Biology subjects

Werner, M. S.

Publications and source records attributed to Werner, M. S..

6 recordsLinked to original sources

bioRxiv2024

Developmental transcriptomics in Pristionchus reveals the logic of a plasticity gene regulatory network

Developmental plasticity enables the production of alternative phenotypes in response to different environmental conditions. While significant advances in understanding the ecological and evolutionary implications of plasticity have been made, understanding its genetic basis has lagged. However, a decade of genetic screens in the model nematode Pristionchus pacificus has culminated in 30 genes which affect mouth-form plasticity. We also recently reported the critical window of environmental sensitivity, and therefore have clear expectations for when differential gene expression should matter. Here, we collated previous data into a gene-regulatory network (GRN), and performed developmental transcriptomics across different environmental conditions, genetic backgrounds, and mouth-form mutants to assess the regulatory logic of plasticity. We found that only two genes in the GRN (eud-1 and seud-1/sult-1) are sensitive to the environment during the critical window. Interestingly, the time points of their sensitivity differ, suggesting that they act as sequential checkpoints. We also observed temporal constraint upon the transcriptional effects of mutating the GRN and revealed unexpected feedback between mouth-form genes. Surprisingly, expression of seud-1/sult-1, but not eud-1, correlated with mouth form biases across different strains and species. Finally, a comprehensive analysis of all samples identified metabolism as a shared pathway for regulating mouth-form plasticity. These data are presented in a Shiny app to facilitate gene-expression comparisons across development in up to 14 different conditions. Collectively, our results suggest that mouth-form plasticity evolved a constrained, two-tiered logic to integrate environmental information leading up to the final developmental decision.

genetics↗

bioRxiv2024

Developmental small RNA transcriptomics reveals divergent evolution of the conserved microRNA miR-100 and the let-7-complex in nematodes

BackgroundOver the last three decades, small RNAs have emerged as one of the key post-transcriptional regulators, and are categorised into microRNAs (miRNAs), small interfering RNAs (siRNAs), and PIWI-interacting RNAs (piRNAs). Since their discovery as regulators of developmental timing in the nematode Caenorhabditis elegans, some miRNAs have been found to be highly conserved throughout animal evolution. miR-100 is one such highly conserved miRNA, possibly predating the origin of bilaterians. Furthermore, mir-100 is a member of the conserved let-7-complex, a locus containing three conserved miRNA-coding genes (mir-100, let-7, and mir-125/lin-4) that does not exist in the same arrangement in C. elegans. ResultsWe performed small RNA sequencing across four developmental time-points in the satellite nematode Pristionchus pacificus, identifying 64 novel and ontogenically specifically expressed miRNAs. We also identified miR-100 as the most abundant miRNA in post-embryonic juvenile stages. Additionally, P. pacificus exhibited a novel constellation of the let-7-complex, wherein mir-100 is closely linked to let-7, but without mir-125/lin-4 in the same locus. Mutants of mir-100 and let-7, individually as well as simultaneously, resulted in viable but developmentally defective worms, showing misregulation in collagen and astacin gene families. Interestingly, lin-4 mutants showed a delayed but otherwise normal development. ConclusionThis study provides the first developmental small RNA transcriptome of P. pacificus, providing a community resource for future evo-devo studies. Furthermore, we identify miR-100 as the most abundant ontogenically-specific miRNA, which, along with let-7, regulates normal development in P. pacificus. Lastly, we observe two alternative arrangements of the highly conserved miRNA locus let-7-complex.

evolutionary biology↗

bioRxiv2023

Characterization of the Pristionchus pacificus epigenetic toolkit reveals the evolutionary loss of the histone methyltransferase complex PRC2

