled to varying degrees by non-cognate RDFs provides a path to tunable, non-binary genetic switches.

Evaluation of Interfering RNA Efficacy in Treating Hepatitis B: Is It Promising?

Background: Despite an existing safe and effective vaccine for hepatitis B virus (HBV), it is still a major public health concern. Nowadays, several drugs are used to treat chronic hepatitis B; however, full healing remains controversial. The viral covalently closed circular DNA (cccDNA) formed by HBV forms a major challenge in its treatment, as does the ability of HBV to integrate itself into the host genome, which enables infection reactivation. Interfering RNA (RNAi) is a gene-silencing post-transcriptional mechanism which forms as a promising alternative to treat chronic hepatitis B. The aim of the present review is to assess the evolution of hepatitis B treatment approaches based on using RNA interference. Methods: Data published between 2016 and 2023 in scientific databases (PubMed, PMC, LILACS, and Bireme) were assessed. Results: In total, 76,949 articles were initially identified and quality-checked, and 226 eligible reports were analyzed in depth. The main genomic targets, delivery systems, and major HBV therapy innovations are discussed in this review. This review reinforces the therapeutic potential of RNAi and identifies the need for conducting further studies to fill the remaining gaps between bench and clinical practice.

Prospectives of developing therapeutic HPV vaccines

Human papillomavirus (HPV) represents one of the most serious global public health problems. Malignant female and male diseases mainly result from persistent HPV infection. Cancer belongs to a high mortality rate disease. It has been established that HPV infection causes about 70% vaginal cancer, 50% male genital cancer, 90% anal cancer and 60% head-and-neck cancer. Annually, a large number of people develop various HPV-caused cancer types, dominated by cervical cancer, one of the most common and aggressive types of cancer that threatens health holding the fourth place among most female common cancer worldwide. According to the World Health Organization (WHO), in 2020, about 600 cases of cervical cancer are recorded daily in different countries. Emergence of cervical cancer is closely related to factors such as long-term (persistent) HPV infection and somatic mutations of the host genome. Although HPV infection can be detected and cured early with highly effective screening methods and surgical procedures, the carcinogenic risk of HPV related diseases constantly increases, which elimination faces certain difficulties, especially in low- and mid-developed countries. The most acceptable solution to this is development and implementation of therapeutic vaccines for prevention and treatment of HPV related diseases. Three licensed HPV vaccines based on L1 type virus-like particles (L1-VLPs) technology are available globally: bivalent (HPV-2), quadrivalent (HPV-4) and nonavalent (HPV-9) vaccines. These vaccines demonstrated effectiveness in reducing HPV-related cervical cancer rate by up to 90% worldwide. However, the therapeutic effect of these vaccines on persistent HPV infection and lesions has not been observed. Therapeutic HPV vaccines candidates targeted Ye6/Ye7 cancer proteins activate cellular immunity that eliminates existing HPV infection. Here we review types, mechanisms of action and clinical effects of therapeutic HPV vaccines, as well as current and future developments in the field for prevention and treatment of HPV related diseases.

Revealing host genome–microbiome networks underlying feed efficiency in dairy cows

Ruminants have the ability to digest human-inedible plant materials, due to the symbiotic relationship with the rumen microbiota. Rumen microbes supply short chain fatty acids, amino acids, and vitamins to dairy cows that are used for maintenance, growth, and lactation functions. The main goal of this study was to investigate gene-microbiome networks underlying feed efficiency traits by integrating genotypic, microbial, and phenotypic data from lactating dairy cows. Data consisted of dry matter intake (DMI), net energy secreted in milk, and residual feed intake (RFI) records, SNP genotype, and 16S rRNA rumen microbial abundances from 448 mid-lactation Holstein cows. We first assessed marginal associations between genotypes and phenotypic and microbial traits through genomic scans, and then, in regions with multiple significant hits, we assessed gene-microbiome-phenotype networks using causal structural learning algorithms. We found significant regions co-localizing the rumen microbiome and feed efficiency traits. Interestingly, we found three types of network relationships: (1) the cow genome directly affects both rumen microbial abundances and feed efficiency traits; (2) the cow genome (Chr3: 116.5 Mb) indirectly affects RFI, mediated by the abundance of Syntrophococcus, Prevotella, and an unknown genus of Class Bacilli; and (3) the cow genome (Chr7: 52.8 Mb and Chr11: 6.1–6.2 Mb) affects the abundance of Rikenellaceae RC9 gut group mediated by DMI. Our findings shed light on how the host genome acts directly and indirectly on the rumen microbiome and feed efficiency traits and the potential benefits of the inclusion of specific microbes in selection indexes or as correlated traits in breeding programs. Overall, the multistep approach described here, combining whole-genome scans and causal network reconstruction, allows us to reveal the relationship between genome and microbiome underlying dairy cow feed efficiency.

