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Keywords:

  • deafness;
  • differentiation;
  • G1-phase;
  • neuron;
  • survival

Abstract

  1. Top of page
  2. Abstract
  3. Introduction
  4. Acknowledgments
  5. References

Neuronal network consists of many types of neuron and glial cells. This diversity is guaranteed by the constant cell proliferation of neuronal stem cells following stop cell cycle re-entry, which leads to differentiation during development. Neuronal differentiation occurs mainly at the specific cell cycle phase, the G1 phase. Therefore, cell cycle exit at the G1 phase is quite an important issue in understanding the process of neuronal cell development. Recent studies have revealed that aberrant S phase re-entry from the G1 phase often links cellular survival. In this review we discuss the different types of G1 arrest on the process of neuronal development in Drosophila. We also describe the issue that aberrant S phase entry often causes apoptosis, and the same mechanism might contribute to sensory organ defects, such as deafness.


Introduction

  1. Top of page
  2. Abstract
  3. Introduction
  4. Acknowledgments
  5. References

The human brain consists of more than 100 billion neurons and an even greater number of glial cells. This huge number of diverged cells is produced during only a few months of development. Meanwhile, neuronal precursor cells constantly divide and produce many types of cells, including those of which give rise to neuronal networks. To supply matured neurons and glial cells, neuronal precursor cells must stop their division and differentiate. Therefore, there might be a tight regulation between cell cycle exit and differentiation. The specific cell cycle point of the G1 phase has been shown to have a pivotal role in initiating differentiation events. The studies of cerebral cortical development, for example, a well characterized system, show cell cycle exit at the G1 phase as a cause, rather than a consequence, of the proper population of the neuronal cell types (Caegari & Huttner 2003). During mouse cerebral cortex development, the total length of the neural progenitor cell cycle increased by prolongation of G1-phase, and the G1-lengthing increases the likelihood that daughter cells will exit the cell cycle upon completion of a division. Another study showed that overexpression of G1 cyclin, which caused shortening of the G1 phase, delayed cell cycle exit (Lange et al. 2009; Pilaz et al. 2009). These observations support the idea that G1 phase regulation might have quite an important role for proper neuronal development.

Cell cycle regulation at G1 phase does not only link to the proper developmental process, but also cellular survival. On the course of Purkinje cells development, ectopic expression of E2F-1 caused aberrant S phase entry followed by apoptotic cell death (Athanasiou et al. 1998). In the case of Drosophila, it has been shown that aberrant G1-S progression by overexpressing dE2F1 caused massive apoptosis (Boulton et al. 2000). These pieces of evidence suggest that there might be a common mechanism for the regulation of cellular survival at the G1 phase during neuronal development.

The E2F family of transcription factor plays an important role for G1 progression in the regulation of the cell cycle (Weinberg 1995; Wong et al. 2011). Recent genome-scale measures of gene expression revealed that E2F regulates transcription of many genes important for both S phase and mitosis, including cyclins and cdks (Wong et al. 2011). A growing amount of evidence has shown that cell cycle exit is obtained by both inhibition of Cyclin/Cdk activity by cyclin-dependent kinase inhibitors (CKIs) and the inhibition of E2F transcriptional activity (Brooks et al. 1998). E2F and its partner, DRTF1-polypeptide (DP), has dual function for the transcription; E2F/DP functions as either a transcriptional activator or repressor by associating with co-activators or co-repressors (Cayirlioglu et al. 2003). Among the repressors, Retinoblastoma (Rb) protein or Rb-related pocket proteins inhibits E2F/DP activity by converting co-repressor molecules in the transcriptional complex (Trimarchi & Lees 2002). Notably, a phosphorylated state of Rb, which is done by Cyclin/Cdk complex, regulate the association with E2F, therefore, cell cycle progression often links to the transcriptional activity of E2F.

