Muscle cells in the heart contract in regular rhythms to pump blood around the body. In humans, rats and other mammals, the vast majority of heart muscle cells lose the ability to divide shortly after birth. Therefore, the heart is unable to replace cells that are lost over the life of the individual, for example, during a heart attack. If too many of these cells are lost, the heart will be unable to pump effectively, which can lead to heart failure. Currently, the only treatment option in humans with heart failure is to perform a heart transplant.

Some animals, such as newts and zebrafish, are able to replace lost heart muscle cells throughout their lifetimes. Thus, these species are able to fully regenerate their hearts even after 20% has been removed. This suggests that it might be possible to manipulate human heart muscle cells to make them divide and regenerate the heart. Recent research has suggested that structures called centrosomes, known to be required to separate copies of the DNA during cell division, are used as a hub to integrate the initial signals that determine whether a cell should divide or not.

Here, Zebrowski et al. studied the centrosomes of heart muscle cells in rats, newts and zebrafish. The experiments show that the centrosomes in rat heart muscle cells are dissembled shortly after birth. Centrosomes are made of several proteins and, in the rat cells, these proteins moved to the membrane that surrounded the nucleus. On the other hand, the centrosomes in the heart muscle cells of the adult newts and zebrafish remained intact.

Further experiments found that that breaking apart the centrosomes of heart muscle cells taken from newborn rats stops these cells from dividing. Zebrowski et al.'s findings suggest that the loss of centrosomes after birth is a possible reason why the hearts of adult humans and other mammals are unable to regenerate after injury. In the future, these findings may aid the development of methods to regenerate human heart muscle and new treatments that may limit division of cancer cells.

The adult mammalian heart is considered to be a post-mitotic organ, as mammalian cardiomyocytes lose their ability to proliferate shortly after birth (Li et al., 1996; Soonpaa et al., 1996; Zebrowski and Engel, 2013). This is supported by the limited regenerative capacity of the adult mammalian heart (Senyo et al., 2014) and the fact that primary adult cardiomyocyte-born tumors are extremely rare, if they exist at all (Dell'Amore et al., 2011). Understanding the underlying mechanisms governing the post-mitotic state of adult mammalian cardiomyocytes may clarify whether it is possible to induce cardiac regeneration based on cardiomyocyte proliferation as seen in zebrafish and newts (Poss et al., 2002; Bettencourt-Dias et al., 2003; Gamba et al., 2014). Tremendous efforts have been invested to induce postnatal mammalian cardiomyocyte proliferation. However, besides the fact that cell cycle promoting factors (e.g., Cyclins) are downregulated around birth, while cell cycle inhibitors (e.g., Cyclin-dependent kinase inhibitors) are induced (Ikenishi et al., 2012), little is known about the mechanisms that induce cell cycle exit or establish the post-mitotic state in mammalian cardiomyocytes (van Amerongen and Engel, 2008).

The centrosome is a solitary, juxtanuclear organelle in metazoan cell-types. It consists of a pair of tubulin-based structures, called centrioles, encased in a dense, non-membranous, multi-protein cloud called the pericentriolar matrix (PCM), which is further surrounded by a dispersed array of proteins termed centriolar satellites (Bettencourt-Dias, 2013). Traditionally, the centrosome is known as the microtubule organizing center (MTOC) of the cell, and is required for primary cilium formation (Kim and Dynlacht, 2013). Recently, the centrosome has emerged as a critical signaling hub for the cell cycle regulatory machinery (Doxsey et al., 2005). For instance, centrosome localization of Cyclin E and Cyclin A is required for G1/S cell cycle progression (Pascreau et al., 2011). Consistent with this role, increasing evidence supports the requirement of a functional centrosome for proliferative potential in mammalian cell-types (Doxsey et al., 2005). For instance, cells undergo G0/G1 arrest when (i) centrosomes are removed via laser ablation, (ii) centrosome integrity is disrupted via knockdown of centrosome proteins, or (iii) centriole biogenesis is blocked via a chemical inhibitor (Hinchcliffe et al., 2001; Khodjakov and Rieder, 2001; Srsen et al., 2006; Mikule et al., 2007; Wong et al., 2015).