C→U transition biases in SARS-CoV-2: still rampant 4 years from the start of the COVID-19 pandemic.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in the pandemic and post-pandemic periods has shown a remarkable capacity to adapt and evade human immune responses and increase its human-to-human transmissibility. The genome of SARS-CoV-2 is also increasingly scarred by the effects of multiple C→U mutations from host genome editing as a cellular defense mechanism akin to restriction factors for retroviruses. Through the analysis of large data sets of SARS-CoV-2 isolate sequences collected throughout the pandemic period and beyond, we show that C→U transitions have driven a base compositional change over time amounting to a net loss of C bases and accumulation of U's at a rate of approximately 0.2%-0.25%/decade. Most C→U substitutions occurred in the absence of the preferred upstream-base context or targeting of unpaired RNA bases previously associated with the host RNA editing protein, APOBEC 3A. The analyses provide a series of testable hypotheses that can be experimentally investigated in the future.

Correction: Gene expression and chromatin conformation of microglia in virally suppressed people with HIV.

"Despite ART, we detected occasional microglia containing cell-associated HIV RNA and HIV DNA integrated into open regions of the host's genome (∼0.005%)" should be corrected to: "Despite ART, we detected occasional microglia containing cell-associated HIV RNA and HIV DNA integrated into open regions of the host's genome (∼0.5%)."

Deep mining reveals the diversity of endogenous viral elements in vertebrate genomes

Integration of viruses into host genomes can give rise to endogenous viral elements (EVEs), which provide insights into viral diversity, host range and evolution. A systematic search for EVEs is becoming computationally challenging given the available genomic data. We used a cloud-computing approach to perform a comprehensive search for EVEs in the kingdoms Shotokuvirae and Orthornavirae across vertebrates. We identified 2,040 EVEs in 295 vertebrate genomes and provide evidence for EVEs belonging to the families Chuviridae, Paramyxoviridae, Nairoviridae and Benyviridae. We also find an EVE from the Hepacivirus genus of flaviviruses with orthology across murine rodents. In addition, our analyses revealed that reptarenaviruses and filoviruses probably acquired their glycoprotein ectodomains three times independently from retroviral elements. Taken together, these findings encourage the addition of 4 virus families and the Hepacivirus genus to the growing virus fossil record of vertebrates, providing key insights into their natural history and evolution.

The impact of insertion bias into piRNA clusters on the invasion of transposable elements.

This study challenges the pre-existing understanding of the TE dynamics by investigating the potential adaptive role of insertion bias into piRNA clusters. Using extensive forward simulations, we demonstrate that while insertion bias significantly alters TE invasion dynamics, it is generally not beneficial for the TE's spread in populations. Our results also show that transposable elements (TEs) that avoid piRNA clusters tend to do better than those that are more likely to preferencially insert into these clusters. This work provides novel insights into the complex dynamics between TEs and host genomes, showing the limited scenarios where the insertion bias could be advantageous to TEs. These results open new area for research into TE invasion dynamics and the evolution of host-TE interactions, further contributing to our understanding of genome evolution and stability.

Agrobacterium virulence factors induce the expression of host DNA repair-related genes without promoting major genomic damage

