Characterized as a protein-dense scaffolding structure responsible for the nucleation of α- and β-tubulin, centrosomes arrange and anchor microtubules that form the bipolar spindle in mitosis (reviewed in Wu and Akhmanova, 2017; Gomes Pereira et al., 2021) (Figure 1). The main microtubule nucleator is the γ-tubulin ring complex (γ-TuRC), a highly conserved complex responsible for the capping of microtubule minus ends (Oakley et al., 1990; Zheng et al., 1995). Formed of several proteins including γ-tubulin and actin (Liu et al., 2020; Wieczorek et al., 2020), this complex is located in the pericentriolar material (PCM) and was shown to rely on pericentriolar proteins such as CDK5RAP2 to attach to centrosomes (Fong et al., 2008). The γ-TuRC complex, operating as an organizational template for the nucleation of microtubules, forms the cytoplasmic microtubule array in interphase as well as the mitotic spindle during mitosis and was shown to regulate nucleation dynamics via conformational changes (Consolati et al., 2020). From interphase to mitosis, the function of centrosomes as MTOCs is highly dynamic and supports the ongoing division of proliferating cells (Mazia, 1987). Both the size and function of centrosomes as MTOCs may fluctuate according to the state of a given cell, or even its cell type (Decker et al., 2011). To behave in such a dynamic manner, MTOCs rely on centrosomal components and associated proteins that enrich at the centrosomes to stabilize or release microtubule organization and involve a large array of components that can even selectively enrich to one centriole over the other throughout the cell cycle (Andersen et al., 2003; Jakobsen et al., 2011). Combined together, all these factors allow for a personalized MTOC function specifically catered to cell conditions at a given time to accurately support cell cycle progression through microtubule nucleation.

Aside from its classic function as MTOC, the centrosome also plays crucial roles in cell polarity, shape and migration. When Van Beneden first discovered the centriole in 1883 (Van Beneden, 1883), he hypothesized that the polarity of a cell could be conferred by the orientation of both its nucleus and centrosome (Luxton and Gundersen, 2011). The nuclear-centrosomal (NC) axis exists in the majority of metazoan differentiated cell types, as well as in some unicellular organisms including yeast (Nelson, 2003). The polarity of a cell defined by the orientation of its centrosome is an important feature at the core of many biological processes. Research performed on normal fibroblast to study wound healing reported that both the Golgi apparatus and the centrosome (MTOC) were necessary for directional migration towards the edge of a lesion. Authors speculated that the coordinated orientation of both the MTOC and the Golgi apparatus towards the wound was required to modulate vesicular transport to the edge of the cell, thus leading to the growth of this extremity towards the wound (Kupfer et al., 1982). Akin to this, the centrosome was also reported to play a crucial role in directional mesenchymal cell migration. In a study published in 2017, Zhang and others used micropatterned one-dimensional adhesive strips to study cell polarity in mesenchymal cells and reported that the centrosome was involved in directional cell migration. Specifically, the centrosome was proposed to dynamically localize at the rear of mesenchymal cells to organize the microtubule network and distribute signals related to protrusive activity as a way to establish tail formation during directional migration (Zhang and Wang, 2017).

Asymmetric cell division, a process equally reliant on cell polarity for its occurrence, can also depend on the orientation of centrosomes to effectively reach completion (as reviewed in Chen and Yamashita, 2021). Asymmetric cell division is a common process routinely observed from yeast to humans. In S. cerevisiae, aging determinants are partitioned asymmetrically, resulting in a young daughter bud expanding from an older parental yeast. This process directly impacts the replicative lifespan of both parental and daughter cells, which represents the finite number of divisions a cell can undertake before reaching senescence (Longo et al., 2012). Spindle orientation and other factors established by the cell polarity machinery can guide this asymmetric process, which results in the transfer of new components such as mitochondria, endoplasmic reticulum (ER), vacuoles and rejuvenating factors to the daughter cell whilst a number of older components remain in the parental cell (Higuchi-Sanabria et al., 2014). Moreover, SPBs themselves undergo asymmetric inheritance. The older parental SPB migrates towards the new daughter bud, whilst the daughter SPB remains in the parental yeast cell (see section “MTOC duplication cycle” for more details). The asymmetric SPB segregation was shown to be regulated by the spindle positioning protein Kar9 as well as the SPB component Nud1, via its role in the mitotic exit network (MEN) (Hotz et al., 2012a). Along the same lines, asymmetric division is also a feature broadly reported in stem cells, in which the cell type of resulting cells –one self-renewed stem cell and one differentiating cell –differs. In Drosophila male germ lines, adult stem cells (GSCs) were shown to asymmetrically divide by relying on the inheritance pattern of mother and daughter centrosomes through directional orientation of the mitotic spindle (Yamashita et al., 2003). Using specific labeling techniques, Yamashita and others later observed that the mother centrosome preferentially remains affixed to the GSCs, whilst the daughter centrosome migrates to the differentiating cell (Yamashita et al., 2007). The authors hypothesized that a high number of astral microtubules may be responsible for the anchorage of the mother centrosome to the GSC, thereby keeping them in close proximity during asymmetric cell division. In accordance with this, the predetermined anchoring of the mother centrosome was suggested to act as an orientation mechanism for the mitotic spindle as a way to ensure the success of asymmetric stem cell division and highlights the core role that centrosomes can play in asymmetric stem cell division. Yet, the non-random segregation of mother and daughter centrosomes is not always a prerequisite for spindle alignment and subsequent asymmetric division. After each of the asymmetric divisions undergone by the germline lineage of the nematode C. elegans, centrosome rotation occurs as a way to re-align the spindle to the anterior-posterior (AP) axis. This rotation requires that one of the centrosomes, called the leading centrosome and chosen randomly, travels near the anterior border of the cell (Hyman and White, 1987; O’Connell, 2000). This example demonstrates that the non-random segregation of centrosomes during asymmetric division is a common occurrence in some species and does not represent an essential feature of spindle alignment for asymmetric cell division in all biological systems.

