Encyclopedia of Signaling Molecules

2018 Edition
| Editors: Sangdun Choi


  • João Gonçalves
  • Helena Soares
Reference work entry
DOI: https://doi.org/10.1007/978-3-319-67199-4_551


Historical Background

The major microtubule-organizing center in animal cells is the centrosome which consists of a pair of centrioles surrounded by the pericentriolar matrix. In interphase cells, centrosomes nucleate and organize the microtubule cytoskeleton and are usually maintained at the cell center in close association with the nucleus (Bettencourt-Dias and Glover 2007). This association constitutes a primordial axis of cytoplasmic compartment organization and organelle positioning (e.g., ER and Golgi apparatus). Centrosomes are also involved in mitotic spindle formation, and an aberrant centrosome number has been linked to multipolar spindles and to cancer cells. In the last years, the ability of centrosomes to become basal bodies and assemble cilia emerged also as a critical function (Bettencourt-Dias and Glover 2007). Cilia are now established as having important roles as movement generating and sensory organelles being essential for embryonic development. Critically, cilia malfunction has been associated with abnormal left–right asymmetry, polycystic disease, obesity, neuropathies, blindness, and other ciliopathies (Nigg and Raff 2009).

Proteomic studies have been revealing an increasing number of centrosomal proteins, making it clear that many of them have unknown roles in the cell and can be translocated to different compartments during the cell cycle or in response to environmental cues (Nigg and Raff 2009; Bettencourt-Dias and Glover 2007). This shows that centrosomes concentrate and probably regulate the interactions and the intracellular distribution of a variety of proteins contributing to the establishing of networks of different pathways.

One example of a group of proteins that have been localized at the centrosome is the one that comprises several components of the tubulin-folding pathway that promotes the maturation of tubulin heterodimers, the building blocks of microtubules, as well as their regulatory and related proteins. These proteins being critical for the assembly of tubulin heterodimers competent to polymerize and controlling heterodimer quality and degradation (Lopez-Fanarraga et al. 2001) are central factors in regulating microtubule assembly and dynamics. However, different in vivo studies in different biological models suggest that these proteins may play other functions, not always related to their role in the tubulin-folding pathway. For example, the centrosomal protein TBCCD1 which is a protein related, through its functional domains, to the tubulin cofactor C, plays a role in centrosome positioning and centrioles’ interplay (Feldman and Marshall 2009; Gonçalves et al. 2010). More recently, a study conducted in the African sleeping sickness parasite Trypanosoma brucei proposed that, besides being associated with centrioles and with a centriole-associated Golgi complex structure designated by bi-lobe, TBCCD1 has a non-tubulin-related function. The authors suggest that TBCCD1 is required for the maintenance of non-microtubule filament-based structures in the Trypanosoma cytoskeleton (André et al. 2013).

The Centrosomal/Basal Body Localization of TBCCD1

The TBCCD1 (TBCC domain containing 1) protein, related to the TBCC (tubulin cofactor C) and RP2 (retinitis pigmentosa 2), was firstly described both in Chlamydomonas reinhardtii and human cells (Feldman and Marshall 2009; Gonçalves et al. 2010). The three proteins share two conserved functional domains: TBCC and CARP (Fig. 1). The CARP domain is also described in CAP proteins which regulate actin polymerization. TBCC, together with tubulin cofactor D, acts as β-tubulin GTPase-activating proteins (GAP) during the maturation of tubulin heterodimers (Fontalba et al. 1993; Tian et al. 1999). In vitro, RP2 can also act as a GAP for tubulin, but it does not substitute for TBCC in the tubulin-folding pathway. Nevertheless, a functional overlap was further shown by the capacity of RP2 to partially complement the CIN2 (tbcc homologous gene) deletion in budding yeast (Bartolini et al. 2002). RP2 was also shown to play a role in the traffic of ciliary proteins (Schwarz et al. 2012). Also, it interacts with proteins such as tubulin, polycystin-2, Gβ1 (β-subunit of rod transducin), and the ADP-ribosylation factor-like 3 (Arl3) that is the best-studied RP2-interacting protein (for review, see Schwarz et al. 2012). Arl3 is a member of the ADP-ribosylation factor family of small GTPases that is highly conserved in eukaryotes and plays important roles in the regulation of microtubule-dependent processes. Human Arl3 localizes at centrosomes, midzones, midbodies, and cilia (Zhou et al. 2006). In the mouse, its knockout causes ciliary problems like abnormal development of renal, hepatic, and pancreatic epithelial tubule structures and photoreceptor degeneration (Schrick et al. 2006). It was also shown that Arl3 is necessary for flagellum biogenesis in Leishmania (Cuvillier et al. 2000).
TBCCD1, Fig. 1

Schematic representation of the functional domains of TBCC domain-containing proteins: TBCC, TBCCD1, and RP2. These three proteins share the TBCC and CARP domains