This study aimed to investigate whether the plant DNA damage levels and DNA damage response (DDR) are regulated during Agrobacterium infection and potentially manipulated by Agrobacterium to facilitate T-DNA integration. We investigated the plant genomic response to Agrobacterium infection by measuring gamma H2AX levels, which reflect the levels of double-strand DNA breaks (DSBs), and by characterizing transcription of three major DNA repair marker genes NAC82, KU70, and AGO2. These experiments revealed that, globally, Agrobacterium infection did not result in a major increase in DSB content in the host genome. The transcription of the DNA damage repair genes, on the other hand, was elevated upon the wild-type Agrobacterium infection. This transcriptional outcome was largely negated by a mutation in the bacterial virB5 gene which encodes the virulence (Vir) protein B5, a minor component of Agrobacterium pilus necessary for the translocation of Vir effector proteins into the host cell, suggesting that the transcriptional activation of the cellular DNA damage repair machinery requires the transport into the host cell of the Agrobacterium effectors, i.e., the VirD2, VirD5, VirE2, VirE3, and VirF proteins. Most likely, a combination of several of these Vir effectors is required to activate the host DNA repair as their individual loss- or gain-of-function mutants did not significantly affect this process.

Widespread HCD-tRNA derived SINEs in bivalves rely on multiple LINE partners and accumulate in genic regions

BackgroundShort interspersed nuclear elements (SINEs) are non-autonomous non-LTR retrotransposons widespread across eukaryotes. They exist both as lineage-specific, fast-evolving elements and as ubiquitous superfamilies characterized by highly conserved domains (HCD). Several of these superfamilies have been described in bivalves, however their overall distribution and impact on host genome evolution are still unknown due to the extreme scarcity of transposon libraries for the clade. In this study, we examined more than 40 bivalve genomes to uncover the distribution of HCD-tRNA-related SINEs, discover novel SINE-LINE partnerships, and understand their possible role in shaping bivalve genome evolution.ResultsWe found that bivalve HCD SINEs have an ancient origin, and they can rely on at least four different LINE clades. According to a “mosaic” evolutionary scenario, multiple LINE partner can promote the amplification of the same HCD SINE superfamilies while homologues LINE-derived tails are present between different superfamilies. Multiple SINEs were found to be highly similar between phylogenetically related species but separated by extremely long evolutionary timescales, up to ~ 400 million years. Studying their genomic distribution in a subset of five species, we observed different patterns of SINE enrichment in various genomic compartments as well as differences in the tendency of SINEs to form tandem-like and palindromic structures also within intronic sequences. Despite these differences, we observed that SINEs, especially older ones, tend to accumulate preferentially within genes, or in their close proximity, consistently with a model of survival bias for less harmful, short non-coding transposons in euchromatic genomic regions.ConclusionHere we conducted a wide characterization of tRNA-related SINEs in bivalves revealing their taxonomic distribution and LINE partnerships across the clade. Moreover, through the study of their genomic distribution in five species, we highlighted commonalities and differences with other previously studied eukaryotes, thus extending our understanding of SINE evolution across the tree of life.

Signature of viral fossils: a comparative genomics approach to understand the diversity of endogenous retroviruses in bats

Endogenous retroviruses (ERVs) are traces of past viral infections commonly found in vertebrate genomes. Many ERVs are tightly regulated by the host genomes and co-opted for various functions within the hosts. Bats are the only true volant mammals, with the smallest mammalian genomes and a high fraction of ERVs within the genomes. They are important hosts for various zoonotic viral pathogens and can effectively modulate their immune response to tolerate viral infections. Integrations of retroviruses have been implicated as one of the mechanisms by which bats have co-evolved strategies to combat viral infections. In this study, we investigated the diversity of ERVs in over 40 publicly available bat genomes to understand the distribution and the evolution of ERVs within bats. We observed all classes of ERVs within bat genomes including even the complex lenti retroviruses. Alpha and spuma retroviruses which are, generally considered rare in mammals, were common within bats. We observed a positive correlation between bat genome size and length of ERV elements. Interestingly, nearly 30% of the ERVs within bats are intact suggesting a recent origin or co-option by the host genome. Future studies focusing on comparative genomic and experimental data will be critical to understand the role of these ERVs in host genome evolution.