Comparative approaches have revealed both divergent and convergent paths to achieving shared developmental outcomes. Thus, only through assembling multiple case studies can we understand biological principles. Yet, despite appreciating the conservation - or lack thereof - of developmental networks, the conservation of epigenetic mechanisms regulating these networks is poorly understood. The nematode Pristionchus pacificus has emerged as a model system of plasticity and epigenetic regulation as it exhibits a bacterivorous or omnivorous morph depending on its environment. Here, we determined the "epigenetic toolkit" available to P. pacificus as a resource for future functional work on plasticity, and as a comparison with C. elegans to investigate the conservation of epigenetic mechanisms. Broadly, we observed a similar cast of genes with putative epigenetic function between C. elegans and P. pacificus. However, we also found striking differences. Most notably, the histone methyltransferase complex PRC2 appears to be missing in P. pacificus. We described the deletion/pseudogenization of the PRC2 genes mes-2 and mes-6 and concluded that both were lost in the last common ancestor of P. pacificus and a related species P. arcanus. Interestingly, we observed the enzymatic product of PRC2 (H3K27me3) by mass spectrometry and immunofluorescence, suggesting that a currently unknown methyltransferase has been co-opted for heterochromatin silencing. Altogether, we have provided an inventory of epigenetic genes in P. pacificus to enable reverse-genetic experiments related to plasticity, and in doing so have described the first loss of PRC2 in a multicellular organism.

genetics↗

bioRxiv2020

Development of a Synthetic Poxvirus-Based SARS-CoV-2 Vaccine

Modified Vaccinia Ankara (MVA) is a highly attenuated poxvirus vector that is widely used to develop vaccines for infectious diseases and cancer. We developed a novel vaccine platform based on a unique three-plasmid system to efficiently generate recombinant MVA vectors from chemically synthesized DNA. In response to the ongoing global pandemic caused by SARS coronavirus-2 (SARS-CoV-2), we used this novel vaccine platform to rapidly produce fully synthetic MVA (sMVA) vectors co-expressing SARS-CoV-2 spike and nucleocapsid antigens, two immunodominant antigens implicated in protective immunity. Mice immunized with these sMVA vectors developed robust SARS-CoV-2 antigen-specific humoral and cellular immune responses, including potent neutralizing antibodies. These results demonstrate the potential of a novel vaccine platform based on synthetic DNA to efficiently generate recombinant MVA vectors and to rapidly develop a multi-antigenic poxvirus-based SARS-CoV-2 vaccine candidate.Competing Interest StatementThe authors have declared no competing interest.View Full Text

immunology↗

bioRxiv2019

WDR76 promotes MLL-rearranged leukemia via selective recognition of 5-hydroxymethylcytosine in DNA

Although rare, the distribution of the 5-hydroxymethylcytosine (hmC) modification in mammalian DNA is tissue- and gene-specific, yet distinct from its transcriptionally-repressive methylcytosine (mC) precursor, suggesting unique signaling potential. To examine this possibility, we fractionated mammalian brain extracts to discover binding partners specific for oxidized states of mC. We demonstrate that one such factor, WDR76, is a highly hmC-specific binding protein that modulates gene expression within chromosomal regions enriched in hmC where it binds. We demonstrate direct transcriptional activation of several target genes in mouse embryonic stem cells as a function of hmC levels and contingent upon WDR76. In human cell lines and mouse models, WDR76 recruitment by hmC is critical for the initiation and maintenance of MLL-rearranged leukemias. Beyond its canonical role as an intermediate in mC remediation, we show that hmC can be an epigenetic mark whose recognition drives leukemogenesis, portending analogous signaling pathways for other rare DNA modifications.

biochemistry↗

bioRxiv2018

Adult-specific pheromones differently influence malthusian escape strategies of younger generations

Many animal and plant species respond to high or low population densities by phenotypic plasticity. To investigate if specific age classes and/or cross-generational signaling affect(s) phenotypic plasticity, we developed a dye-based method to differentiate co-occurring nematode age classes. We applied this method to Pristionchus pacificus, which develops a predatory mouth form to exploit alternative resources and kill competitors in response to high population densities. Remarkably, only adult, but not juvenile, crowding induces the predatory morph in other juveniles. Profiling of secreted metabolites throughout development with HPLC-MS combined with genetic mutants traced this result to the production of adult-specific pheromones. Specifically, the P. pacificus-specific di-ascaroside#1 that induces the predatory morph exhibits a binary induction in adults, even though mouth form is no longer plastic in adults. This cross-generational signaling between adults and juveniles may serve as an indication of rapidly increasing population size. Thus, phenotypic plasticity depends on critical age classes.

ecology↗