Biology subjects

Kennedy, S.

Publications and source records attributed to Kennedy, S..

At least 19 recordsLinked to original sources

bioRxiv2026

An NF2-wildtype malignant meningioma cell line for basic and translational science

BackgroundMeningiomas are the most common primary nervous system neoplasm in adults. There are few good cellular models, especially of high grade/malignant meningiomas, to use to identify new therapeutic agents and treatment regimens. The widely available, partially characterized, NF2-wildtype (NF2wt) Grade 3 malignant meningioma cell line IOMM-Lee can help meet this need. MethodsWe generated new data to better characterize IOMM-Lee genomic and mtDNA variants, proliferation rate and colony-forming efficiency and sensitivity to ionizing radiation as a function of ATM kinase activity. A screen of 349 anti-cancer drugs identified multiple, mechanistically distinct clinical use drugs with nanomolar IC50 values and high drug sensitivity prediction scores. ResultsExome sequencing confirmed that IOMM-Lee is NF2wt, and contains a pathogenic TERT-promoter (c.-124C>T) variant. Population doubling times (PDT) were short (19-21 hrs), and colony forming efficiency (CFE) high, of up to 87%. IOMM-Lee is comparatively radiosensitive with a D10 of [~]3.9 Gy, and could be radiosensitized by AZD-1390-mediated ATM kinase inhibition. Thirty-four anti-cancer compounds spanning several mechanistic classes were identified that potently suppressed cell proliferation at sub-micromolar IC50 values with high Breeze 2.0 Drug Sensitivity Scores. Importance of the StudyWe provide new data to better characterize IOMM-Lee, the most widely used cell line model of human Grade 3 malignant meningioma. These data identify and characterize IOMM-Lee genomic alterations and mtDNA variants; quantify growth kinetics and ionizing radiation sensitivity; and identify multiple mechanistically distinct, clinical use drugs with nanomolar IC50 values, high drug sensitivity prediction scores and potential as meningioma systemic therapies. Our data more clearly locate IOMM-Lee in the landscape of genomically-defined meningiomas, and will aid better use of this experimentally tractable cell line model to understand meningioma biology and identify more effective malignant meningioma therapies and treatment regimens. Key pointsO_LIIOMM-Lee lacks NF2 mutations, though is clearly related to but distinct from many other meningiomas and meningioma cell lines. C_LIO_LIIOMM-Lee grows rapidly, is comparatively radio-sensitive, and can be suppressed by several mechanistic classes of anti-cancer agents at clinically achievable, sub-micromolar IC50 values with high Drug Sensitivity Scores. C_LIO_LIThe experimental tractability, simplicity and versatility of IOMM-Lee can facilitate analyses of many aspects of meningioma biology and therapeutic development across a wide range of in vitro, high throughput and in vivo xenograft/organoid protocols. C_LI

cancer biology↗

bioRxiv2026

SroA links SigS-dependent stress signaling to metabolic remodeling in Staphylococcus aureus

Staphylococcus aureus encounters diverse environmental conditions during colonization and infection, including fluctuations in nutrient availability, oxidative stress, and oxygen limitation. Adaptation to these environments requires regulatory systems that coordinate stress responses with metabolic remodeling. The extracytoplasmic function sigma factor SigS contributes to stress adaptation and virulence in S. aureus and directly activates expression of the sroAB operon, which encodes the small proteins SroA and SroB. While previous work demonstrated that SroA participates in feedback regulation of sigS expression, the broader physiological role of SroA has remained unclear. To define the regulatory functions of SroA, we performed RNA sequencing following inducible overexpression of sroA in S. aureus. Transcriptome analysis revealed extensive remodeling of gene expression, with approximately 200 transcripts significantly altered. Transcriptome analysis revealed coordinated repression of metabolic pathways (including nitrate respiration and nucleotide biosynthesis) alongside activation of stress-response and nutrient acquisition genes. Northern blot and quantitative RT-PCR analysis confirmed repression of narG and narJ transcripts following SroA overexpression. Consistent with these transcriptional changes, nitrate reduction assays demonstrated that SroA overexpression reduces nitrate respiration activity. In addition to repression of nitrate respiration genes, SroA overexpression broadly suppressed genes involved in de novo purine and pyrimidine biosynthesis. In contrast, transcripts associated with stress responses and nutrient acquisition, including the SOS-associated gene sosA and the phosphate transport gene pstS, were upregulated. Together, these findings identify SroA as a regulator that links stress-responsive signaling to metabolic remodeling in S. aureus, particularly through modulation of nitrate respiration pathways. ImportanceStaphylococcus aureus must rapidly adapt its metabolism to survive the diverse environments encountered during colonization and infection, including conditions where oxygen availability is limited. In this study, we identify a previously uncharacterized role for the small protein SroA in regulating metabolic adaptation in S. aureus. Transcriptome analysis revealed that SroA strongly represses genes involved in nitrate respiration, a pathway that enables bacteria to maintain energy production when oxygen is scarce. Consistent with these transcriptional changes, SroA overexpression reduced nitrate respiration activity. These findings reveal a regulatory link between stress-responsive signaling pathways and respiratory metabolism, expanding our understanding of how S. aureus adapts to oxygen-limited environments encountered during infection.