To date, the role of centrosome integrity in cell proliferation has always been studied in the context of centrosome component mutants. Here, we show that centrosome integrity is developmentally regulated in mammalian cardiomyocytes, revealing a novel mechanism that renders cells post-mitotic. Our findings might have important implications for efforts to induce therapeutic cardiomyocyte proliferation in adult mammalian hearts.

A normal diploid cell in G0/G1-phase contains one centrosome with two paired centrioles. During S-phase the centrosome duplicates whereby the two parental centrioles form daughter centrioles. Around the transition from G2-phase to mitosis, parental centrioles ‘split’ (loss of cohesion) resulting in two separated centrosomes that become part of the spindle poles during mitosis (Doxsey et al., 2005). When centrosome integrity is compromised during G0/G1-phase, centrioles can lose cohesion (i.e., become unpaired) adopting a premature ‘split’ configuration, defined as a distance > 2 µm between centrioles (Graser et al., 2007). To determine if centrosome integrity changes in cardiomyocytes during development, we analyzed centriole configuration using antibodies against γ-tubulin, a marker of both the centriole and the PCM (Sonnen et al., 2012). Centrioles were observed to be in a typical paired configuration in the majority of cultured cardiomyocytes isolated from embryonic day (E) 15, E18, and postnatal day (P) 0 rat hearts (Figure 1A,B) as well as in cardiomyocytes from E15 and P0 rat heart sections (Figure 1—figure supplement 1A,B). In contrast, shortly after birth, centrioles were split in the vast majority of P3 and P5 cardiomyocytes in vitro (Figure 1A,B) and in vivo (Figure 1—figure supplement 1A,B). Analysis of mother and daughter centriole markers (Odf2 and Centrobin, respectively) in isolated rat cardiomyocytes verified that each γ-tubulin signal represented a single centriole (Figure 1—figure supplement 1C). As an internal control, cardiac non-myocytes from the same cultures and tissue sections were examined. The majority of non-myocytes in the heart are fibroblasts, endothelial cells, and smooth muscle cells that all have the ability to proliferate. At all examined developmental stages, non-myocytes showed a typical paired-centriole configuration indicating that the split-centriole phenotype was cardiomyocyte-specific (Figure 1B and Figure 1—figure supplement 1A,B). Collectively, these data demonstrate that centriole cohesion, and thus centrosome integrity, is lost in mammalian cardiomyocytes shortly after birth.

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To identify an underlying cause of the split-centriole phenotype, the cellular localization of various centrosome proteins was assessed in isolated cardiomyocytes from different developmental stages. The PCM proteins Pericentrin and Cdk5Rap2 have previously been shown to be required for centriole-cohesion (Graser et al., 2007; Matsuo et al., 2010). Consistent with this function, both PCM proteins localized to the centrosome in E15-isolated cardiomyocytes (Figure 1C,D and Figure 1—figure supplement 1D,E). In contrast, both proteins were localized to the nuclear envelope in P3-isolated cardiomyocytes (Figure 1C,D and Figure 1—figure supplement 1D,E). Although remnants of Pericentrin and Cdk5Rap2 could be observed at the centriole in P3-isolated cardiomyocytes (Figure 1C,D and Figure 1—figure supplement 1D,E), their presence was significantly reduced when centrioles were split (Figure 1E and Figure 1—figure supplement 1F). Pcnt siRNA-mediated knockdown in P0-isolated cardiomyocytes resulted in an increase of split-centrioles (Figure 1F,G), confirming that Pericentrin is required for centriole-cohesion in cardiomyocytes. In contrast to PCM proteins, the centriole-associated proteins CEP135, Odf2, and Centrobin were not observed at the nuclear envelope in P3-isolated cardiomyocytes (Figure 1C and Figure 1—figure supplement 1C,D). Collectively, these results indicate that loss of centriole cohesion is accompanied by redistribution of centrosome proteins to the nuclear envelope.

