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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.
- Tubulin acetylation and the cellular mechanosensing and stress responsePublication . Carmona, Bruno; Delgado, Inês L. S.; Nolasco, Sofia; Marques, Rita; Gonçalves, João; Soares, Helena; Halasa, M.; Wawruszak, A.Microtubule (MT) acetylation has emerged as a critical regulator of cellular stress responses, integrating mechanical and oxidative stimuli to support cellular adaptability and survival. This post-translational modification (PTM) enhances MT flexibility and resilience, enabling cells to withstand mechanical challenges such as changes in extracellular matrix stiffness and applied forces. Through its impact on MT physical properties, acetylation minimizes cytoskeletal breakage, reducing the need for constant remodeling and supporting cellular integrity under mechanical stress. Furthermore, tubulin acetylation regulates intracellular trafficking by modulating interactions with molecular motors, allowing for efficient cargo transport and precise spatial organization without disrupting the MT network. In the context of oxidative stress, tubulin acetylation responds to redox imbalances by stabilizing MTs and influencing cellular pathways that regulate reactive oxygen species (ROS). This modification is linked to enhanced antioxidant responses, autophagy regulation, and mitochondrial dynamics, highlighting its role in maintaining cellular homeostasis under oxidative conditions. The dual function of tubulin acetylation, responding to and integrating signals from mechanical and oxidative stress, acts as a bridging mechanism between physical and chemical signaling pathways. Consequently, it has the potential to be a therapeutic target in diseases characterized by dysregulated stress responses, including neurodegenerative disorders, cancer, and cardiovascular conditions. Despite significant progress having been made, unanswered questions persist, particularly regarding the molecular mechanisms by which acetylated MTs encode spatial and functional information and their interplay with other tubulin PTMs.
- Tubulin acetylation: a critical regulator of microtubule functionPublication . Delgado, Inês L.; Carmona, Bruno; Nolasco, Sofia; Marques, Rita; Gonçalves, João; Soares, Helena; Halasa, M.; Wawruszak, A.The cytoskeleton is conserved throughout the eukaryotic lineage and consists of a complex dynamic network mainly composed of three distinct polymers: microtubules (MTs), actin filaments, and intermediate filaments. MTs are polymers of α/β-tubulin heterodimers, playing a myriad of distinct cellular functions, and are the main components of complex structures like the mitotic spindle, cilia, and centrioles. Post-translational modifications (PTMs) regulate the function and increase the complexity of the α/β-tubulin heterodimer pools. One of the PTMs that has been extensively studied is the acetylation of lysine 40 (K40) on α-tubulin, which specifically occurs inside the MT lumen. Acetylation plays a crucial role in controlling the stability and function of MTs in response to signals from within and outside the cell. It impacts the cytoplasm's 3D arrangement and important cellular activities like intracellular transport, cell division, polarity, and migration. Recent research has also emphasized the significance of this PTM in regulating the mechanical properties of MTs and cellular sensing. The levels and activity of MT acetyltransferases and deacetylases are tightly regulated through various transcriptional, post-transcriptional, and post-translational mechanisms, including miRNAs, phosphorylation, protein-protein interactions, and regulated localization between the nucleus and cytoplasm. These regulatory processes involve components of diverse signaling pathways, and their deregulation has been implicated in numerous diseases, including neurological disorders, cancer, and cardiac conditions.