microbiology↗

bioRxiv2026

Closely related, yet phenotypically different - Genome assemblies of two sister species of widow spiders: Latrodectus hasselti and L. katipo, Theridiidae

Widow spiders of the genus Latrodectus are important animals for biomedical, pest and conservation research. Here, we present the assembled genomes of two closely related Latrodectus species: the Australian L. hasselti and the New Zealand endemic L. katipo. The genome of L. katipo consists of 13 scaffolds likely corresponding to chromosomes (90% of the total length) and 1267 short scaffolds (10%). It has a total length of 1.5 Gbp and BUSCO of 94.9%. The genome of L. hasselti consists of 379 scaffolds and has a total length of 1.7 Gbp and a BUSCO score of 95.4%. The repeat content is very similar in both genomes with a total proportion of 37.2% for L. katipo and 39.9% for L. hasselti. Genome annotation predicted 12706 and 15111 genes for L. katipo and L. hasselti respectively. An ortholog analysis shows large overlap between orthogroups suggesting either duplication events in L. hasselti or loss of genes in L. katipo.

genomics↗

bioRxiv2026

Comparison of environmental DNA and bulk DNA metabarcoding for assessing terrestrial arthropod diversity across three habitat types on Guam

DNA based methods offer a rapid and cost-effective way for detecting species occurrence and monitoring biodiversity; among them bulk DNA metabarcoding is well-established, and recently developed environmental DNA (eDNA)-based methods offer a non-destructive alternative. With a goal to develop suitable methods for assessing insect biodiversity in ecosystems for which DNA reference libraries are not well developed and incomplete, such as remote islands, we compared established bulk DNA metabarcoding methods with eDNA across three replicated terrestrial ecosystem types (limestone forest, degraded forest, and grassland) in Guam. Using two mitochondrial COI primer pairs, we performed bulk DNA metabarcoding of standard entomological collection methods (malaise traps, pan traps, vegetation beating), and compared the assessment of biodiversity with that from different eDNA sources (flowers, spider webs, leaves, tree trunks). In our samples, eDNA and bulk DNA metabarcoding both detected a large proportion of overall taxa (OTUs, 86.6% and 60.3%, respectively). Although DNA metabarcoding detected significantly more taxa, eDNA proved to be a reasonable non-destructive alternative. As expected, because of limitations in existing reference databases for remote habitats, species-level identification was achieved for only a few OTUs. Overall, the sampling approach was the dominant driver of arthropod diversity, explaining [~]17% of the observed variation, while habitat type accounted for [~]4%. Thus, each sampling approach captured some unique diversity signals and contributed to the complementary effect of maximizing detection. For rapid insect biodiversity surveys of terrestrial arthropods, we recommend an integrated metabarcoding approach, and in sensitive habitats where insect capture is undesirable, eDNA offers a powerful alternative to monitor diversity and community change.