Centrosome localization of Pericentrin is dependent on the centriole satellite protein PCM1 (Dammermann and Merdes, 2002; Barenz et al., 2011). Consistent with this, PCM1 localized at the centrosome and the nuclear envelope in E15- and P3-isolated cardiomyocytes, respectively (Figure 1C,H and Figure 1—figure supplement 1D). Further, nuclear envelope localization of PCM1 occurred by P0 (Figure 1H), prior to that of Pericentrin (Figure 1D). Occasionally, PCM1 was observed in a semi-belt pattern at the nuclear envelope proximal to the centrosome in E15-isolated cardiomyocytes (Figure 1—figure supplement 1G), indicating a transitional state. These data suggest that in cardiomyocytes, PCM1 is also required for the localization of Pericentrin to the centrosome. To test this, we overexpressed the major isoform of Pericentrin associated with the centrosome, Pericentrin B, in E15-, P0-, and P3-isolated cardiomyocytes and non-myocytes. Pericentrin B-GFP localized to centrioles in non-myocytes as well as E15-isolated cardiomyocytes (Figure 1—figure supplement 2A,B). In contrast, Pericentrin B-GFP did not localize to the centrioles in the vast majority of P0- and P3-isolated cardiomyocytes but rather created cellular aggregates which were not observed in E15 cardiomyocytes or non-myocytes. These data indicate that the machinery required for centrosome integrity is lost in postnatal cardiomyocytes.

To determine if relocalization of centrosome proteins to the nuclear envelope and loss of centriole-cohesion is caused by cell autonomous mechanisms, long-term culturing experiments were performed. Long-term culturing of E15-isolated and P0-isolated cardiomyocytes resulted in PCM1 and Pericentrin relocalization to the nuclear envelope (Figure 1I) and loss of paired-centrioles (Figure 1J), respectively. Further, mouse iPSC-derived cardiomyocytes had split-centrioles and the centrosome proteins PCM1 and Pericentrin localized to the nuclear envelope (Figure 1—figure supplement 2C). These results indicate that loss of centrosome integrity begins during fetal development and progresses in a cell autonomous manner.

Previously, it has been demonstrated that Pcnt is alternatively spliced (Miyoshi et al., 2006), resulting in two isoforms; Pericentrin B which resembles the human Pericentrin Kendrin, and Pericentrin S, which lacks an N-terminus region. Pericentrin B is ubiquitously expressed at all developmental stages. In contrast, expression of Pericentrin S starts late in fetal development and was found to be specific for adult heart and skeletal muscle (Miyoshi et al., 2006). Therefore, we speculated that expression of the Pericentrin S isoform may coincide with changes in centrosome integrity during the development of cardiomyocytes. RT-PCR analysis revealed the appearance of Pcnt S expression shortly before birth in the heart (Figure 1K). Antibodies specific for the Pericentrin B isoform indicated that Pericentrin S, and not Pericentrin B, was the predominant Pericentrin isoform at the nuclear envelope in P3-isolated cardiomyocytes (Figure 1L). These results indicate that Pcnt is alternatively spliced during perinatal heart development and this change is related to its relocalization to the nuclear envelope.

