Biology subjects

Willemsen, A.

Publications and source records attributed to Willemsen, A..

8 recordsLinked to original sources

bioRxiv2026

A multilayered network reveals the centrality of newly discovered Nucleocytoviricota in wastewater treatment plant communities

Viruses of the phylum Nucleocytoviricota are paradigm-shifting entities due to their exceptionally large genomes and complex gene repertoires, which blur the lines between viral and cellular life. Previous research has leveraged computational approaches to map their extensive diversity, while experimental work has started to elucidate the intricate networks they form with hosts, bacterial and other symbionts, co-infecting virophages and other mobile genetic elements. Here, we analyzed deeply sequenced metagenomes sampled from wastewater treatment plants in Denmark, an environment with rapid abiotic changes and known to be a hotbed of dense microbial communities. We discovered 61 novel nucleocytoviruses, 15 virophages and 14 polinton-like viruses. By integrating them with microbial contigs into a multilayered interaction network, we explore the role of these entities on a mesocosm scale. We demonstrate the centrality of nucleocytoviruses, positioning them as important players shaping microbial community structure and evolution in wastewater treatment plants.

microbiology↗

bioRxiv2025

Giant virus transcription and translation occur at well-defined locations within amoeba host cells

Many giant viruses replicate in the cytoplasm in so-called viral factories. How exactly these viral factories are established and where the different steps of the replication cycle occur remains largely obscure. We have developed a single-molecule messenger RNA fluorescence in situ hybridisation (smFISH) protocol for giant viruses in an Acanthamoeba host. Combined with other labelling techniques (FUNCAT, DiD, rRNA FISH, DAPI), we show the Mimivirus transcription and translation sites during an infection cycle in the amoeba host cell. While viral mRNA localisation changes depending on the infection stage, transcription occurs at well-defined spots within the viral factory. The original viral cores released within the cytoplasm most likely define these spots. When transported outside of the viral factory, the translation of viral mRNA takes place in a well-defined ring surrounding it. With this study, we obtained novel insights into giant virus replication, of which the methods are widely applicable to other viruses for the visualisation and quantification of RNA molecules.

microbiology↗

bioRxiv2024

Giant virus creates subcellular environment to overcome codon-tRNA mismatch

Codon usage consonant with the cellular tRNA pool is important for efficient translation. However, many eukaryotic viruses, including amoeba-infecting mimiviruses, have codon usage that is largely deviated from that of their host, despite using the host machinery for translation. This raises the question of how these viruses cope with the mismatch between tRNA supply and demand. Here we show that Acanthamoeba castellanii mimivirus generates a subcellular area in the host cells to translate virus mRNAs. A combination of genome-wide ribosome profiling and RNA sequencing showed that ribosomes traversed along viral mRNAs with fewer pausing events than were observed on amoeba mRNAs. Frequently used codons in viral mRNAs had higher tRNA accessibility than the same type of codons in amoeba mRNAs. tRNA sequencing showed that the tRNA pool was not greatly altered during the infection even though the virus encodes tRNA genes. Instead, by in situ labelling, we found that viral mRNAs and newly synthesized proteins were localized at the periphery region of the viral factory, likely creating a unique environment to facilitate viral translation. Our data provide a perspective on how local translation assists the virus in overcoming the mismatch between tRNA supply and demand.

microbiology↗

bioRxiv2024

Novel high-quality amoeba genomes reveal widespread codon usage mismatch between giant viruses and their hosts

The need for high-quality protist genomes has prevented in-depth computational and experimental studies of giant virus-host interactions. In addition, our current knowledge of host range is highly biased due to the few hosts used to isolate novel giant viruses. This study presents six high-quality amoeba genomes from known and potential giant virus hosts belonging to two distinct eukaryotic clades: Amoebozoa and Discoba. We employ their genomic data to investigate the predictability of giant virus host range. Using a combination of long and short-read sequencing, we obtained highly contiguous and complete genomes of Acanthamoeba castellanii, Acanthamoeba griffini, Acanthamoeba terricola, Naegleria clarki, Vermamoeba vermiformis, and Willaertia magna, contributing to the collection of sequences for the eukaryotic tree of life. We found that the six amoebae have distinct codon usage patterns and that, contrary to other virus groups, giant viruses often have different and even opposite codon usage with their known hosts. Conversely, giant viruses with matching codon usage are frequently not known to infect or replicate in these hosts. Interestingly, analyses of integrated viral sequences in the amoeba host genomes reveal potential novel virus-host associations. Matching of codon usage preferences is often used to predict virus-host pairs. However, with the broad-scale analyses performed in this study, we demonstrate that codon usage alone appears to be a poor predictor of giant virus host range. We discuss the potential strategies that giant viruses employ to ensure high viral fitness in non-matching hosts. Moreover, this study emphasises the need for more high-quality protist genomes. Finally, the amoeba genomes presented in this study set the stage for future experimental studies to better understand how giant viruses interact with different host species.

evolutionary biology↗

bioRxiv2022

The non-neutral, multi-level, phenotypic impact of synonymous mutations revealed through heterologous gene expression in human cells.