ecology↗

bioRxiv2025

Transposon Regulation in the Caenorhabditis elegans Germline and Soma

Transposons are parasitic nucleic acids present in most genomes. The ability of transposons to mobilize makes them a source of genetic diversity and a threat to genome integrity. Interestingly, mutations in the C. elegans gene mut-2/rde-3 increases the rate of transposition in the germline, but not in the soma, suggesting that the C. elegans germline and soma employ different strategies to regulate Tc1 transposition. Here, we develop fluorescence reporters for studying DNA transposon regulation in living C. elegans in a tissue-specific manner and we use candidate gene testing and genetic screening approaches to identify factors that regulate Tc1 mobility in the germline and/or the soma of the animal. We find that both cytoplasmic and nuclear components of the RNA interference (RNAi) pathway silence Tc1 in the germline, but not in the soma. We identify a novel pathway involving the C. elegans ortholog of HNRNPC, and a gene we term suppressor of transposon mobilization (stm)-1, which regulates Tc1 primarily in the soma, likely by binding Tc1 RNA and preventing its splicing. Our findings reveal tissue-specific strategies for regulating parasitic nucleic acids and pave the way for future studies exploring how and why different tissues adopt different transposon silencing systems.

genetics↗

bioRxiv2025

Viral RNA pUGylation Promotes Antiviral Immunity in C. elegans

RNA interference (RNAi) is a component of the innate immune systems of many eukaryotes, including C. elegans. During RNAi in C. elegans, the nucleotidyltransferase RDE-3 modifies the 3 termini of mRNAs with polyUG (pUG) tails, which recruit RNA-dependent RNA Polymerase (RdRP) enzymes that drive gene silencing by synthesizing antisense small interfering (si)RNAs. During normal growth and development, RDE-3 pUGylates transposon RNAs to silence transposons and protect genomic integrity. How C. elegans identifies specific RNAs for pUGylation and whether the pUGylation system is used for other biological purposes is not yet known. Here we show that pUGylation contributes to antiviral immunity in C. elegans: During infection of C. elegans with Orsay virus, RDE-3 adds pUG tails to viral RNAs, which converts these RNAs into templates for RdRP-based antiviral siRNA production, thereby limiting viral replication. We present evidence that MUT-15 is critical for viral pUGylation because it interacts with RDE-3 and the NYN domain-containing endonuclease RDE-8, thus bridging the enzymes that cleave and pUGylate viral RNA, ensuring efficient antiviral immunity. We conclude that pUGylation promotes antiviral immunity in C. elegans and we provide molecular insights into how C. elegans identifies and neutralizes its internal and external parasitic threats. ImportanceViruses are a threat to all organisms. Therefore, organisms have evolved numerous systems to recognize and neutralize viruses. Many of these systems, which are referred to as innate immune systems, function by recognizing unique molecular characteristics of viral genetic material. One such innate immune system is RNA interference. RNA interference uses double stranded RNA, which is an obligatory byproduct of replication for many viruses, as a weapon to fight viruses. In this work, we provide molecular insights into how the nematode C. elegans uses RNA interference and viral double stranded RNAs to defend itself against viral invaders.

genetics↗

bioRxiv2025

Novel biomarkers in mutation-specific gene profiles highlight the involvement of COL9A3 in cancer cell plasticity in uveal melanoma

UM is a deadly ocular malignancy with well-described genetic alterations that predict disease outcome. However, our current understanding of the biological underpinnings of high-risk uveal melanoma progression remains relatively limited. On the basis of RNA expression profiles, we identified 12 novel biomarkers associated with high-risk UM, with protein expression validation of the 2 top markers RBFOX2 and COL9A3. Moreover, we investigated the functional contribution of COL9A3 via its overexpression in low-risk and high-risk UM cell lines. Our data suggest that COL9A3 enhances cell motility and cell proliferation and alters morphology specifically in high-risk UM cells. Furthermore, zebrafish xenograft studies revealed improved cell dissemination in a low-risk UM cell line upon overexpression of COL9A3, whereas a high-risk UM cell line remained under wild-type conditions. Interestingly, RNA sequencing revealed a high-stress profile in high-risk UM cell lines overexpressing COL9A3 and suggested the upregulation of plasticity markers, such as CD44, Nestin, EZH2, ABCB5, PAX3 and CD166, as a coping mechanism. These markers are known as differentiation markers during the development of melanocyte biogenesis and are implicated in plasticity in other high-risk cancers. The relationship between COL9A3 and plasticity was validated by multiplex immunohistochemistry of FFPE tissue, which demonstrated colocalization of COL9A3, CD44 and Nestin. BAP1mut-UM samples presented higher levels of COL9A3, CD44 and Nestin expression than EIF1AXmut- or SF3B1mut UM samples did. A positive correlation between COL9A3 and Nestin expression in triple-positive cells, regardless of the UM subtype, was observed. Together, our data suggest that COL9A3 is a novel high-risk biomarker that may be involved in UM cell plasticity and is most prevalent in BAP1mut UM.