Loss of centrosome integrity in postnatal cardiomyocytes suggested that centrosome function might be compromised as well. Nearly all cell-types studied to date are capable of forming a primary cilium when arrested in G0/G1-phase (Bowser and Wheatley, 2000, Nigg and Stearns, 2011). This suggests that cardiomyocytes, which are arrested shortly after birth in G0/G1 phase (Takeuchi, 2014), should be capable of ciliogenesis. Yet, given that PCM1, which is required for ciliogenesis (Kim et al., 2008), is lost from the centrosome during development, we speculated that ciliogenesis is suppressed in cardiomyocytes. Induction of ciliogenesis via serum starvation resulted in the formation of a primary cilium in E15-isolated cardiomyocytes and non-myocytes (Figure 2A). However, the frequency of cardiomyocytes capable of ciliogenesis decreased with heart development (Figure 2B) with less than 1% of isolated postnatal cardiomyocytes, which were arrested in G1/G0 as confirmed by lack of Ki67 expression and FACS analysis (Figure 2—figure supplement 1A–D), forming a primary cilium. Further, no postnatal binucleated cardiomyocytes were observed to be capable of ciliogenesis (Figure 2A). In contrast to cardiomyocytes, the frequency of cardiac non-myocytes capable of ciliogenesis was high at all developmental stages investigated (Figure 2B).

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Pericentrin and Cdk5Rap2 are required for the centrosome to function as the cellular MTOC (Takahashi et al., 2002; Choi et al., 2010). As these proteins localize to the nuclear envelope during neonatal development, we hypothesized that the cellular MTOC is transferred from the centrosome to the nuclear envelope. In accordance with this hypothesis, microtubules were found to predominantly emanate from the centrosome in E15-isolated cardiomyocytes as also observed in non-myocytes. In contrast, in P3-isolated cardiomyocytes microtubules were found to predominantly emanate from the nuclear envelope (Figure 2C). The local shift of the MTOC during development was confirmed by a MTOC-regrowth assay (Figure 2D and Figure 2—figure supplement 1E,F), which further demonstrated that microtubules do not emanate from centrioles in P3-isolated cardiomyocytes (Figure 2E and Figure 2—figure supplement 1G,H). Finally, siRNA-mediated knockdown of Pcnt in P3-isolated cardiomyocytes demonstrated that Pericentrin is required for a functional MTOC at the nuclear envelope (Figure 2F and Figure 2—figure supplement 1I). Collectively, these results indicate that, in addition to centrosome integrity, centrosome function is progressively compromised in cardiomyocytes during development.

Given that traditional centrosome functions of ciliogenesis and microtubule organization are lost in postnatal cardiomyocytes, we hypothesized that there would be a relationship between centrosome integrity and proliferative potential. To test this, the cell cycle marker Ki67 was assessed in cardiomyocytes from different developmental stages. Cardiomyocyte proliferative potential decreased with neonatal development (Figure 3A). Moreover, P3-isolated cardiomyocytes with paired-centrioles exhibited greater proliferative potential than those with split centrioles (Figure 3B,C) at a frequency similar to that of E15- and P0-isolated cardiomyocytes (Figure 3A,C)—of which the vast majority have paired-centrioles (Figure 1B). In contrast, cardiac non-myocyte proliferative potential, as well as the percentage of non-myocytes with paired centrioles, did not decrease during neonatal development (Figure 3—figure supplement 1A). Subsequently, we tested whether centrosome integrity is required for cardiomyocyte proliferative potential. Centrosome integrity was disrupted by either siRNA-mediated knockdown of Pcnt, or overexpression of a RFP-tagged dominant negative C-terminal Pericentrin (RFP-Peri^CT), which displaces endogenous Pericentrin localization (Gillingham and Munro, 2000; Mikule et al., 2007). Immunofluorescence analysis confirmed that RFP-Peri^CT localizes to the centrioles in P0-isolated cardiomyocytes (Figure 3—figure supplement 1B). Both methods used to disrupt centrosome integrity suppressed P0-isolated cardiomyocyte proliferative potential (Figure 3D–F). Taken together, these data indicate that loss of centrosome integrity promotes G0/G1 cell cycle arrest in mammalian cardiomyocytes.