Another important function for centrosomes as MTOCs can also be observed in neuronal development. A decisive part of neuronal differentiation lies in axon specification, a process through which one of the neurites matures into a functional axon. This is of high importance for the fate of a neuron, as this process permanently defines its polarization and connectivity. In the current literature, the contribution of centrosomes to this specific stage of neuronal development has met some controversy (as reviewed in Meka et al., 2020). Several reports describe a key role for the centrosome in axonal outgrowth and specification (Rivas and Hatten, 1995; Schaar and McConnell, 2005; Tsai and Gleeson, 2005; Higginbotham and Gleeson, 2007; Kuijpers and Hoogenraad, 2011), whilst other studies seem to contradict such statement and rule out a potential requirement for centrosome function throughout this neuronal process (Esch et al., 1999; Andersen and Bi, 2000; Bradke and Dotti, 2000). In cultured hipoccampal neurons, growing axons were reported to organize microtubule arrays in a centrosome-independent way once axon specification is complete. This observation is supported by the fact that during axonal elongation, centrosome ablation was shown to have no effect on axon extension or regeneration and suggests that centrosomal function may be required only in the earlier stages of neuronal development (Stiess et al., 2010). Recent studies also argue for a role for centrosomes as F-actin organization centers in developing cultured neurons (Meka et al., 2019). Disruption of centrosome function was shown to alter the content of somatic F-actin and decreased peripheral F-actin matter in neuronal growth cones, suggesting a key role for the centrosome in F-actin organization (Meka et al., 2019). During neuronal differentiation, centrosomes as MTOCs can have various other functions. The most classical and well-known function of MTOCs in neuron biology is probably cargo transport across dendrites and axons, a function performed in partnership with motor proteins (Kapitein et al., 2010). In mouse and chick neural tube cells, centrosomes were also shown to influence neuronal delamination, a process by which novel neurons detach from the neuroepithelium throughout differentiation and morphogenesis. For delamination, the centrosome has to be retained in the newborn neuron and nucleates a wheel-like microtubule organization that supports apical abscission. In this process, the centrosome is thus of high importance in mediating microtubule activity and is involved in nervous system growth and expansion (Kasioulis et al., 2017). Another interesting function for centrosomes in neuron biology is in neuronal activity. Using fluorescent microscopy, Hu and others reported that microtubules also have the propensity to invade dendritic protrusions. This observation suggests that MTOCs, through microtubules, may have an implication in the operative exchanges between neurons. An increase in neuronal activity was notably shown to correlate with an increased number of spines occupied by microtubules, as well as with an increased contact time between microtubules and dendritic protrusions (Hu et al., 2008). However, more work is needed to establish the precise function of these microtubules in neuronal plasticity. Taken together, these examples display the various ways in which centrosomes as MTOCs can impact neuronal development and highlight the specialized –and still debated– contribution of this organelle in neuron biology.

Despite lacking a finite membrane border, the centrosome maintains its unique tri-dimensional shape via centrosome-interacting proteins, 500 of which have been identified to date (Andersen et al., 2003; Gupta et al., 2015; Gheiratmand et al., 2019). Throughout the cell cycle, its size and composition vary, allowing for diverse arrangements in microtubule organization (Devi et al., 2021; Gomes Pereira et al., 2021). Centrosomes contain centrioles, a pair of cylindrical organelles perpendicularly positioned to one another (Figure 1A). Surrounding the centrioles is the pericentriolar material (PCM), a fibrous coiled-coil protein platform (Schatten, 2008) formed by the main microtubule nucleator γ-tubulin, γ-turc, actin (Liu et al., 2020; Wieczorek et al., 2020) and pericentrin proteins (Salisbury, 1995; Levy et al., 1996; Lutz et al., 2001; Martinez-Campos et al., 2004; Wu and Akhmanova, 2017). This platform allows for sustained or transient anchoring of specific signaling proteins, such as the Nuclear Mitotic Apparatus Protein (NuMA), a key effector of the mitotic machinery. Similar to NuMA, centriolin was also reported to connect to the centrosomes during specific phases of mitosis to facilitate cell cycle progression and cytokinesis (Gromley et al., 2003). Importantly, the size of the PCM varies according to levels of γ-tubulin recruited to centrosomes in a way that supports the ongoing cell cycle state. Accordingly, the PCM is a smaller and tighter structure during interphase and becomes much larger during mitosis to support spindle formation through γ-tubulin nucleation (Robbins et al., 1968; Khodjakov and Rieder, 1999).