cancer biology↗

bioRxiv2025

An RNA Splicing System that Excises Transposons from Animal mRNAs

All genomes harbor mobile genetic parasites called transposable elements (TEs). Here we describe a system, which we term SOS splicing, that protects C. elegans and human genes from DNA transposon-mediated disruption by excising these TEs from host mRNAs. SOS splicing, which operates independently of the spliceosome, is a pattern recognition system triggered by base-pairing of inverted terminal repeat elements, which are a defining feature of the DNA transposons. We identify three factors required for SOS splicing in both C. elegans and human cells; AKAP17A, which binds TE-containing mRNAs; the RNA ligase RTCB; and CAAP1, which bridges RTCB and AKAP17A, allowing RTCB to ligate mRNA fragments generated by TE excision. We propose that SOS splicing is a novel, conserved, and RNA structure-directed mode of mRNA splicing and that one function of SOS splicing is to genetically buffer animals from the deleterious effects of TE-mediated gene perturbation.

genetics↗

bioRxiv2023

Germ granule compartments coordinate specialized small RNA production

Germ granules are biomolecular condensates present in most animal germ cells. One function of germ granules is to help maintain germ cell totipotency by organizing mRNA regulatory machinery, including small RNA-based gene regulatory pathways. The C. elegans germ granule is compartmentalized into multiple subcompartments whose biological functions are largely unknown. Here, we identify a new subcompartment of the C. elegans germ granule, which we term the E compartment. The E compartment is nonrandomly positioned within the germ granule. We identified five proteins that localize to the E compartment, including the RNA-dependent RNA polymerase (RdRP) EGO-1, the Dicer-related helicase DRH-3, the Tudor domain-containing protein EKL-1, and two intrinsically disordered proteins, EGC-1 and ELLI-1. Localization of EGO-1 to the E granule enables synthesis of a specialized class of 22G RNAs, which derive exclusively from 5 regions of a subset of germline-expressed mRNAs. Defects in E compartment assembly elicit disordered production of endogenous siRNAs, which disturbs fertility and the RNAi response. Our results define a new subcompartment of the C. elegans germ granule and suggest that one function of germ granule compartmentalization is to facilitate the localized production of specialized classes of small regulatory RNAs.

cell biology↗

bioRxiv2023

A multivariate outcome test of covariance

Multivariate analysis is becoming central in studies investigating high-throughput molecular data, yet, some important features of these data are seldom explored. Here, we present MANOCCA (Multivariate Analysis of Conditional CovAriance), a powerful method to test for the effect of a predictor on the covariance matrix of a multivariate outcome. The proposed test is by construction orthogonal to tests based on the mean and variance, and is able to capture effects that are missed by both approaches. We first compare the performances of MANOCCA with existing correlation-based methods and show that MANOCCA is the only test correctly calibrated in simulation mimicking omics data. We then investigate the impact of reducing the dimensionality of the data using principal component analysis when the sample size is smaller than the number of pairwise covariance terms analysed. We show that, in many realistic scenarios, the maximum power can be achieved with a limited number of components. Finally, we apply MANOCCA to 1,000 healthy individuals from the Milieu Interieur cohort, to assess the effect of health, lifestyle and genetic factors on the covariance of two sets of phenotypes, blood biomarkers and flow cytometry-based immune phenotypes. Our analyses identify significant associations between multiple factors and the covariance of both omics data.

bioinformatics↗

bioRxiv2023

Facilitating taxonomy and phylogenetics: An informative and cost-effective protocol integrating long amplicon PCRs and third-generation sequencing