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Disruption of centrosome integrity promotes p38MAP kinase (p38)-mediated G1/S cell cycle arrest (Mikule et al., 2007; Pascreau et al., 2011). Previously, it has been demonstrated that p38 inhibition (p38i) allows postnatal cardiomyocyte proliferation (Engel et al., 2005). This raised the question whether p38i can over-ride post-mitotic arrest in cardiomyocytes with split-centrioles. p38i enhanced the proliferative potential of P3-isolated cardiomyocytes with split-centrioles (Figure 3C). These results indicate that suppression of the p38-mediated stress-activated pathway can promote cell cycle progression in cardiomyocytes that lack centriole-cohesion.

A bipolar mitotic spindle is required for high fidelity chromosome segregation (Pihan, 2013). Thus, we determined if the observed changes in centrosome integrity affects spindle pole number during mitosis in postnatal cardiomyocytes when cell cycle progression is induced. In response to serum stimulation, P0- and P3-isolated cardiomyocytes exhibited a typical metaphase consisting of two γ-tubulin foci (Figure 3G,H). p38i had no effect on the number of γ-tubulin foci in P0-isolated cardiomyocytes during metaphase (Figure 3H). In contrast, p38i resulted in a significant increase in metaphases containing multiple γ-tubulin foci in P3-isolated cardiomyocytes (Figure 3G,H). Similarly, adult-isolated cardiomyocytes induced to re-enter the cell cycle by p38i + FGF1 also exhibited multiple γ-tubulin foci during metaphase (Figure 3I). These data indicate that over-riding cell cycle arrest in cardiomyocytes results in multiple spindle poles.

In contrast to mammals, adult newts and zebrafish can regenerate their heart through cardiomyocyte proliferation (Poss et al., 2002; Bettencourt-Dias et al., 2003). This suggests that they either do not establish a post-mitotic state by disassembling their centrosomes or they are able to reverse this mechanism upon injury. To test which scenario occurs in these species, we analyzed their centrosomes. Paired centrioles could readily be identified in adult newt and zebrafish cardiomyocyte nuclei (Figure 4A). Further, the frequency of newt and zebrafish cardiomyocyte nuclei with intact centrosomes was similar to that of non-myocyte nuclei (Figure 4B). In addition, MTOC-regrowth assays demonstrated that centrosomes in zebrafish and newt cardiomyocytes are functional (Figure 4C and Figure 4—figure supplement 1A,B). Collectively, these results suggest that, unlike mammalian cardiomyocytes, newt and zebrafish cardiomyocytes maintain centrosome integrity throughout adulthood.

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We then sought to determine if zebrafish cells require centrosome integrity for proliferation in vivo. To determine this, we injected RNA encoding RFP-tagged dominant negative C-terminal Pericentrin (RFP-Peri^CT) into one-cell stage zebrafish embryos. Compared to control embryos (RFP RNA-injected), a significantly larger number of RFP-Peri^CT RNA-injected embryos showed developmental delay at 24 hr post fertilization (hpf) and a significantly reduced number of H3P-positive cells (Figure 4—figure supplement 1C–F). Since RFP-Peri^CT RNA did not cause necrosis or lethality, it is plausible that the observed developmental delay was caused, at least partly, by reduced proliferation of RFP-Peri^CT-expressing cells. To further test an effect on proliferation, we injected DNA constructs driving RFP or dominant negative Pericentrin (RFP-Peri^CT) expression from the CMV promoter into one-cell stage embryos and analyzed the size of RFP-positive clones of cells at 20 hpf. Embryos were sorted into four classes according to the size of the clones, namely no visible clones (class I), few scattered cells (class II), medium-sized clones (class III), and large clones (class IV). RFP DNA-injected controls exhibited comparable numbers of embryos displaying no clones (class 1), scattered (class II), medium-sized (class III), and larger clones (class IV). In contrast, the majority of RFP-Peri^CT DNA-injected embryos contained either no clones (class I) or few scattered clones (class II), while larger clones (class III and IV) were rarely seen (Figure 4—figure supplement 1G,H). This suggests that clonal expansion of RFP-Peri^CT expressing cells is reduced, which substantiates the hypothesis that expression of RFP-Peri^CT reduces cell proliferative potential. These results suggest that zebrafish cells require centrosome integrity for proliferation in vivo.