Aside from this core centrosomal structure comprised of centrioles and their surrounding PCM, other accessory structures including centriolar appendages and satellites positioned around the PCM further decorate centrosomes and provide this essential organelle with extra key features. The mother and daughter centrioles are different in that additional appendages can only be found on the mother centriole. Distal appendages (DAPs), existing at the distal end of mother centrioles across eukaryotic species except for C. elegans and D. melanogaster (Azimzadeh, 2014), are required for the docking of the centriole to the membrane and for the process of ciliogenesis (Tanos et al., 2013; Ye et al., 2014). Subdistal appendages (sDAPs) are found in close proximity to DAPs and are also involved in cilia formation and microtubule anchoring. In the literature, the relationship between DAPs and sDAPs remains elusive but recent evidence suggests that DAPs are important for sDAPs functionality and positioning (Chong et al., 2020). Apart from these appendages, centrosomes are also surrounded by centriolar satellites, small particles that congregate around the PCM of centrosomes (Kubo et al., 1999). These satellites are mainly composed of proteins involved in the maintenance of centrosomes, neurogenesis and ciliogenesis (reviewed in Prosser and Pelletier, 2020; Odabasi et al., 2020). Centriolar satellites can also play key roles in the transduction of several other biological cues and vary in form and function throughout the cell cycle and across cell types (Kubo and Tsukita, 2003; reviewed in Tollenaere et al., 2015).

Analogous to centrosomes, SPBs of lower eukaryotes act as key microtubule organizing centers but differ dramatically in their mechanism-of-action and structural features (Jaspersen, 2021). Across yeast species, SPBs are functionally conserved but display key architectural differences. Here, we provide a brief description of both budding yeast and fission yeast SPBs as we compare and contrast their organizational features.

In comparison to the more diffuse centrosomal organization, budding yeast S. cerevisiae SPBs are tightly embedded in the nuclear membrane through three highly organized interconnected disk-like structures (see Figure 2A for a detailed representation of budding yeast SPB structure) (Robinow and Marak, 1966; Bullitt et al., 1997; Rüthnick and Schiebel, 2018). The outer plaque is responsible for the nucleation of cytoplasmic microtubules, whereas the inner plaque generates nuclear microtubules. The central plaque anchors the SPB into the nuclear membrane and connects to the half-bridge, an important structure for SPB duplication (Figure 2A). Two tightly packed and organized disks, called intermediate layer 1 (IL1) and intermediate layer 2 (IL2), act as spacers between the outer plaque and the central plaque. In budding yeast, 17 proteins have been identified as SPB structural components (Figure 2A), six of which constitute the core of the spindle pole body: Spc42, Cnm67, Nud1, Spc72, Spc29 and Spc110 (Adams and Kilmartin, 1999; Francis and Davis, 2000; Viswanath et al., 2017). Through reciprocal interactions, these SPB components are integral for creating and maintaining the core SPB structure (Jaspersen and Winey, 2004; Jaspersen, 2021). Most SPB genes are essential for viability and single point mutations in these genes often result in temperature-sensitivity or even lethality.

Fission yeast S. pombe SPBs are bipartite structures which, akin to budding yeast SPBs, are implanted in the nuclear membrane. In opposition to budding yeast SPBs, the cytoplasmic domain of fission yeast SPBs represents the bulk of its structure. The architecture of S. pombe SPBs also differs from that of budding yeast in that it lacks intermediate spacers and does not contain multiple separate strata except from the outer (cytoplasmic), central and inner (nuclear) layers. Despite these architectural differences, fission yeast SPBs nucleate both cytoplasmic and nuclear microtubules as budding yeast SPBs do and encompass a half bridge required for SPB duplication (Ito and Bettencourt-Dias, 2018). Several components of S. pombe SPBs were classified as confirmed or probable homologues of S. cerevisiae SPBs, denoting a certain degree of functional and structural conservation in terms of SPB constituents across these species. These include (S. cerevisiae/S. pombe): Tub4/Gtb1; Spc97/Alp4; Spc98/Alp6; Spc110/Pcp1; Spc72/Mto1; Spc42/Ppc89; Cmd1/Cam1; Cnm67/Sid4; Nud1/Cdc11; Cdc31/Cdc31; Sfi1/Sfi1; Mps3/Sad1; Mps2/Kms2. For more details on S. pombe SPB and its structural intricacies, we direct readers towards the study of Bestul et al. (2017).