In this review, we summarize the cell cycle regulation in Drosophila and divergent regulatory systems of the G1 phase during neuronal development. We further discuss the linkage between G1 phase regulation and cellular survival, which might lead to issues of human sensory disorders, such as deafness.

Cell cycle regulations in Drosophila

There appears to be four types of cell cycle regulation during Drosophila development (Fig. 1). Each cell cycle event is highly synchronous, therefore, it is an ideal system to understand the relationship between cell cycle regulation and induction of differentiation. Drosophila first instar larvae consists of approximately 50 000 cells, which are derived from cell division during an approximately 24-h period of embryogenesis. The early pattern formation of Drosophila embryo begins at syncytium, which is derived from 13 nuclear divisions from a fertilized egg. This rapid nuclear division (~10 min intervals) has only DNA synthesis (S) and mitosis (M) (Fig. 1). A G2 phase is emerged after 14 nuclear divisions in embryos (Fig. 1). At this time, G2 to M phase progression is regulated by the patterning cue, which might be coming through string/cdc25 (Edgar & O'Farrel 1990; Edgar et al. 1994). After the 15th cell division, cells are divided into two groups based on the cell cycle patterning. One group consists only of an S and Gap phase and there is no intervening mitosis; this group of cells comprises larval tissue (Fig. 1). In this case, nutritional signaling seems to act through CycE to regulate S phase reentry (Pierce et al. 2004). This system is ideal for producing a large size (or mass) of larval tissue: larvae increase their mass about 200-fold in just 4 days of their development. Another group of cells, such as imaginal discs, germ cells, and stem cells, obtain G1 phase and their division seems to be regulated by the patterning and growth signaling, which might come through E2F and cycE (Prober & Edgar 2000).

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Figure 1. Four different types of cell cycle regulations in Drosophila.

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Figure 2. Components of Rb/E2F/DPs and cyclin-dependent kinase inhibitors (CKIs) in human and Drosophila.

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Regulatory machinery of Rb/E2F/DP pathway

In mammalian cells, there are eight E2F genes, three DP family members (DP1, DP2 and DP3), three Rb families (Rb, p107 and p130) and seven CKIs (p15, p16, p18, p19, p21, p27 and p57) (van den Heuvel & Dyson 2008) (Fig. 2). Each E2F/DP complex is acting as either activator or repressor for the target genes expression. These cell cycle machineries are evolutionarily conserved; Drosophila produces two E2F family proteins, dE2F1 and dE2F2, one Dp (dDp), two Rb family proteins (retinoblastoma family protein 1 [RBF1] and RBF2), and one CKI (dacapo[dap]; a member of p21/p27 family) (Duronio et al. 1995; Du & Dyson 1999; van den Heuvel & Dyson 2008) (Fig. 2). Therefore, Drosophila provides us with a simple system to understand the functions of the Rb/E2F/DP pathway at the G1-S during development.

To understand the regulation of the Rb/E2F/DP pathway, RBF1 associated protein complex has been analyzed, and Drosophila dE2F2, dDP and RBF1 proteins are identified in a large complex including Myb oncogene containing proteins called dREAM/MMB (Drosophila RBF, dE2F2, and Myb-interacting proteins or Myb-MuvB) (Beall et al. 2002; Korenjak et al. 2004; Lewis et al. 2004). The dREAM/MMB complex is evolutionarily conserved; human has an equivalent complex called DREAM/LINC complex (Schmit et al. 2007). A growing body of evidence supports the idea that the dREAM/MMB complex has distinct functions and components in different cellular contexts (Georlette et al. 2007; Debruhl et al. 2013).