Phylogenetic inference has become a standard technique in integrative taxonomy and systematics, as well as in biogeography and ecology. DNA barcodes are often used for phylogenetic inference, despite being strongly limited due to their low number of informative sites. Also, because current DNA barcodes are based on a fraction of a single, fast-evolving gene, they are highly unsuitable for resolving deeper phylogenetic relationships due to saturation. In recent years, methods that analyse hundreds and thousands of loci at once have improved the resolution of the Tree of Life, but these methods require resources, experience and molecular laboratories that most taxonomists do not have. This paper introduces a PCR-based protocol that produces long amplicons of both slow- and fast-evolving unlinked mitochondrial and nuclear gene regions, which can be sequenced by the affordable and portable ONT MinION platform with low infrastructure or funding requirements. As a proof of concept, we inferred a phylogeny of a sample of 63 spider species from 20 families using our proposed protocol. The results were overall consistent with the results from approaches based on hundreds and thousands of loci, while requiring just a fraction of the cost and labour of such approaches, making our protocol accessible to taxonomists worldwide. HighlightsO_LIDNA barcoding is an invaluable tool for fast and accurate taxonomic classification C_LIO_LIExisting DNA barcodes are still insufficient for obtaining well-supported phylogenies C_LIO_LIWe present a protocol that produces long amplicons of unlinked loci for spiders C_LIO_LIAmplicons are sequenced at very low cost per specimen with ONT MinION C_LIO_LIOur recovered phylogeny is largely consistent with that of high-cost approaches. C_LI

evolutionary biology↗

bioRxiv2023

Spatially-resolved metabolomic identifies colibactin-specific principles of reprogrammed lipid metabolism to promote cancer progression.

Intratumoral bacteria locally contribute to cellular and molecular tumor heterogeneity that support cancer stemness through poorly understood mechanisms. This study aims to explore how Colibactin-producing Escherichia coli (CoPEC) flexibly alters the tumor microenvironment in right-sided colorectal cancer (CRC). Metabolomic and transcriptomic spatial profiling uncovered that CoPEC colonization establishes a high-glycerophospholipid microenvironment within the tumor that is conducive to exhaustion of infiltrated CD8+ T cell and has a lowered prognostic value in right-sided CRC. Mechanistically, the accumulation of lipid droplets in infected cancer cells relied on the production of colibactin as a measure to limit genotoxic stress and supply with sufficient energy for sustaining cell survival and lowering tumor immunogenicity. Specifically, a heightened phosphatidylcholine remodeling of CoPEC-infected cancer cells by the enzyme of the Lands cycle coincided with a lowered accumulation of proapoptotic ceramide and lysophosphatidylcholine. Consequently, a reduced infiltration of CD8+ T lymphocytes that produce the cytotoxic cytokines IFN-{gamma} was found where invading bacteria have been geolocated. By contrast, such an immunosuppressive dysmetabolic process was not observed when human colon cancer cells were infected with the mutant strain that did not produce colibactin (11G5{delta}ClbQ). This work revealed an unexpected property of CoPEC on lipid overload within tumors that could locally provide an inflammatory environment leading to immunosuppressive mechanisms and tumor expansion. This may pave the way for improving chemoresistance and subsequently outcome of CRC patients who are colonized by CoPEC.

cancer biology↗

bioRxiv2022

Multiple paths towards repeated phenotypic evolution in the spiny-leg adaptive radiation (Tetragnatha; Hawaii)