Centrosome duplication occurs once per cell cycle and is a semi-conservative process (i.e., the two centrioles present in each cell duplicate to generate two pairs where one template centriole is older than the newly generated copy; Figure 1B) (Gomes Pereira et al., 2021). Importantly, the process of centriole assembly occurs throughout three full cell cycles. At the onset of replication, procentrioles (new centrioles) separate and by S¹ phase start their assembly (Figures 1B, step 2). Both procentrioles elongate throughout S¹ phase, G2¹ phase and M¹ phase and grow perpendicularly from their template side (Figures 1B, step 3). At the beginning of M¹ phase, additional PCM is built around each pair of centrioles as they start to separate (Figures 1B, step 4). To form the mitotic spindle, procentrioles and their developing centrosome separate in early prophase of M¹. This event, mainly achieved by motor proteins, is supported by push-and-pull forces mediated by the kinesin motor Kif1 and the minus end-directed dynein motor complex. In the literature, the mechanistic intricacies of dynein function in MTOC positioning and separation remained elusive for many years (Holzbaur and Vallee, 1994; Vallee and Sheetz, 1996; Gönczy et al., 1999). Recent studies performed in one-cell C. elegans embryos report that different pools of dynein, localized at the cell cortex and on the nuclear surface, can influence centrosome separation. Whilst the pool of dynein located on the nuclear surface moves centrosomes by sliding the centrosome-associated microtubules, the pool of dynein at the cell cortex pulls centrosomes through MT-mediated cortical tugging forces. In this process, dynein was shown to behave as a coupling device that transfers forces produced by polarized actomyosin cortical flows to centrosomes, thereby promoting centrosome separation (De Simone and Gönczy, 2017; Torisawa and Kimura, 2020). Along with this, the plus end-directed kinesin-related motor protein Eg5 creates outward pushing forces by tethering to plus-end antiparallel MTs (Kapitein et al., 2005). Thus, dynein and Eg5 have the ability to create opposite forces that further promotes centrosome separation (Raaijmakers et al., 2012; Agircan et al., 2014). At the end of M¹ phase, procentrioles are separated and individually assemble their PCM. This occurrence, termed centriole disengagement, signifies that the mother and daughter procentrioles are not in close association anymore. Thus, at this stage of centrosome duplication, disengaged procentrioles can be defined as daughter centrioles. From G1² to S² phase of the following cell cycle, daughter centrioles acquire appendages and further increase in length. Upon entry into S² phase of the second cell cycle, each newly formed daughter centriole begins its own cycle of centrosome duplication once more. During this process, the younger mother centriole persistently accumulates additional PCM from S² phase to G2², until its PCM resembles the older mother centriole PCM prior to M² phase. In G2² phase of the second cell cycle, the younger daughter centriole still develops and acquires distal appendages (DAPs) and subdistal appendages (sDAPs). These appendages will evolve and mature until the G2³ phase of the third cell cycle, after which the corresponding round of centriole assembly is complete (Sullenberger et al., 2020).

Centrosome duplication produces two spindle poles that localize perpendicular to the plane of cell division. Achieving this precise orientation is required to support balanced chromosome segregation in mitosis (Kaseda et al., 2012; Silkworth et al., 2012; Nunes et al., 2020). Accordingly, defects in centrosome duplication can have drastic consequences for the cell. If the process of duplication fails and generates extra centrosomes, a resulting scenario may be multipolar mitosis. In multipolar mitosis, chromosomes are segregated to more than two poles during cell division and often leads to gross aneuploidy, chromosome instability (CIN) and clonal evolution (Kwon et al., 2008; Yi et al., 2011; Yang et al., 2012; Telentschak et al., 2015; LoMastro and Holland, 2019). In some cases, clustering mechanisms allow for the formation of a functional bipolar spindle despite the presence of additional centrosomes (Kwon et al., 2008). In other cases, centrosomes may gather at the center of the cell to form a monopolar spindle, a scenario equally threatening to the maintenance of genomic integrity (Chatterjee et al., 2020). Many factors can influence the organization of the mitotic spindle following defective centrosome duplication. Overall, accurate centrosome duplication and partitioning in mitosis is decisive in the maintenance of genome stability and prevention of tumorigenesis.

Like centrosome duplication, SPB duplication is a prerequisite for effective cell division in lower eukaryotes, however, since dynamic exchanges between new and old components occur throughout duplication, SPB duplication cannot be viewed as fully conservative. In budding yeast, the half-bridge elongates in early G1 and remains connected to the central plaque and the IL2 spacer throughout the duplication process (Figures 2B, step 1) (Byers and Goetsch, 1974). Once sufficiently elongated, the daughter SPB is built from satellite material (Figures 2B, step 2), developing into a duplication structure formed by Cnm67, Nud1 and Spc72 through Spc42-directed self-assembly (Winey et al., 1991; Adams and Kilmartin, 1999) (Figures 2B, step 3), after which the half-bridge retracts, allowing for the duplication plaque to embed itself into the membrane. At the end of G1 phase, the parental and daughter SPBs are leveled and connected through a full bridge (Figures 2B, step 4), the disassembly of which allows the parental SPB to preferentially migrate into the daughter bud (Figures 2B, step 5), and form a bipolar metaphase spindle (Roof et al., 1992; Jaspersen, 2021). Following the initial formation of both spindle poles, additional material is added to each SPB in a dynamic manner, hence why SPBs are considered to be dynamic: their growth process should not be viewed as exclusively conservative (Lengefeld and Barral, 2018). Instead, the continuing SPB maturation increases the ability to maintain functional integrity and has been proposed to be a mechanism for SPB repair (Jaspersen and Winey, 2004).