Multiomics integration unravels genotype‐microbiome interactions shaping the conjunctival transcriptome

AbstractGene expression is a molecular phenotype shaped by the interplay between genotype and environment. The microbiome represents a critical environmental exposure for the host. However, the genotype‐microbiome interactions (GMIs) shaping the host transcriptome remain largely unexplored. Here, we integrated conjunctival multiomics data from 120 pairs of twins to investigate GMIs. We identified 272,972 expression quantitative trait loci associated with 5946 genes and 241,073 genotype‐controlled correlations between gene expression and microbial abundance. We developed a modeling approach, MicroGenix, that screens for GMIs from host genome, transcriptome, and microbiome data and identifies GMIs associated with the disease through gene‐based association tests. We applied MicroGenix to the conjunctival data set and found that incorporating GMIs into gene expression prediction models significantly increased prediction accuracy. The genes with increased accuracy were overrepresented by those encoding cell adhesion molecules. We further used MicroGenix to predict the transcriptome from conjunctival metagenomes and identified GMIs associated with ocular surface disease. Our work provides a resource for studying host‐microbiome interactions at the conjunctiva and a computational approach to investigate GMIs using multiomics data.

Direct genome sequencing of respiratory viruses from low viral load clinical specimens using the target capture sequencing technology.

The use of metagenomic next-generation sequencing technology to obtain complete viral genome sequences directly from clinical samples with low viral load remains challenging-especially in the case of respiratory viruses-due to the low copy number of viral versus host genomes. To overcome this limitation, target capture sequencing for the enrichment of specific genomes has been developed and applied for direct genome sequencing of viruses. However, as the efficiency of enrichment varies depending on the probes, the type of clinical sample, etc., validation is essential before target capture sequencing can be applied to clinical diagnostics. In this study, we evaluated the utility of target capture sequencing with a comprehensive viral probe panel for clinical respiratory specimens collected from patients diagnosed with SARS-CoV-2 or influenza type A. We focused on clinical specimens containing low copy numbers of viral genomes. Target capture sequencing yielded approximately 180- and 2,000-fold higher read counts of SARS-CoV-2 and influenza A virus, respectively, than metagenomic sequencing when the RNA extracted from specimens contained 59.3 copies/µL of SARS-CoV-2 or 625.1 copies/µL of influenza A virus. In addition, the target capture sequencing identified sequence reads in all SARS-CoV-2- or influenza type A-positive specimens with <26 RNA copies/µL, some of which also yielded >70% of the full-length genomes of SARS-CoV-2 or influenza A virus. Furthermore, the target capture sequencing using comprehensive probes identified co-infections with viruses other than SARS-CoV-2, suggesting that this approach will not only detect a wide range of viruses but also contribute to epidemiological studies.IMPORTANCETarget capture sequencing has been developed and applied for direct genome sequencing of viruses in clinical specimens to overcome the low detection sensitivity of metagenomic next-generation sequencing. In this study, we evaluated the utility of target capture sequencing with a comprehensive viral probe panel for clinical respiratory specimens collected from patients diagnosed with SARS-CoV-2 or influenza type A, focusing on clinical specimens containing low copy numbers of viral genomes. Our results showed that the target capture sequencing yielded dramatically higher read counts than metagenomic sequencing for both viruses. Furthermore, the target capture sequencing using comprehensive probes identified co-infections with other viruses, suggesting that this approach will not only detect a wide range of viruses but also contribute to epidemiological studies.

Chromosomal integrons are genetically and functionally isolated units of genomes.

Integrons are genetic elements that increase the evolvability of bacteria by capturing new genes and stockpiling them in arrays. Sedentary chromosomal integrons (SCIs) can be massive and highly stabilized structures encoding hundreds of genes, whose function remains generally unknown. SCIs have co-evolved with the host for aeons and are highly intertwined with their physiology from a mechanistic point of view. But, paradoxically, other aspects, like their variable content and location within the genome, suggest a high genetic and functional independence. In this work, we have explored the connection of SCIs to their host genome using as a model the Superintegron (SI), a 179-cassette long SCI in the genome of Vibrio cholerae N16961. We have relocated and deleted the SI using SeqDelTA, a novel method that allows to counteract the strong stabilization conferred by toxin-antitoxin systems within the array. We have characterized in depth the impact in V. cholerae's physiology, measuring fitness, chromosome replication dynamics, persistence, transcriptomics, phenomics, natural competence, virulence and resistance against protist grazing. The deletion of the SI did not produce detectable effects in any condition, proving that-despite millions of years of co-evolution-SCIs are genetically and functionally isolated units of genomes.

EVE-X: Software to Identify Novel Viral Insertions in Wild-Caught Arthropod Hosts From Next-Generation Short Read Data.