RP2 mutations are implicated in the retinitis pigmentosa pathology characterized by a progressive photoreceptor cell degeneration leading to the loss of peripheral vision and eventually to blindness that affects about 1.8 million individuals worldwide. In fact, 15–20% of the cases of retinitis pigmentosa result from mutations in the rp2 gene (Schwahn et al. 1998). RP2 associates with the plasma membrane of photoreceptors, including the outer segments, inner segments, cell bodies, and synapses in human retinas (for review, see Schwarz et al. 2012). The GAP activity of RP2 and TBCC relies on the TBCC domain, and many of the RP2 mutations involved in this pathology occur in this domain (Schwahn et al. 1998). For example, this activity is abolished if RP2 is mutated in the only arginine residue conserved between RP2 (Arg-118) and TBCC (Arg-262), that also abolishes TBCC’s GAP activity (Veltel et al. 2008). Human, Chlamydomonas, and Trypanosoma TBCCD1 proteins do not contain this critical arginine residue, suggesting that its TBCC domain lost the GAP activity. This hypothesis is supported by the observation that human TBCCD1 is unable to rescue the phenotypes of CIN2 deletion in yeast (Gonçalves et al. 2010).

In human cells, TBCCD1 is localized in the centrosome throughout the cell cycle being also detected in the spindle midzone and in the midbody at the end of cell division (Fig. 2A). TBCCD1 was also found in the basal bodies of primary and motile cilia. Interestingly, Chlamydomonas TBCCD1 also localizes at centrioles/basal bodies and in rhizoplasts, structures that connect centrioles to each other and to the nucleus (Fig. 2B) (Feldman and Marshall 2009). A similar localization was also observed in Trypanosome where TBCCD1 is detected in the microtubule barrels of mature and probasal bodies (André et al. 2013).
TBCCD1, Fig. 2

Subcellular localization of TBCCD1 in human cells,Chlamydomonas, andTrypanosoma. (A) Human HEK-293T cells were transfected with a plasmid containing TBCCD1-RFP and immunostained using antibodies against γ-tubulin (a–c) or poly-glutamylated tubulin (d). DNA was stained with DAPI. (a) Interphase cell. (b) Cell in anaphase. (c) Cell in cytokinesis. (d) Cell cycle arrested cell after serum starvation for 24 h showing a primary cilium. Arrowheads point to TBCCD1-RFP at centrosomes. Arrows indicate spindle midzone (b), midbody (c), and basal body (d). (B) Schematic representation of the localization of TBCCD1 in human cells, in the algae Chlamydomonas, and in the parasite protozoan Trypanosoma. TBCCD1 cellular localization is shown in red in the centrosome (human cells); in the region between the rhizoplasts, structures that are responsible for the connection between the basal bodies and the nucleus (Chlamydomonas cells); and in association with the complex filament-based tripartite attachment complex that attaches mature and probasal bodies to the kinetoplast and localizes to the “bi-lobe,” a structure that connects the centriole to the Golgi complex and finally at the anterior end of the cell body (Trypanosoma cells)

TBCCD1: The Nucleus–Centrosome Connection

TBCCD1 silencing by RNAi in human RPE-1 cells caused a remarkable separation of the centrosome from the nucleus being the former often located at the cell periphery (Gonçalves et al. 2010). Similarly, in Chlamydomonas, TBCCD1 loss of function by an insertion disrupting the TBCCD1 gene caused centriole positioning defects, which leads to the assembly of the mitotic spindle with incorrect orientation (Feldman and Marshall 2009). In Trypanosoma, which present cell shape and space organization mainly based on the microtubule cytoskeleton, TBCCD1 was observed proximally to the kinetoplast (single unit-copy mitochondrial genome) (André et al. 2013). Specifically TBCCD1 is associated with the complex filament-based tripartite attachment complex (TAC) that attaches mature and probasal bodies to the kinetoplast by crossing inner and outer mitochondrial membranes. TBCCD1 is also localized to the “bi-lobe,” a structure that connects the centriole to the Golgi complex and at the anterior end of the cell body. Accordingly and similar to human cells, the depletion of Trypanosoma TBCCD1 leads to the disorganization of the structurally complex bi-lobe and loss of centriole connection to the kinetoplast of the parasite. Interestingly, in Trypanosoma dividing cells treated with RNAi that already possess segregated basal bodies but only one kinetoplast, the basal body from which the new axoneme extended presents little or no association with kinetoplast DNA suggesting that TBCCD1 is required for normal segregation of kinetoplast DNA (André et al. 2013).