In fission yeast, the process of SPB duplication differs from that observed in budding yeast. The interphase SPB of S. pombe localizes in the cytoplasm, in close proximity to the nuclear envelope (NE), and embeds itself in the nuclear membrane only upon mitotic entry. In the literature, the timing of fission yeast SPB duplication throughout the cell cycle has been controversial for many years. Older studies state that SPB duplication occurs in G2/M (Ding et al., 1997), whereas newer studies suggest that the process instead begins in G1/S phase of the cell cycle (Uzawa et al., 2004). When describing SPB duplication and maturation, Uzawa and others separate the maturation process of S. pombe SPB into early and late SPB maturation. Early maturation, reported to occur upon S phase completion, represents growth of the lamellae bodies (laminated structure corresponding to the premature SPB) on the half-bridge, nuclear membrane invagination and gathering of material linking the premature SPB to the nuclear membrane. Akin to what is reported in budding yeast, the early event of SPB duplication giving rise to the lamellae bodies in fission yeast relies on the elongation of the half-bridge. The latter, without which SPB duplication could not take place, is required to support the development of the premature SPB. The newly created laminated structure, still undergoing maturation, remains linked to the mother SPB through an ellipsoid bridge (Ding et al., 1997). Late maturation, shown to take place in M phase of the cell cycle, encompasses the separation of mother and daughter SPBs, NE fenestration for SPB insertion and establishment of the mitotic spindle (Uzawa et al., 2004). While individual steps of SPB duplication differ in some respects across yeast species, the process remains broadly conserved overall.

Although centrosomes are significantly larger in size than SPBs (Gräf, 2018), they share several characteristics in duplication modes and main MTOC functions (see Figure 3 for centrosome/SPB homologs and orthologs). For example, Kendrin and CG-NAP are human orthologs of yeast Spc110 that localize at the PCM (Flory et al., 2000; Takahashi et al., 2002). Likewise, coiled-coil domains required to establish interactions with analogous binding partners are conserved across yeast Nud1 and human centriolin, both of which are important players in cell cycle progression, mitotic exit and cytokinesis (Gromley et al., 2003; Fraschini, 2019) (Figure 3). However, microtubules nucleated by the centrosome uniquely enables motility, subcellular trafficking, and anchoring of receptors at the surface of the cell (Bettencourt-Dias, 2013), whereas yeast SPBs remain restricted to roles as MTOCs and docking stations for various signaling events.

The polo-like kinase (PLK) family, comprised of PLK1-PLK5 in humans, are serine/threonine kinases that regulate fundamental aspects of cell cycle progression (Zitouni et al., 2014; Iliaki et al., 2021). Within this family, polo-like kinase 1 (PLK1) is arguably the most prominent effector of cell cycle events. PLK1 and its functional homologs in budding and fission yeasts (Cdc5 and Plo1, respectively) require phosphorylation by Cdk1/Cdc28/Cdc2 kinases and/or Aurora kinases for full activation in vivo. Following this initial activation stage, PLK1 and its yeast counterparts play crucial roles in the regulation of mitotic entry, spindle assembly, chromosome condensation, sister chromatid segregation, cytokinesis, and adaptation to DNA damage (Toczyski et al., 1997; St-Pierre et al., 2009; Ratsima et al., 2011; Zitouni et al., 2014). Importantly, PLK1 function is also essential for centrosome maturation and aberrant PLK1 activity can lead to serious diseases in humans, including cancer (Liu et al., 2017).

All PLKs share a C-terminal polo-box domain (PBD) and a highly conserved multi-domain structure with an N-terminal kinase domain (KD) that harbors a T-loop with an activating phosphorylation site (Rodriguez-Rodriguez et al., 2016; reviewed in; Serrano and D’Amours, 2016). To recognize pre-phosphorylated substrates including CDK1/Cdc28 targets, members of the PLK family use their PBD as a signal amplification module to locate and hyperphosphorylate aforementioned targets. However, the distinctive tripartite architecture of PLK4 PBD differs from the PBDs described across other PLK members and was shown to operate in a phospho-independent manner, making PLK4 PBD an exception on that matter (Slevin et al., 2012). The PBD also behaves as a subcellular targeting domain that allows PLKs to recognize and bind specific structures –such as centrosomes and SPBs– and promote specialized cell cycle functions (Colicino and Hehnly, 2018).