The repeated evolution of phenotypes is ubiquitous in nature and offers some of the clearest evidence of the role of natural selection in evolution. The genomic basis of repeated phenotypic evolution is often complex and can arise from a combination of gene flow, shared ancestral polymorphism and de novo mutation. Here, we investigate the genomic basis of repeated ecomorph evolution in the adaptive radiation of the Hawaiian spiny-leg Tetragnatha. This radiation comprises four ecomorphs that are microhabitat-specialists, and differ in body pigmentation and size (Green, Large Brown, Maroon, and Small Brown). Using 76 newly generated low-coverage, whole-genome resequencing samples, coupled with population genomic and phylogenomic tools, we studied the evolutionary history of the radiation to understand the evolution of the spiny-leg lineage and the genetic underpinnings of ecomorph evolution. Congruent with previous works, we find that each ecomorph has evolved twice, with the exception of the Small Brown ecomorph, which has evolved three times. The evolution of the Maroon and the Small Brown ecomorphs likely involved ancestral hybridization events, whereas the Green and the Large Brown ecomorphs likely evolved because of either standing genetic variation or de novo mutation. Pairwise comparisons of ecomorphs based on the fixation index (FST) show that divergent genomic regions include genes with functions associated with pigmentation (melanization), learning, neuronal and synapse activity, and circadian rhythms. These results show that the repeated evolution of ecomorphs in the Hawaiian spiny-leg Tetragnatha is linked to multiple genomic regions and suggests a previously unknown role of learning and circadian rhythms in ecomorph.

evolutionary biology↗

bioRxiv2022

Optimizing bioink composition for human chondrocyte expression of lubricin

The surface zone of articular cartilage is the first area impacted by cartilage defects, commonly resulting in osteoarthritis. Chondrocytes in the surface zone of articular cartilage synthesize and secrete lubricin, a proteoglycan that functions as a lubricant protecting the deeper layers from shear stress. 3D bioprinting is a tissue engineering technique that uses cells encapsulated in biomaterials to fabricate 3D constructs. Gelatin methacrylate (GelMA) is a frequently used biomaterial for 3D bioprinting cartilage. Oxidized methacrylated alginate (OMA) is a chemically modified alginate designed for its tunable degradation rate and mechanical properties. To determine an optimal combination of GelMA and OMA for lubricin expression, we used our novel high-throughput human articular chondrocyte reporter system. Primary human chondrocytes were transduced with PRG4 (lubricin) promoter-driven Gaussia luciferase, allowing for temporal assessment of lubricin expression. A lubricin expression driven Design of Experiment screen and subsequent validation identified 14% GelMA/2% OMA for further study. Therefore, 14% GelMA/2% OMA, 14% GelMA and 16% GelMA were 3D bioprinted. The combination of lubricin protein expression and shape retention over the 22 days in culture, determined the 14% GelMA/2%OMA to be the optimal formulation for lubricin secretion.

bioengineering↗

bioRxiv2022

Correction of age-associated defects in dendritic cell functions enables CD4+ T cells to eradicate tumors in the elderly

Defective host defenses later in life are associated with changes in immune system activity. The means to correct immune defects to ensure immunity in the elderly are undefined. In this study, we found that CD8+ T cells, which are necessary for anti-tumor immunity in young mice, are not required to eradicate the same cancers later in life. Rather, CD4+ T cells drive anti-tumor immunity in elderly mice. The generation of anti-tumor CD4+ T cells requires multiple dendritic cell (DC) activities that are elicited by immune agonists known as hyperactivators, but not by adjuvants that model those used clinically. DC hyperactivators correct age-associated defects in DC migration and T cell co-stimulation while enabling NLRP3 inflammasome activities within living cells. These combined activities enable DCs to induce TH1-skewed T cells that persist into old age and eliminate implantable tumors. These results raise the possibility of correcting age-associated immune defects through DC manipulation.

