Biology subjects

Buescher, J.

Publications and source records attributed to Buescher, J..

3 recordsLinked to original sources

bioRxiv2025

Exogenous dsRNA made accessible to Dicer by two eukaryotic RNA-dependent RNA polymer-ases

Discrimination of self from non-self RNA is a critical requirement for any cell to respond to infections and to maintain cellular integrity. We report novel functions for two RNA-dependent RNA polymerases (RDRs) in Paramecium in the detection of exogenous RNA. In the RNA interference (RNAi) mechanism, RDRs are normally involved in the production of large amounts of secondary siRNAs in response to an initial primary siRNA. To characterize the function of RDRs in context of exogenous RNA recognition, we developed a novel dsRNA delivery system using dextran nanoparticles to deliver heteroduplex dsRNA to cells as food particles, mimicking the natural phagosomal entry pathway. Subsequent small RNA sequencing allows us to dissect siRNAs produced from exogenous RNA or RDR transcripts. Contrary to expectations of how dsRNA-induced silencing is triggered, our data show that Dicer is unable to directly cleave exogenous dsRNA while two RDRs are required for the initial steps of dsRNA-induced RNAi. Paradoxically, these two RDRs must replicate dsRNA to make it available for Dicer cleavage and primary siRNA accumulation. Our data also show that this system works efficiently also with exogenous ssRNA, although RDR2 is dispensable for ssRNA conversion. The function of RDRs in this protist is in contrast to that in animals, plants and fungi and extends the functional diversity of these polymerases. RDR-associated complexes appear to control the entry of food and symbiont-derived RNA into the RNAi machinery, enabling complex RNA interactions with the environment.

genetics↗

bioRxiv2024

PDK-1/S6K and mTORC1 bypass systemic growth restrictions to promote regeneration

Inflammation triggers systemic signals that induce growth restrictions in distant organs, a process well characterized in tumor cachexia. While mechanisms allowing tumors to circumvent these systemic growth restrictions have been established, the physiological processes that overcome inflammation-induced growth restrictions during regeneration remain largely unexplored. In our study, we use a model of tissue inflammation and regeneration in developing Drosophila imaginal discs to dissect key metabolic and signaling adaptations that, in physiological settings, both induce and overcome systemic growth restrictions. We find that expression of eiger, the Drosophila TNF- homolog, induces systemic insulin restriction, as evidenced by reduced expression of dILP2 and dILP5, as well as elevated nuclear dFOXO, and reduced protein translation and proliferation in peripheral tissues. Proliferating cells overcome systemic insulin restriction by upregulating Pdk1, which is both necessary and sufficient to promote phosphorylation of ribosomal protein S6 and protein translation via an Insulin/Akt-independent mechanism. We further demonstrate that JAK/STAT signaling acts upstream to increase Pdk1 levels, delineating a novel JAK/STAT-Pdk1-S6K axis essential for regenerative proliferation. The upregulation of amino acid transporters in the proliferative domain further suggests that regenerating cells preferentially import amino acids, fueling mTORC1 activation. Similar metabolic signatures are observed in a Drosophila RasV12, scrib tumor model, suggesting that tumors may co-opt these metabolic pathways to sustain growth in an insulin-restricted environment. Our findings reveal a specialized metabolic program that integrates systemic nutrient mobilization with local metabolic reprogramming, with important implications for understanding of physiological tissue repair but also pathologies such as chronic wounds and cancer.

developmental biology↗

bioRxiv2019

Tissue ingrowth markedly reduces mechanical anisotropy and stiffness in fibre direction of highly aligned electrospun polyurethane scaffolds

PurposeThe lack of long-term patency of synthetic vascular grafts currently available on the market has directed research towards improving the performance of small diameter grafts. Improved radial compliance matching and tissue ingrowth into the graft scaffold are amongst the main goals for an ideal vascular graft. MethodsBiostable polyurethane scaffolds were manufactured by electrospinning and implanted in subcutaneous and circulatory positions in the rat for 7, 14 and 28 days. Scaffold morphology, tissue ingrowth, and mechanical properties of the scaffolds were assessed before implantation and after retrieval. ResultsTissue ingrowth after 24 days was 96.5 {+/-} 2.3% in the subcutaneous implants and 77.8 {+/-} 5.4% in the circulatory implants. Over the 24 days implantation, the elastic modulus at 12% strain decreased by 59% in direction of the fibre alignment whereas it increased by 1379% transverse to the fibre alignment of the highly aligned scaffold of the subcutaneous implants. The lesser aligned scaffold of the circulatory graft implants exhibited an increase of the elastic modulus at 12% strain by 77% in circumferential direction. ConclusionBased on the observations, it is proposed that the mechanism underlying the softening of the highly aligned scaffold in the predominant fibre direction is associated with scaffold compaction and local displacement of fibres by the newly formed tissue. The stiffening of the scaffold, observed transverse to highly aligned fibres and for more a random fibre distribution, represents the actual mechanical contribution of the tissue that developed in the scaffold.

bioengineering↗