The other members of the PLK family perform distinct but sometimes overlapping functions in cell biology. PLK2, involved in centriole duplication, is dynamically expressed throughout the cell cycle and peaks at the G1/S transition of the cell cycle (Warnke et al., 2004). Given its implication in centriole biology, PLK2 was reported to endogenously localize at the centrosomes throughout the cell cycle. The expression of PLK2 varies widely across tissues and, given its importance in mammary gland development, was shown to be particularly high in mammary tissues (Villegas et al., 2014). On the other hand, PLK3 is more steadily expressed throughout the cell cycle and its function mostly relates to stress response pathways involving p53 during DNA damage and spindle disruption (Donohue et al., 1995; Xie et al., 2001). PLK4, derived from PLK1 (Carvalho-Santos et al., 2010) and sharing with PLK2 a role in centriole duplication, is characterized as a master regulator of MTOC formation and centrosome amplification (Habedanck et al., 2005). The last member of the PLK family, PLK5, has a slightly different structure than other members of its group in that it completely lacks a kinase domain in humans. Opposite to its other orthologs, the expression of PLK5 was shown to be very low throughout cell division and high in quiescent cells. PLK5 expression is highest in brain tissues and plays a core function in the nervous system, including neuron differentiation (de Cárcer et al., 2011a). For more information regarding the PLK family, its family members and its evolution across species, we direct readers towards reviews covering these topics (Archambault and Glover, 2009; de Cárcer et al., 2011b).

MTOCs are crucial scaffolding structures used by PLKs to reach specific substrates and promote cell division (Figure 4). In S. cerevisiae, Cdc5 decorates the nuclear surface of duplicating SPBs from late S phase to early anaphase and is also located in the nucleus. In late anaphase, Cdc5 enriches specifically on the cytoplasmic side of the parental SPB segregated to the daughter bud as well as on the bud neck (Botchkarev and Haber, 2017). Once the cell cycle is completed, Cdc5 is degraded by the anaphase-promoting complex (APC) throughout the G1 phase of the next cell cycle (Visintin et al., 2008).

In fission yeast, the polo-related kinase Plo1 shows equally important roles in cell cycle progression and displays high levels of functional overlap with budding yeast Cdc5 and human Plk1 (Lee et al., 2005). Amongst its key roles, Plo1 is required for mitotic entry, formation of the mitotic spindle, establishment of the actin ring prior to cytokinesis as well as septation activation preceding mitotic completion (Ohkura et al., 1995). Similar to Cdc5 and Plk1, Plo1 requires the SPBs as a docking platform and transiently enriches on the structure in a spatiotemporal manner throughout the cell cycle. Similar to the enrichment of Cdc5 at the SPBs, which is low in S-phase but high in G2/M (Simpson-Lavy and Brandeis, 2011), the enrichment of Plo1 on the SPBs is high during mitosis but absent in interphase. Additionally, Plo1 activity at the SPBs is highly reliant on the activity of the kinase Cdc2 (Mulvihill et al., 1999). Upon Cdc2 activation, Plo1 enriches at the SPBs and remains until spindle breakdown whilst keeping steady expression levels throughout the cell cycle (Lee et al., 2005).

The enrichment of Plo1 at the SPBs plays a pivotal role in the commitment to cell division and mitotic progression. The process of mitotic commitment is tightly regulated by M-phase promoting factor (MPF) (Ohi and Gould, 1999), composed of the regulatory subunit cyclin B and the catalytic subunit Cdc2. Following its recruitment to the SPBs in G2 phase (Alfa et al., 1989; Decottignies et al., 2001; Grallert et al., 2013), MPF activity can promote mitotic entry at any point during the cell cycle. Consequently, its activity must remain strictly restrained to the instant where cell division is timely and suitable. The kinase Wee1, via the phosphorylation of Cdc2, is responsible for such inhibitory effect on MPF activity (Russell and Nurse, 1987). Once all conditions for mitotic progression have been fulfilled, the phosphatase Cdc25 removes the inhibitory phosphorylation on Cdc2 and thus promotes cell division. Once MPF is activated, the complex creates a positive feedback loop that further promotes mitotic commitment through increased Cdc25 activity and Wee1 inhibition. Downstream of this feedback loop instigated by MPF, Plo1 interacts with the SPB component Cut12 in a way that supports entry into mitosis. The NIMA-related kinase Fin1, along with Plo1, was also reported to contribute to this positive feedback loop (Grallert and Hagan, 2002).

Apart from its involvement in mitotic entry, Plo1 is equally important throughout cell division. Plo1 shows two mechanistically distinct activity peaks during mitotic progression: First during prophase, where the formation of the actin ring occurs; second in late mitosis, corresponding to septum formation (Tanaka et al., 2001). Indeed, Plo1 was reported to localize to the medial ring structures as soon as they arise, a subcellular zone that correlates with its key function in the setting of partition sites (Bähler et al., 1998). Akin to Cdc5 in budding yeast, fission yeast Plo1 relies on the APC for its disassociation from the SPBs upon mitotic completion (Mulvihill et al., 1999). Overall, the enrichment of Plo1 at the SPBs is reflective of its implication in the spatial organization of mitotic processes and represents an essential step in the regulation of mitotic entry and cell cycle progression (Lee et al., 2005; Grallert et al., 2013).