Differences in codon frequency between genomes, genes, or positions along a gene, modulate transcription and translation efficiency, leading to phenotypic and functional differences. Here, we present a multiscale analysis of the effects of synonymous codon recoding during heterologous gene expression in human cells, quantifying the phenotypic consequences of codon usage bias at different molecular and cellular levels, with an emphasis on translation elongation. Six synonymous versions of an antibiotic resistance gene were generated, fused to a fluorescent reporter, and independently expressed in HEK293 cells. Multiscale phenotype was analysed by means of: quantitative transcriptome and proteome assessment, as proxies for gene expression; cellular fluorescence, as a proxy for single-cell level expression; and real-time cell proliferation in absence or presence of antibiotic, as a proxy for the cell fitness. We show that differences in codon usage bias strongly impact the molecular and cellular phenotype: (i) they result in large differences in mRNA levels and in protein levels, leading to differences of over fifteen times in translation efficiency; (ii) they introduce unpredicted splicing events; (iii) they lead to reproducible phenotypic heterogeneity; and (iv) they lead to a trade-off between the benefit of antibiotic resistance and the burden of heterologous expression. In human cells in culture, codon usage bias modulates gene expression by modifying mRNA availability and suitability for translation, leading to differences in protein levels and eventually eliciting functional phenotypic changes. IMPORTANCESynonymous codons encode for the same amino acid, but they are not neutral regarding gene expression or protein synthesis. Bias between synonymous codons have evolved naturally and are also applied in biotechnology protein production. We have studied the multilevel impact of codon usage on a human cell system. We show that differences in codon usage lead to transcriptomic, proteomic and functional changes, modulating gene expression and cellular phenotype.

systems biology↗

bioRxiv2020

Ecological opportunity as a driving force of radiation events and time-dependent evolutionary rates in papillomaviruses

Papillomaviruses (PVs) have a wide host range, infecting mammals, birds, turtles, and snakes. The recent discovery of PVs in different fish species allows for a more complete reconstruction of the evolutionary history of this viral family. In this study we perform phylogenetic dating to analyse evolutionary events that occurred during PV evolution, as well as to estimate speciation and evolutionary rates. We have used four different data sets to explore and correct for potential biases that particular taxa combinations may introduce during molecular time inference. When considering the evolution of substitution rates we observed that short-term rate estimates are much higher than long-term rate estimates, also known as the time-dependent rate phenomenon. When considering the evolution of viral branching events (as a proxy for speciation rates), we show that these have not been constant through time, suggesting the occurrence of distinct evolutionary events such as adaptive radiations and/or changes in the available host niches. In a joint analysis with host speciation rates, we identified at least four different evolutionary periods, suggesting that the evolution of PVs has been multiphasic, and thus refining the previously suggested biphasic evolutionary scenario. Thanks to the discovery of novel PVs in basal hosts and to the implementation of a time-dependent rate model for molecular dating, our results provide new insights into the evolutionary history of PVs. In this updated evolutionary scenario, ecological opportunity appears as one main driving force for the different radiation and key-innovation events we observe.

evolutionary biology↗

bioRxiv2018

Origin and evolution of papillomavirus (onco)genes and genomes

Papillomaviruses (PVs) are ancient viruses infecting vertebrates, from fish to mammals. Although the genomes of PVs are small and show conserved synteny, PVs display large genotypic diversity and ample variation in the phenotypic presentation of the infection. Most PVs genomes contain two small early genes E6 and E7. In a bunch of closely related human PVs, the E6 and E7 proteins provide the viruses with oncogenic potential.\n\nThe recent discoveries of PVs without E6 and E7 in different fish species place a new root on the PV tree, and suggest that the ancestral PV consisted of the minimal PV backbone E1-E2-L2-L1.\n\nBayesian phylogenetic analyses date the most recent common ancestor of the PV backbone to 424 million years ago (Ma). Common ancestry tests on extant E6 and E7 genes indicate that they share respectively a common ancestor dating back to at least 184 Ma. In AlphaPVs infecting primates, the appearance of the E5 oncogene 53-58 Ma concurred with i) a significant increase in substitution rate, ii) a basal radiation, and iii) key gain of functions in E6 and E7. This series of events was instrumental to build the extant phenotype of oncogenic human PVs.\n\nOur results assemble the current knowledge on PV diversity and present an ancient evolutionary timeline punctuated by evolutionary innovations in the history of this successful viral family.

evolutionary biology↗

bioRxiv2018

Genome plasticity in Papillomaviruses and de novo emergence of E5 oncogenes

The clinical presentations of papillomavirus (PV) infections come in many different flavors. While most PVs are part of a healthy skin microbiota and are not associated to physical lesions, other PVs cause benign lesions, and only a handful of PVs are associated to malignant transformations linked to the specific activities of the E5, E6 and E7 oncogenes. The functions and origin of E5 remain to be elucidated. These E5 ORFs are present in the genomes of a few polyphyletic PV lineages, located between the early and the late viral gene cassettes. We have computationally assessed whether these E5 ORFs have a common origin and whether they display the properties of a genuine gene. Our results suggest that during the evolution of Papillomaviridae, at least four events lead to the presence of a long non-coding DNA stretch between the E2 and the L2 genes. In three of these events, the novel regions evolved coding capacity, becoming the extant E5 ORFs. We then focused on the evolution of the E5 genes in AlphaPVs infecting humans. The sharp match between the type of E5 protein encoded in AlphaPVs and the infection phenotype (cutaneous warts, genital warts or anogenital cancers) supports the role of E5 in the differential oncogenic potential of these PVs. In our analyses, the best-supported scenario is that the five types of extant E5 proteins within the AlphaPV genomes may not have a common ancestor. However, the chemical similarities between E5s regarding amino acid composition prevent us from confidently rejecting the model of a common origin. Our evolutionary interpretation is that an originally non-coding region entered the genome of the ancestral AlphaPVs. This genetic novelty allowed to explore novel transcription potential, triggering an adaptive radiation that yielded three main viral lineages encoding for different E5 proteins, and that display distinct infection phenotypes. Overall, our results provide an evolutionary scenario for the de novo emergence of viral genes and illustrate the impact of such genotypic novelty in the phenotypic diversity of the viral infections.

evolutionary biology↗