Increasing evidence supports the requirement of a functional centrosome for cellular proliferative potential (Doxsey et al., 2005). For instance, somatic cell-types arrest in G0/G1 when centrosomes are removed or disrupted (Hinchcliffe et al., 2001; Khodjakov and Rieder, 2001; Srsen et al., 2006; Mikule et al., 2007; Wong et al., 2015). Further, centrosome degradation occurs during female meiosis—a process believed to inhibit parthenogenesis—with mitotic cycles restored upon fertilization when the sperm donates a centrosome during zygote formation (Klotz et al., 1990; Clift and Schuh, 2013). Moreover, relocalization of the MTOC to non-centrosomal loci has been described for several post-mitotic, highly differentiated, cell-types (Bartolini and Gundersen, 2006). This body of literature coupled with our results suggests that centrosome integrity can be developmentally regulated to achieve a post-mitotic state.

Centrosome disassembly appears to be a very effective way to achieve a post-mitotic state. But why do cardiomyocytes disassemble their centrosomes? Upon birth, the neonatal heart, and the cardiomyocytes therein, undergo increased hemodynamic stress. Effective cardiomyocyte function in response to increased hemodynamic stress may require a cytoskeletal architecture more conducive to handling postnatal physical stresses (e.g., a nuclear envelope-based MTOC). Thus, centrosome disassembly may be a result of cytoskeletal reorganization. In this scenario, proliferative potential might be sacrificed for postnatal function.

It has long been considered that skeletal myoblasts and cardiomyocytes have distinct molecular mechanisms to control their proliferative growth. This theory has been primarily based on the observation that proliferation and differentiation (contraction) are mutually exclusive in skeletal muscle but occur in parallel in cardiomyocytes (Ueno et al., 1988). However, our results together with others (Tassin et al., 1985; Bugnard et al., 2005; Srsen et al., 2006; Fant et al., 2009; Zaal et al., 2011) challenges this dogma, as establishment of a post-mitotic state in both muscle cell-types coincides with MTOC relocalization to the nuclear envelope and centrosome disassembly.

The observation that p38i promotes cardiomyocyte proliferative potential in the absence of a functional centrosome provides some optimism for heart regeneration via proliferation of endogenous cardiomyocytes. However, we observe that an absence of centrosome integrity correlates with multiple spindle poles. Although multiple spindle poles are generally resolved into a semi-clustered/pseudo-bipolar conformation, this process is nevertheless highly prone to the formation of merotelic spindle-kinetochore attachments, which can promote chromosome missegregation (Ganem et al., 2009). Indeed, postnatal cardiomyocytes induced to proliferate exhibit chromosome segregation abnormalities (Engel et al., 2006). While cell viability can exist when aneuploidy is limited (e.g., as seen in Down Syndrome), aneuploidy is generally not well tolerated (Torres et al., 2008). Therefore, proliferation of adult cardiomyocytes may not necessarily result in viable daughter cells. Thus, in the absence of a functional centrosome, whether a particular manipulation induces aneuploidy, and to what degree, may be a critical factor in determining its regenerative therapeutic potential.

The origin of multiple spindle poles (i.e., multiple γ-tubulin foci) during cardiomyocyte mitosis is not entirely clear. Interestingly, in addition to centriole-cohesion, Pericentrin and Cdk5Rap2 are also required for centriole-engagement (Barrera et al., 2010; Lee and Rhee, 2012). During S-phase, centrioles are duplicated, with each mother centriole forming a daughter centriole, which remains closely attached (i.e., engaged) until mid-anaphase (Kuriyama and Borisy, 1981; Sluder, 2013). When mother-daughter centrioles are engaged, only the mother centriole exhibits a strong γ-tubulin signal (Wang et al., 2011). Thus, one speculation is that when cells that lack centriole-cohesion enter mitosis, they have an increased likelihood of premature loss of centriole-engagement, thus accounting for the four γ-tubulin signals observed at metaphase. However, it has to be considered that induction of mitosis in bi-nucleated cardiomyocytes, which increasingly appear after birth, results in four γ-tubulin signals in metaphase corresponding to four duplicated centrioles.