In human cells, the Aurora-A kinase, in complex with its co-factor Bora, phosphorylates Plk1 on a conserved residue located in the T-loop of its kinase domain (T210). This G2 phase phosphorylation event uniquely occurs at the centrosomes (Bruinsma et al., 2015). Throughout the cell cycle, Plk1 localization and activity varies greatly. In late G2/early prophase, Plk1 preferentially enriches at the centrosomes to promote mitotic entry and then becomes enriched at the kinetochores to support microtubule-kinetochore connections in prometaphase, with lower Plk1 levels remaining at the centrosomes to instruct spindle assembly.

The dynamic localization of Cdc5/Plk1 at MTOCs has major implications for signal transduction events during the cellular response to DNA damage. Upon DNA damage, cells initiate a checkpoint response that allows time for DNA repair by preventing the G2/M transition (Sandell and Zakian, 1993; Rhind and Russell, 1998; Cagney et al., 2006; Chao et al., 2017). After successful DNA damage repair, cells resume cycling through a process termed checkpoint recovery (Vaze et al., 2002). However, not all DNA lesions can be safely repaired, and the extent of damage suffered determines the fate of the damaged cells. When DNA damage is too extensive, apoptotic signals lead to programmed cell death thereby preventing the transfer of deleterious genomic errors to daughter cells. When DNA damage is less extensive, cells can resume their cell cycle through checkpoint adaptation (or bypass) despite the presence of “permanent” DNA damage (Sandell and Zakian, 1993; Toczyski et al., 1997; Lee et al., 1998; Vidanes et al., 2010; Ratsima et al., 2011). Consequently, the process of checkpoint adaptation postpones the repair of DNA lesions to subsequent phases of the cell cycle.

The exact signaling pathway responsible for the adaptation response to persistent DNA damage is still not fully understood. In both human and yeast cells, PLK activity is required for adaptation, and Cdc5 enrichment at the SPBs is both necessary and sufficient to promote adaptation to persistent DNA damage in budding yeast cells (Ratsima et al., 2016). These observations suggest that SPBs function as docking platforms for Cdc5 to execute the adaptation response. How this is achieved is unclear, however a possible link connecting PLK, BRCA1 and centrosomes was recently proposed in human cancers (Yoshino et al., 2021). In some cases, aberrant expression of the tumor suppressor gene BRCA1 in mammary tissues can dysregulate centrosome duplication and generate a higher centriole number in vivo. This reported process requires the tethering of BRCA1 to centrosomes via RACK1. This protein also acts as a scaffolding factor that promotes Aurora A and PLK1 interaction in S phase. Previous literature linked RACK1 overexpression to centriole overduplication and involved BRCA1 as a component in this process (Yoshino et al., 2019; Yoshino et al., 2020). This centriole overduplication event was shown to stem from higher levels of phosphorylated PLK1, resulting in kinase hyperactivity at centrosomes. The reported centrosome aberration phenotype in response to PLK1 hyperactivity is intriguingly reminiscent of the supernumerary SPB and polyploidy/multinucleation phenotypes observed in adaptation-defective cdc5-16 mutants (Ratsima et al., 2011; Ratsima et al., 2016) and in cells overexpressing CDC5 (Song et al., 2000; Bartholomew et al., 2001). However, more research is needed to assess whether there are cross-species phenotypic similarities between these two cellular processes and how this might be related to the adaptation response to unrepairable DNA damage. Despite the impact of BRCA1 aberrations reported above in centrosome amplification (Yoshino et al., 2019; Yoshino et al., 2020), other studies demonstrated that mutations in BRCA1 can induce a variety of phenotypes that do not always result in amplified centrosomes. To further evaluate the influence of BRCA1 in centrosome biology in vivo, Kais and others explored the effect of a subset of mutations in the BRCA1 locus on centrosome behavior. Remarkably, these mutations induced a range of phenotypes affecting two separate branches of centrosome biology, namely centriole pairing and centrosome number. This result suggests that BRCA1 regulates these two branches of centrosome duplication separately, and nicely underlines the separation-of-function aspect of certain BRCA1 mutations (Kais et al., 2012). Thus, some mutations in BRCA1 can affect functions unrelated to centrosome number and do not always correlate with centrosome amplification in transformed cells.