In the last years, the importance of the nucleus–centrosome connection has been recognized not only for the intracellular organization of the cytoplasm but also has a key factor implicated, for example, in directed cell migration, adhesion and polarity, crucial events in development, and cancer. This connection is dynamic and able to be remodeled in response to several signals, like those involved in cell differentiation (e.g., differentiating neurons). However, the mechanisms underlying the dynamic nucleus–centrosome connection are far from being understood. In this context, TBCCD1 being required for centrosome positioning and correct mitotic spindle orientation is an emerging key factor in the nucleus–centrosome interplay and in the centrosome activities that depend on it. Supporting this view are the observations that in human cells silenced for TBCCD1, the centrosome mispositioning is accompanied by a dramatic disorganization of the Golgi apparatus and a G1 cell cycle delay. Moreover, TBCCD1-depleted cells are larger and less efficient in primary cilia assembly, and their migration is slower in wound-healing assays.

Elegant experiments growing mammalian cells in patterned surfaces have shown that geometrical constraints imposed by the substratum (extracellular matrix) and/or cell–cell interactions play a crucial role in centrosome positioning and cytoplasmic organization (Pouthas et al. 2008). However, this is not the sole responsible factor since microtubules and forces exerted on them are also critical (for review, see Burakov and Nadezhdina 2013). Indeed, the centrosome position, relative to the nucleus, is regulated by the balance between the forces that “pull” and “push” the two organelles. In interphase cells, the centrosome organizes a radial microtubule network, with a similar microtubule distribution in all directions. The balance between the polymerization/depolymerization of microtubules creates forces that “push” the centrosome in all directions, eventually promoting its location at the cell center. In this system of forces, the motor protein dynein is also a critical factor by “pulling” microtubules at the cell cortex (Burakov and Nadezhdina 2013). Partial inhibition of dynein and alterations in microtubule dynamics (due to microtubule depolymerization or stabilization) cause increasing distances between the nucleus and the centrosome in different cell lines. On the other hand, centripetal forces dependent on myosin acting in actin filament contraction also participate in regulation of nucleus and centrosome positioning at the center of the cell (Burakov and Nadezhdina 2013).

The existence of a physical link between the centrosome and the nuclear envelope is supported by a variety of data. Indeed, proteins such as Zyg-12, emerin, and Samp1 are implicated in connecting the centrosome to the nuclear envelope (for review, see Burakov and Nadezhdina 2013; Buch et al. 2009). This interaction seems to be regulated by a group of distinct kinases like the p160ROCK Rho-associated and the Polo/Greatwall (Gwl) mitotic kinases (Chevrier et al. 2002; Archambault et al. 2007).

In Chlamydomonas, the absence of TBCCD1 can lead to aberrant numbers of centrioles and flagella. TBCCD1 mutant cells (asq2) can have up to seven flagella which show that the protein is not essential for the formation of these structures. Additionally, the increased number of centrioles suggests that, in Chlamydomonas, TBCCD1 could have a regulatory role in de novo centriole assembly pathway (Feldman and Marshall 2009). This phenotype has not been detected in human cells depleted of TBCCD1 which may be related to remaining levels of TBCCD1 in these cells that impair the appearance of more dramatic effects.

Marshal and coworkers (Feldman and Marshall 2009) have also shown that the linkage between centrioles in asq2 mutant cells has been affected. Interestingly, in moving RPE-1 centrin-GFP TBCCD1 knockdown cells analyzed by time-lapse microscopy, the centrioles are less dynamic that in control cells. In fact, in control moving cells, the centrioles present a dynamic behavior in which they temporarily distance slightly from each other, suggesting that TBCCD1 is also involved in centriole–centriole interaction.

Summary and Perspectives

The characterization of TBCCD1 in human cells, Chlamydomonas, and Trypanosoma shows that TBCCD1 is a centrosomal/basal body protein conserved throughout evolution that is involved in the connection of these complex structures to DNA genomes be them nuclear or mitochondrial and affecting the activities depending on it. Also the protein is critical for Golgi apparatus organization showing to be a key factor for the maintenance of cell architecture.

The roles of TBCCD1 only now started to be revealed, and much more investigation is required to clarify its functions. In the immediate, it would be important to understand if this protein establishes a physical link between the centrosome/basal body and the nucleus or, alternatively, if it regulates this interaction. Therefore, the clarification if TBCCD1 presents or not a GAP activity, and also the identification of its partners would be a decisive step to clarify this question.


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Copyright information

© Springer International Publishing AG 2018

Authors and Affiliations

  • João Gonçalves
    • 1
    • 2
  • Helena Soares
    • 3
    • 4
  1. 1.Centro de Química e Bioquímica, Faculdade de CiênciasUniversidade de LisboaLisboaPortugal
  2. 2.Lunenfeld-Tanenbaum Research InstituteTorontoCanada
  3. 3.Departamento de Química e BioquímicaCentro de Química e Bioquímica, Faculdade de Ciências, Universidade de LisboaLisboaPortugal
  4. 4.Escola Superior de Tecnologia da Saúde de LisboaLisboaPortugal