The ability of zebrafish and newts to regenerate their heart has gained extensive interest in recent years. One major question is what distinguishes mammalian cardiomyocytes from those of zebrafish and newts with regards to their proliferative potential. Our data demonstrate that the state of cellular differentiation of cardiomyocytes from various species is not evolutionary conserved. The fact that adult zebrafish and newt cardiomyocytes maintain their centrosome integrity indicates that factors promoting adult zebrafish cardiomyocyte proliferation might not necessarily induce adult mammalian cardiomyocyte proliferation.

Recently, it has been shown that planarians can develop and regenerate in the absence of centrosomes (Azimzadeh et al., 2012). This has questioned the requirement of a centrosome for cell proliferation during development and regeneration. However, there are significant differences between planarians and vertebrates in cell cycle control. For example, planarians express only one single repressive E2F, whereas mammals and zebrafish express several repressive and activating E2Fs. In addition, planarians do not have Cyclin E or Cyclin A homologs. Further, cyclin-dependent kinase CDK2 (to which Cyclin E/A usually binds) is expressed at such low levels in planarians that it is considered functionally dead (Zhu and Pearson, 2013). This is important as in mammals centrosome localization of Cyclin E and Cyclin A is required for G1/S cell cycle progression (Pascreau et al., 2011). Thus, at least in the case of Cyclin E and Cyclin A, planarians lack centrosome-regulated cell cycle factors. In addition, our data indicate that centrosome integrity is required for proliferation during zebrafish development. This is in agreement with the recent observation that plk4 depletion in zebrafish impairs centriolar biogenesis during development and increases premature cell cycle exit (independent of ciliogenesis defects) resulting in reduced zebrafish size (Martin et al., 2014). Other examples of centriole-associated genes whose depletion causes cell cycle defects resulting in impaired development are stil (Pfaff et al., 2007; Vulprecht et al., 2012; Sun et al., 2014) and cetn2 (Delaval et al., 2011). While there is no evidence for the requirement of functional centrosomes in cardiac regeneration in zebrafish, there are data that indirectly suggest that centrosome function might be required for cardiac regeneration. For example it has been shown that inhibition of mps1 and plk1, factors implicated in centrosome assembly and maturation (Pike and Fisk, 2011; Joukov et al., 2014; Kong et al., 2014), impairs cardiac regeneration in zebrafish after apex resection (Poss et al., 2002; Jopling et al., 2010). Thus, it will be interesting in the future to test if destruction of centrosome integrity will indeed abolish the ability of zebrafish to regenerate their heart.

Taken together, this study suggests that relocalization of the MTOC disrupts centrosome integrity which, in turn, promotes a post-mitotic state in mammalian cardiomyocytes. Given the increasing role of the centrosome in cell cycle control, understanding how centrosome integrity is regulated during development may reveal new mechanisms to regulate cell proliferation with implications for regeneration and cancer.

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Author details

1. David C Zebrowski

   1. Experimental Renal and Cardiovascular Research, Department of Nephropathology, Institute of Pathology, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany
   2. Department of Cardiac Development and Remodeling, Max Planck Institute for Heart and Lung Research, Bad Nauheim, Germany

   Contribution

   DCZ, Conception and design, Acquisition of data, Analysis and interpretation of data, Drafting or revising the article

   For correspondence

   david.zebrowski@gmail.com

   Competing interests

   The authors declare that no competing interests exist.