The process of DNA damage-induced centrosome amplification (DDICA) (Zou et al., 2014) represents another intriguing link connecting DNA damage responses, PLKs and MTOCs. After treatment with the DNA crosslinker mitomycin C, higher levels of BRCA1 and PLK1 were detected at centrosomes alongside increased centrosome amplification. How DDICA might enhance genomic stability and/or survival remains unclear to this day. On one hand, DDICA may promote the elimination of cells bearing extensive amounts of DNA damage through mitotic catastrophe, whilst contributing to DNA damage repair via local increase of DNA repair factors at the centrosomes (Yoshino et al., 2021). The rationale behind this is that an increased amount of DNA repair factors at the centrosomes stemming from DDICA could constitute an extra source of DNA repair proteins available for relocation from the centrosomes to nuclear sites of DNA damage, consequently supporting nuclear DNA repair as well as DDICA processing. This theory, however, remains to be proven and is a work in progress in the current literature. On the other hand, this process was suggested to be beneficial for cancer cells seeking a proliferative advantage in specific growth environments, as centrosome amplification in p53-deficient cancer cells can encourage chromosome mis-segregation (Yoshino et al., 2021), a key promoter of genomic heterogeneity. The mitotic catastrophe phenotype resulting from DDICA, observed primarily in breast cancer cells, is intriguingly evocative of the phenotype reported in budding yeast with the adaptation-defective cdc5-16 allele. In response to DNA damage, this mutant fails to enrich at the SPB and gradually fragments its SPB, akin to DDICA (Ratsima et al., 2011). It would be informative for future research to explore the mechanistic similarities between Cdc5-related SPB fragmentation in yeast and PLK1/BRCA1-related DDICA in breast cancer cells.

In damaged cells, the generation of extra centrosomes can also be an outcome of circumstances unrelated to PLK1 or BRCA1 expression. Dodson and others notably reported that centrosome amplification can ensue an extended G2 phase caused by DNA damage checkpoint activation, in which DNA replication is paused but centrosome duplication remains. Interestingly, the small portion of cells able to override this G2/M cell cycle arrest were shown to contain a normal number of centrosomes (Dodson et al., 2004). Other potential causes of centrosome amplification also include cytokinesis failures, as well as cell-cell fusion (reviewed in Godinho and Pellman, 2014). Overall, the relationship between centrosome amplification and DNA damage is an ongoing work in progress in the field of centrosome biology and its intricacies are yet to be fully uncovered.

Nek2 is known to phosphorylate ß-catenin, a multifunctional Wnt-pathway effector implicated in a wide array of biological contexts including centrosome-related cellular processes (Kaplan et al., 2004; Bahmanyar et al., 2008; Vora et al., 2020). Throughout mitosis, Nek2-mediated ß-catenin phosphorylation prevents its degradation, a mechanism required to maintain high levels of centrosomal ß-catenin (Mbom et al., 2014) and associated with accurate centrosome disjunction. Nek2 kinase activity at the start of mitosis relies on Plk1 (Mardin et al., 2011), however, ß-catenin enrichment at the centrosomes is independent of its phosphorylation state (Mbom et al., 2014). Outside of the centrosome, the Nek2b isoform forms a complex with T-cell factor (TCF4) to drive ß-catenin-dependent cell proliferation, a mechanism associated with tumor cell invasion and metastasis (Shin et al., 2017; Zhang et al., 2017; Shen et al., 2019).

Nek2 also phosphorylates dishevelled (DVL), a scaffold protein involved in both the canonical and non-canonical branches of the Wnt pathway. In higher organisms, three genes encode DVL isoforms –DVL1, DVL2 and DVL3. These isoforms, broadly expressed in mammalian cells, were reported to have partly overlapping functions with high levels of redundancy (Lee et al., 2008). The phosphorylation event mediated by Nek2 on DVL isoforms is essential to promote interactions between DVL and several centrosomal linker proteins, liberating these from the centrosomes and ultimately promoting centrosomal separation. Indeed, lack of DVL impedes the dissolution of centrosomal linkers, resulting in an absence of centrosomal separation (Cervenka et al., 2016). Nek2 can also positively modulate the pool of DVL available at the centrosomes to upregulate the canonical Wnt/β-catenin pathway (Cervenka et al., 2016).

Apart from its implication in centrosome separation, the Wnt signaling pathway was also reported to play a role in cell motility. In response to exosome-transported signaling molecules named planar cell polarity (PCP) proteins, the Wnt pathway stimulates breast cancer cell (BCC) motility at the cell cortex. For this event to occur, the association of a centrosomal module is required. Specifically, DVL2 isoform was shown to mediate the assembly of this module, composed of the human centrosomal protein CEP192 and PLK4/AURKB, to promote protrusive activity in BCCs. This centrosomal module coordinates the exchange of formin DAAM1 for formin DAAM2 at the cell cortex, resulting in increased cell migration (Luo et al., 2019). This sequence of events may partly explain why aberrant expression of PLK4, AURKB and DAAM2 in breast cancer was shown to correlate with poor prognosis and increased cancer aggressiveness (http://www.cbioportal.org). Interestingly, the function of this centrosomal module was reported to be independent of centrosomes or microtubules and elegantly highlights how contextual Wnt signalling in cancer cells has the power to initiate processes such as cell migration as a means to augment metastatic potential.

In the developing Drosophila eye, the relationship between Nek2 and Wnt/Wingless is more direct. In a setting where the anaphase-promoting complex (APC) is inactivated, Nek2 accumulation causes hyperactivation of Wnt signaling and blocks retinal differentiation. Conversely, when Nek2 is degraded by APC, local Wnt signaling is suppressed and retinal differentiation proceeds (Martins et al., 2017). Taken together, these studies highlight how Nek2 operates in partnership with the Wnt pathway throughout the entire cell, including at the centrosomes.