Eukaryotic genomes harbour sequences derived from non-retroviral RNA viruses, known as endogenous viral elements (EVEs) or non-retroviral integrated RNA virus sequences (NIRVS). These sequences represent a record of past infections and have been implicated in host anti-viral response. We have created a program to identify viral sequences integrated in a host genome. It begins with a specimen BAM file and outputs candidate NIRVS, along with putative host insertion sites and overlapping genomic features of the host genome in XML and visual formats, with minimal intermediary intervention. We ran through this software short-read data derived from the genomes of 222 wild-caught A. aegypti mosquitoes, from a dozen geographical regions, and located putative NIRVS from seven virus families. This program is as accurate as currently available software for NIRVS detection, and represents a significant improvement in adaptability and user-friendliness. Furthermore, the flexibility of this pipeline allows the user to search for sequence integrations across the genome of any organism, as long as a query sequence database and a reference genome is provided. Potential extended applications include identification of integrated transgenic sequences used for research or vector control strategies.

Multi-omics insights into the energy compensation of rumen microbiota of grazing yaks in cold season.

The ability of yaks to adapt to the extreme environment of low temperatures and hypoxia at cold seasons on the Qinghai-Tibet Plateau (QTP) is related to the host genome; however, the convergent evolution of rumen microbiomes in host adaption is unknown. Here, we conducted a multi-omics study on the rumen fluid of grazing yaks from warm (July) and cold (December) seasons on the QTP to evaluate the convergent evolution of rumen microbiomes in the adaptation of grazing yaks to cold-seasons environments. The results showed that grazing yaks at cold seasons had higher fibrolytic enzyme activities and volatile fatty acids (VFAs) concentrations, and the relative abundance of Firmicutes and the ratio Firmicutes to Bacteroidetes was significantly higher than that of yaks at warm seasons. Macrogenomic analyses showed that genes involved in forming VFAs and arginine were significantly enriched in cold-season yaks. Transcriptome analyses of the rumen epithelium showed that 72 genes associated with VFAs absorption and transport were significantly upregulated in cold-season yaks. Metabolomic analyses showed that the levels of ornithine, related to efficient nitrogen utilization, were significantly upregulated in cold-season yaks. The synergistic role of rumen microbiomes in the adaptation of grazing yaks to extreme environments at cold seasons was revealed by multi-omics study.

Landscape of evolutionary arms races between transposable elements and KRAB-ZFP family

Transposable elements (TEs) are mobile parasitic sequences that have expanded within the host genome. It has been hypothesized that host organisms have expanded the Krüppel-associated box-containing zinc finger proteins (KRAB-ZFPs), which epigenetically suppress TEs, to counteract disorderly TE transpositions. This process is referred to as the evolutionary arms race. However, the extent to which this evolutionary arms race occurred across various TE families remains unclear. In the present study, we systematically explored the evolutionary arms race between TE families and human KRAB-ZFPs using public ChIP-seq data. We discovered and characterized new instances of evolutionary arms races with KRAB-ZFPs in endogenous retroviruses. Furthermore, we found that the regulatory landscape shaped by this arms race contributed to the gene regulatory networks. In summary, our results provide insight into the impact of the evolutionary arms race on TE families, the KRAB-ZFP family, and host gene regulatory networks.

Gene expression and chromatin conformation of microglia in virally suppressed people with HIV.

The presence of HIV in sequestered reservoirs is a central impediment to a functional cure, allowing HIV to persist despite life-long antiretroviral therapy (ART), and driving a variety of comorbid conditions. Our understanding of the latent HIV reservoir in the central nervous system is incomplete, because of difficulties in accessing human central nervous system tissues. Microglia contribute to HIV reservoirs, but the molecular phenotype of HIV-infected microglia is poorly understood. We leveraged the unique "Last Gift" rapid autopsy program, in which people with HIV are closely followed until days or even hours before death. Microglial populations were heterogeneous regarding their gene expression profiles but showed similar chromatin accessibility landscapes. Despite ART, we detected occasional microglia containing cell-associated HIV RNA and HIV DNA integrated into open regions of the host's genome (∼0.005%). Microglia with detectable HIV RNA showed an inflammatory phenotype. These results demonstrate a distinct myeloid cell reservoir in the brains of people with HIV despite suppressive ART. Strategies for curing HIV and neurocognitive impairment will need to consider the myeloid compartment to be successful.