2. Silvia Vergarajauregui

   Experimental Renal and Cardiovascular Research, Department of Nephropathology, Institute of Pathology, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany

   Contribution

   SV, Conception and design, Acquisition of data, Analysis and interpretation of data

   Competing interests

   The authors declare that no competing interests exist.

3. Chi-Chung Wu

   Institute for Biochemistry and Molecular Biology, University of Ulm, Ulm, Germany

   Contribution

   C-CW, Conception and design, Acquisition of data, Analysis and interpretation of data

   Competing interests

   The authors declare that no competing interests exist.

4. Tanja Piatkowski

   Department of Cardiac Development and Remodeling, Max Planck Institute for Heart and Lung Research, Bad Nauheim, Germany

   Contribution

   TP, Acquisition of data, Analysis and interpretation of data, Contributed unpublished essential data or reagents

   Competing interests

   The authors declare that no competing interests exist.

5. Robert Becker

   Experimental Renal and Cardiovascular Research, Department of Nephropathology, Institute of Pathology, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany

   Contribution

   RB, Acquisition of data, Analysis and interpretation of data

   Competing interests

   The authors declare that no competing interests exist.

6. Marina Leone

   1. Experimental Renal and Cardiovascular Research, Department of Nephropathology, Institute of Pathology, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany
   2. Department of Cardiac Development and Remodeling, Max Planck Institute for Heart and Lung Research, Bad Nauheim, Germany

   Contribution

   ML, Acquisition of data, Analysis and interpretation of data

   Competing interests

   The authors declare that no competing interests exist.

7. Sofia Hirth

   Department of Medicine II, University of Ulm, Ulm, Germany

   Contribution

   SH, Acquisition of data, Analysis and interpretation of data

   Competing interests

   The authors declare that no competing interests exist.

8. Filomena Ricciardi

   Department of Developmental Genetics, Max Planck Institute for Heart and Lung Research, Bad Nauheim, Germany

   Contribution

   FR, Acquisition of data, Analysis and interpretation of data

   Competing interests

   The authors declare that no competing interests exist.

9. Nathalie Falk

   Department of Biology, Animal Physiology, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany

   Contribution

   NF, Acquisition of data, Analysis and interpretation of data

   Competing interests

   The authors declare that no competing interests exist.

10. Andreas Giessl

    Department of Biology, Animal Physiology, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany

    Contribution

    AG, Analysis and interpretation of data, Contributed unpublished essential data or reagents

    Competing interests

    The authors declare that no competing interests exist.

11. Steffen Just

    Department of Medicine II, University of Ulm, Ulm, Germany

    Contribution

    SJ, Analysis and interpretation of data, Drafting or revising the article

    Competing interests

    The authors declare that no competing interests exist.

12. Thomas Braun

    Department of Cardiac Development and Remodeling, Max Planck Institute for Heart and Lung Research, Bad Nauheim, Germany

    Contribution

    TB, Analysis and interpretation of data, Contributed unpublished essential data or reagents

    Competing interests

    The authors declare that no competing interests exist.

13. Gilbert Weidinger

    Institute for Biochemistry and Molecular Biology, University of Ulm, Ulm, Germany

    Contribution

    GW, Conception and design, Analysis and interpretation of data, Drafting or revising the article

    Competing interests

    The authors declare that no competing interests exist.

14. Felix B Engel

    1. Experimental Renal and Cardiovascular Research, Department of Nephropathology, Institute of Pathology, Friedrich-Alexander-Universität Erlangen-Nürnberg, Erlangen, Germany
    2. Department of Cardiac Development and Remodeling, Max Planck Institute for Heart and Lung Research, Bad Nauheim, Germany

    Contribution

    FBE, Conception and design, Analysis and interpretation of data, Drafting or revising the article

    For correspondence

    felix.engel@uk-erlangen.de

    Competing interests

    The authors declare that no competing interests exist.

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