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- Characterization of a MOB1 homolog in the Apicomplexan parasite toxoplasma gondiiPublication . Delgado, Inês L. S.; Tavares, Alexandra; Francisco, Samuel; Santos, Dulce; Coelho, João; Basto, Afonso P.; Zúquete, Sara; Müller, Joachim; Hemphill, Andrew; Meissner, Markus; Soares, Helena; Leitão, Alexandre; Nolasco, S.Monopolar spindle One Binder1 (MOB1) proteins are conserved components of the tumor-suppressing Hippo pathway, regulating cellular processes such as cytokinesis. Apicomplexan parasites present a life cycle that relies on the parasites’ ability to differentiate between stages and regulate their proliferation; thus, Hippo signaling pathways could play an important role in the regulation of the apicomplexan life cycle. Here, we report the identification of one MOB1 protein in the apicomplexan Toxoplasma gondii. To characterize the function of MOB1, we generated gain-of-function transgenic lines with a ligand-controlled destabilization domain, and loss-of-function clonal lines obtained through CRISPR/Cas9 technology. Contrary to what has been characterized in other eukaryotes, MOB1 is not essential for cytokinesis in T. gondii. However, this picture is complex since we found MOB1 localized between the newly individualized daughter nuclei at the end of mitosis. Moreover, we detected a significant delay in the replication of overexpressing tachyzoites, contrasting with increased replication rates in knockout tachyzoites. Finally, using the proximity-biotinylation method, BioID, we identified novel members of the MOB1 interactome, a probable consequence of the observed lack of conservation of some key amino acid residues. Altogether, the results point to a complex evolutionary history of MOB1 roles in apicomplexans, sharing properties with other eukaryotes but also with divergent features, possibly associated with their complex life cycle.
- Tubulin post-translational modifications: the elusive roles of acetylationPublication . Carmona, Bruno; Marinho, H. Susana; Matos, Catarina Lopes; Nolasco, S.; Soares, HelenaMicrotubules (MTs), dynamic polymers of α/β-tubulin heterodimers found in all eukaryotes, are involved in cytoplasm spatial organization, intracellular transport, cell polarity, migration, division, and cilia biology. MTs functional diversity depends on the differential expression of distinct tubulin isotypes and is amplified by a vast number of different post-translational modifications (PTMs). The addition/removal of PTMs to α- or β-tubulins is catalyzed by specific enzymes and allows combinatory patterns largely enriching the distinct biochemical and biophysical properties of MTs, creating a code read by distinct proteins, including microtubule-associated proteins (MAPs), which allow cellular responses. This review is focused on tubulin-acetylation, whose cellular roles continue to generate debate. We travel through the experimental data pointing to α-tubulin Lys40 acetylation role as being an MT stabilizer and a typical PTM of long-lived MTs, to the most recent data, suggesting that Lys40 acetylation enhances MT flexibility and alters the mechanical properties of MTs, preventing MTs from mechanical aging characterized by structural damage. Additionally, we discuss the regulation of tubulin acetyltransferases/desacetylases and their impacts on cell physiology. Finally, we analyze how changes in MT acetylation levels have been found to be a general response to stress and how they are associated with several human pathologies.
- Ageing affects the CD4+ T cell polarization and mucosal tropism induced by TLR2/TLR4-activated dendritic cellsPublication . Zúquete, Sara; Ferreira, Mariana; Delgado, Inês L.; Rosa, Maria teresa; Mendes, Ana Catarina; Santos, Dulce; Nolasco, Sofia; Graça, Luís; Leitão, Alexandre; Basto, Afonso P.Toll-like receptor (TLR)2 activation induces aldehyde dehydrogenase enzymes in non-mucosal dendritic cells (DCs), enabling them to metabolize vitamin A into all-trans retinoic acid, which induces the expression of mucosal-homing molecules (α4β7 and CCR9) in the activated T cells. Recently, we have shown that the simultaneous activation of non-mucosal DCs through TLR2 and TLR4 maintains such capacity while reinforcing the polarization of primed CD4+ T cells towards Th1. Here, we observed that TLR2/TLR4 stimulation of aged DCs leads to the production of less TNFα and more IL-10, and that CD4+ T cells primed by those DCs express lower levels of the mucosal homing receptor CCR9 and produce less type-1 (IFNγ) and more type-2 (IL-4 and IL-13) cytokines. These results emphasize the importance of considering the age-related alterations in DC function when developing novel immunomodulation strategies that rely on the DC-T cell crosstalk through stimulation of pattern recognition receptors.