Simonsen, H. T. | Search bioRxiv

bioRxiv2025

Functional Characterization of Target of Rapamycin (TOR) Signalling in Physcomitrella

Target of rapamycin (TOR) is a conserved protein kinase and an important signalling hub in eukaryotes. The moss Physcomitrella (Physcomitrium patens) is a model organism for plant physiology, development, and evolution. However, little is known about TOR signalling in non-vascular plants, including Physcomitrella. Here, we report the effects of inhibiting TOR signalling in Physcomitrella. We identified and characterized Physcomitrella 12-kD FK506-binding protein (FKBP12), which binds TOR in the presence of rapamycin. Whereas the growth of wild-type protonema is unaffected by rapamycin, overexpressing endogenous FKBP12 rendered the plant susceptible to the inhibitor in a dose-dependent manner. Likewise, protonema growth was inhibited when the TOR-specific inhibitor AZD8055 was present in the culture. We show that rapamycin and AZD8055 have pleiotropic effects, as they delay cell cycle progression and development, induce chlorosis, inhibit photosynthesis, and alter total protein content. Additionally, we identified and characterized PpTOR, PpLST8, and PpRAPTOR, key components of TOR complex 1 (TORC1), whereas RICTOR and mSIN1 of TORC2 are not encoded by the P. patens genome. We found that PpLST8 substitutes its homolog in yeast to allow cell growth. Physcomitrella mutants were generated with a conditional downregulation of PpTOR, PpLST8, and PpRAPTOR, respectively. They were impaired in growth. Finally, we show TOR-dependent phosphorylation of a well-known TOR phospho-target, the ribosomal protein RPS6, via LC-MS/MS. Collectively, our results show that Physcomitrella growth and development is positively controlled by a conserved TOR kinase. We suggest to further dissect TOR signalling in Physcomitrella in order to elucidate signalling integration via TORC1 in plants. Key messageThe evolutionary conserved TOR kinase positively controls growth and development of the moss Physcomitrella, development and function of its chloroplasts, its protein synthesis and cell cycle progression.

plant biology↗

bioRxiv2022

Dietary non-starch polysaccharides impair immunity to enteric nematode infection

The influence of diet on immune function and resistance to enteric infection and disease is becoming ever more established. Highly processed, refined diets can lead to inflammation and gut microbiome dysbiosis, whilst health-promoting dietary components such as phytonutrients and fermentable fibres are thought to promote a healthy microbiome and balanced mucosal immunity. Chicory (Cichorium intybus) is a leafy green vegetable rich in fibres and bioactive compounds that may promote gut health. Unexpectedly, we here show that incorporation of chicory into semisynthetic AIN93G diets renders mice susceptible to infection with enteric helminths. Mice fed a high level of chicory leaves (10% dry matter) had a more diverse gut microbiota, but a diminished type-2 immune response to infection with the intestinal roundworm Heligmosomoides polygyrus. Furthermore, the chicory-supplemented diet significantly increased burdens of the caecum-dwelling whipworm Trichuris muris, concomitant with a highly skewed type-1 immune environment in caecal tissue. The chicory-supplemented diet was rich in non-starch polysaccharides, particularly uronic acids (the monomeric constituents of pectin). In accordance, mice fed pectin-supplemented AIN93G diets had higher T. muris burdens and reduced IgE production and expression of genes involved in type-2 immunity. Importantly, treatment of pectin-fed mice with exogenous IL-25 restored type-2 responses and was sufficient to allow T. muris expulsion. Collectively, our data suggest that increasing levels of fermentable, non-starch polysaccharides in refined diets compromises immunity to helminth infection in mice. This diet-infection interaction may inform new strategies for manipulating the gut environment to promote resistance to enteric parasites.

immunology↗