immunology↗

bioRxiv2022

DNA repair strategy sets transposon mobilization rates in Caenorhabditis elegans

Transposons are parasitic nucleic acids that threaten genome integrity in all cells. In the metazoan model organism Caenorhabditis elegans, DNA transposons are active in the soma where they are reported to exhibit mobilization rates {cong}1000 fold higher than in germ cells. How and why DNA transposons might be so highly active in the C. elegans soma is a mystery. To better understand this question, we constructed reporter genes that label cells in which Tc1 has mobilized with fluorescent protein. The reporters recapitulate the known properties of DNA transposons in C. elegans and allow transposon activity to be monitored in intact, living animals. Using these reporters, we identify cytoplasmic and nuclear factors that limit transposition in the germline. Interestingly, none of these factors limit transposition in the soma. Rather, we identify a gene (nhj-1/scb-1), which we show is required for 99.9% of Tc1 mobilization events in somatic tissues, but does not influence mobilization in the germline. nhj-1/scb-1 encodes a nematode-specific component of the non-homologous end joining (NHEJ) DNA repair machinery. Mutations in the other components of the NHEJ machinery (cku-70, cku-80, and lig-4) also suppress Tc1 mobilization in the C. elegans soma by {cong}1000 fold. The data show that the use of NHEJ to repair transposon-induced DNA breaks in the soma dramatically increases the rate of transposon mobilization in this tissue. And because C. elegans germ cells use homology-based repair, and not NHEJ, to fix transposon-induced breaks, we propose that the 1000-fold difference in transposon mobility reported for the C. elegans soma and germline can, in large part, be explained by tissue-specific differences in DNA repair strategy. Author SummaryTransposons are common parasitic genetic elements that threaten all genomes. For example, half of the human genome is made up of transposons. Transposon mobilization can disrupt gene function, causing disease, so transposon activity needs to be tightly regulated to prevent harm to the host. Transposons are typically less active in the soma than in the germline, because somatic transposition benefits neither host or transposon. Surprisingly, in the nematode model organism Caenorhabditis elegans, transposons are reported to be 1000-fold more active in the soma than the germline. Here, we develop a system to investigate transposon regulation in an intact live animal, and show that, in large part, tissue-specific differences in transposon activity in C. elegans is due to the use of different DNA repair pathways by these tissues, highlighting the importance of DNA repair strategy in determining outcomes of transposon excision events. Given that DNA repair factors have been linked to transposon regulation in other eukaryotes, we propose that DNA repair choice likely contributes to transposon mobilization in all eukaryotes.

genetics↗

bioRxiv2021

An Atypical RNA Quadruplex Marks RNAs as Vectors for Gene Silencing

The addition of poly(UG) ("pUG") repeats to 3' termini of mRNAs drives gene silencing and trans-generational epigenetic inheritance in the metazoan C. elegans.1 pUG tails promote silencing by recruiting an RNA-dependent RNA Polymerase (RdRP) that synthesizes small interfering (si)RNAs.1 Here we show that active pUG tails require a minimum of 11.5 repeats and adopt a quadruplex (G4)2 structure we term the pUG fold. The pUG fold differs from known G4s in that it has a left-handed backbone similar to Z-RNA3,4, no consecutive guanosines in its sequence, and three G quartets and one U quartet stacked non-sequentially. Its biological importance is emphasized by our observations that porphyrin molecules bind to the pUG fold and inhibit both gene silencing and binding of RdRP. Moreover, specific N7-deaza RNA substitutions that do not adopt the pUG fold neither bind RdRP nor induce RNA silencing. These data define the pUG fold as a previously unrecognized RNA secondary structure motif that drives gene silencing. The pUG fold can also form internally within larger RNA molecules. Approximately 20,000 pUG-fold sequences are found in non-coding regions of human RNAs, suggesting the fold likely has biological roles beyond gene silencing.

molecular biology↗

bioRxiv2021

Phenogenomic characterization of a newly domesticated and novel species from the genus Verrucosispora

The concept of bacterial dark matter stems from our inability to culture most microbes and represents a fundamental hole in our knowledge of microbial diversity. Herein we present the domestication of such an organism: a previously uncultured, novel species from the rare-Actinomycetes genus Verrucosispora. Although initial recovery took >4 months, isolation of phenotypically distinct, domesticated generations occurred within weeks. Two isolates were subjected to phenogenomic analyses, revealing domestication correlated with enhanced growth rates in nutrient-rich media, but diminished capacity to metabolize diverse amino acids. This is seemingly mediated by genomic decay through the pseudogenization of amino acids metabolism genes. Conversely, later generational strains had enhanced spore germination rates, potentially through the reversion of a sporulation-associated kinase from pseudogene to true gene status. We observed that our most wild-type isolate had the greatest potential for antibacterial activity, which correlated with extensive mutational attrition of biosynthetic gene clusters in domesticated strains. Comparative analyses revealed wholesale genomic reordering in strains, with widespread SNP, indel and pseudogene mutations observed. We hypothesize that domestication of this previously unculturable organism resulted from the shedding of genomic flexibility required for life in a dynamic marine environment, parsing out genetic redundancy to allow for a newfound cultivable amenability.

microbiology↗