Browsing by Author "Lince-Faria, Mariana"
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- Centrosome positioning and development of ciliopathies: role of the human centrosomal protein TBCCD1Publication . Carmona, Bruno; Camelo, Carolina; Mehraz, Manon; Lemullois, Michel; Ferreira, David C.; Nolasco, Sofia; Lince-Faria, Mariana; Marinho, H. Susana; Bettencourt-Dias, Mónica; Tassin, Anne-Marie; Soares, HelenaAims/Context: Primary cilia are specialized microtubule-based signaling organelles that convey extracellular signaling and cellular polarity into a cellular response. Defects in primary cilia assembly/function cause severe diseases known as ciliopathies, typified by clinical manifestations, like infertility, obesity, brain problems, blindness, and kidney cysts. Primary cilia assembly entails centrosome migration to the plasma membrane where a centriole docks, maturates into a basal body (BB), and assembles the cilia axoneme. The human centrosomal TBCCD1 is a critical factor in centrosome positioning previously identified by us. Our aim is to discover the mechanisms/signals required for the correct positioning of the centrosome during cilia assembly, and how these mechanisms, when compromised, are related to ciliopathies. Methods: The proximity-dependent identification (BioID) assay was used to screen for TBCCD1 interactors. Immunofluorescent and super-resolution microscopy, as well as Western blot, were used to study the levels and cellular localization of the identified TBCCD1 interactors in human RPE1 cells overexpressing or depleted of TBCCD1. To study the impact of TBCCD1 knockdown in motile cilia the ciliate Paramecium, containing ∼3,000 motile cilia, was used. Results: Our BioID screen for TBCCD1 interactors identified several well-known proteins encoded by ciliopathy genes, e.g. the centrosomal protein OFD1 involved in the Orofacial-Digital Syndrome. We show that TBCCD1 knockdown and overexpression in RPE1 cells affects OFD1 distribution. Super-resolution microscopy shows TBCCD1 is localized at the distal region of the centrosome and that its depletion dramatically affects the centrosome subdistal protein CEP170, a component of cilia basal feet. In Paramecium, the TBCCD1 knockdown causes abnormal BB-associated structures organization and anomalous BB positioning/anchoring defects. Conclusions: Our data support a role for TBCCD1 in the maintenance of centrosome structure and in BB anchoring at the cell membrane during ciliogenesis. TBCCD1 is emerging as a novel protein with a role in human ciliopathies.
- From centrosomal microtubule anchoring and organization to basal body positioning: TBCCD1 an elusive proteinPublication . Carmona, Bruno; Camelo, Carolina; Mehraz, Manon; Lemullois, Michel; Ferreira, David C.; Nolasco, Sofia; Lince-Faria, Mariana; Marino, H. Susana; Bettencourt-Dias, Mónica; Tassin, Anne-Marie; Koll, France; Soares, HelenaCilia are microtubule-based organelles that protrude from the cell surface and fulfill critical motility and sensory functions being required for normal embryonic development and for homeostasis of human adult tissues. Cilia loss or dysfunction is associated with human ciliopathies. At their base cilia have a centriole/basal body (BB), which can be derived from the centrosome and assembles the ciliary axoneme. This process requires the correct positioning/anchoring of the centrosome’s mother centriole/BB to the cell membrane. A clear picture of the different signals and players involved in centrosome positioning/anchoring is still not available. Published work from our group identified a new centrosomal TBCC domain-containing human protein (TBCCD1) that is involved in centrosome correct positioning and primary cilia assembly. In mammalian cells, TBCCD1 is observed at pericentriolar satellites, in basal bodies of primary and motile cilia and at primary cilia ciliopathy hot domain, the transition zone. Super-resolution microscopy shows that TBCCD1 is localized at the distal region of the centrosome and its depletion dramatically affects the centrosome subdistal protein CEP170, a component of primary and motile cilia basal feet. By doing a proximity-dependent biotin identification (BioID-MS) screen for TBCCD1 interactors several well-known proteins encoded by ciliopathy genes were identified, e.g. the centrosomal proteins OFD1 and Moonraker/KIAA0753 associated with Digital Syndrome 1 and Joubert syndrome, respectively. OFD1 and Moonraker are required for the maintenance of centrosome structure and both proteins localization is dramatically disturbed by TBCCD1 depletion. To clarify the role of human TBCCD1 in cilia biogenesis we used the ciliate Paramecium. Noteworthy, in Paramecium TBCCD1 knockdown causes abnormal basal body associated rootlets organization, anomalous BB positioning/anchoring defects. Our data using human cells and the ciliate Paramecium support a role of TBCCD1 in centrosome structure maintenance and BB anchoring at the cell membrane. The Paramecium phenotypes confirm that TBCCD1 is a new candidate to a ciliopathic gene probably by founding the TBCCD1/Moonraker/OFD1 functional conserved module required for cilia assembly.
- TBCCD1: a new player in the development of ciliopathies?Publication . Carmona, Bruno; Camelo, Carolina; Mehraz, Manon; Lemullois, Michel; Ferreira, David C.; Nolasco, Sofia; Lince-Faria, Mariana; Marinho, H. Susana; Bettencourt-Dias, Mónica; Tassin, Anne-Marie; Koll, France; Soares, HelenaCilia are hair-like appendages, consisting of a microtubule (MT)-based ciliary axoneme, which fulfill critical motility and sensory functions required for normal embryonic development and also for homeostasis of adult tissues. At their base, cilia have a centriole/basal body, which can be derived from the centrosome, and that nucleates the ciliary axoneme. Centrosomes consist of a pair of centrioles surrounded by the pericentriolar matrix that nucleate/organize the cytoskeleton and are implicated in cell migration, adhesion, and polarity, while during mitosis they assist spindle pole formation. Centriolar satellites are cytoplasmic granules that are located and move around the centrosome. These particles are involved in centrosome assembly and primary cilium formation by delivering cytoplasmatic centriolar/centrosomal components to the centrosome. Mutations in genes encoding centrosome and/or centriolar satellite components and regulators lead to various human disorders such as ciliopathies. Ciliopathies are typified by often overlapping clinical manifestations, e.g. infertility, obesity, brain and skeletal developmental problems, blindness and kidney cysts.