Using a dsRNA-based knockdown system, the modifier screening for E2F transcriptional activity has been done, and identified l(3)malignant brain tumor (l(3)mbt) (Lu et al. 2007). Biochemical analysis revealed that L(3)mbt is in the dREAM/MMB complex and regulates cellular growth by modifying transcriptional activity of E2F. L(3)mbt was originally identified as a tumor suppressor gene in the 1970s, and has been shown to have PcG-like chromatin modification activity (Gateff 1975; Tomotsune et al. 1999). These results suggest that epigenetic regulation might have an important role for the control of Rb/E2F activity. To support this idea, several reports showed that the Rb/E2F pathway interacts with Polycomb Group (PcG) silencing complexes in Drosophila. The PcG silencing complex maintains a repressed state of gene expression by regulating the methylated state of histone H3, and this complex plays important roles in epigenetic processes such as Hox gene repression, genomic imprinting, stem cell maintenance, and X chromosome inactivation (Simon & Kingston 2009). It has been reported that Polycomb responsive elements (PRE), a DNA sequence that is recognized by the PcG complex, have been identified in the promoter and coding region of several cell-cycle related genes, and PcG proteins are indeed found at the promoters of dCycB, dDp, dE2F1, and Rbf1 in Drosophila embryo (Martinez et al. 2006; Oktaba et al. 2008). To support this result, Ji et al. performed modifier screening of loss-of E2F1 activity using a dsRNA based system, and identified several PcG complex genes (Ji et al. 2012). They found that inhibition of the PcG molecule upregulated several E2F1 target genes expression, suggesting that PcG downregulates E2F1 target genes. These studies support the idea that PcG is regulating cell proliferation through multiple cell cycle genes expression during development.

G1 phase regulation on the process of neural development

A growing number of studies have shown that there is a link between G1 arrest and neural development. Here we would like to take a look at those regulatory systems at the G1 phase during neuronal formation in Drosophila.

Embryo neural development

In the Drosophila central nervous system (CNS), at the stage 9–11 embryo, ventral epidermal cells produce neuroblasts (NBs) (Homen & Knoblich 2012). After delamination from epidermal cells, NBs start dividing shortly afterwards to generate another NB and smaller ganglion mother cell (GMC) (Fig. 3A). GMCs produce ganglion cells (GCs), which become neuron or glia cells after dividing once. Afterwards GCs produce neurons. In the previous study, it has been shown that overexpressing cycE caused self-renewing GMCs in the embryo, suggesting that G1 arrest might have an important role for the self-renewing GMC. The cell cycle exit and initiation of differentiation is performed by the homeodomain protein, Prospero (Pros). Pros is proposed to arrest cell cycle in GMCs by inhibiting the expression of cell-cycle regulators such as cyclin A, cyclin E and E2F (Lee & Vaessin 2000) (Fig. 3B). Recent study has provided a more complicated situation: cell cycle independent function of cycE inhibits the function of Pros and facilitates its cellular localization (Berger et al. 2010). These results suggest that there might be a reciprocal regulatory system between cycE and Pros.

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Figure 3. Neuronal diversity and cell cycle regulation at G1 in Drosophila. (A) A Drosophila neurogenesis in embryo. Neuroblast (NB) delaminates from neuroepithelia. NB divides to generate a ganglion mother cell (GMC) that divides once more, producing two neurons. (B) Dual function of Prospero (Pros) against different target genes. (C) A brain at 3rd instar larva. Ventral nerve cord (VNC) consist of thoracic neuroblasts (thoracic NBs) and abdominal NBs. Brain lobes consist of optic lobe NBs (OL NBs), central brain mushroom body NBs (MB NBs), type I NBs, and type II NBs. (D) A cell division pattern of Type I NB in larval brain. (E) A G1 phase regulation of GCs in type I NBs by Pros. DPN, Deadpan.

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Surprisingly, Pros is necessary for the expression of a subset of differentiation genes, such as zfh1 and Lim1, which specify neuronal subtype, and the adhesion molecules Facicllin I (Fas I), Fasciclin II (Fas II) and Netrin-B, those of which have roles in axon guidance (Choksi et al. 2006) (Fig. 3B). These data suggest that Pros is an activator for transcription of differentiation genes, and acts as a binary switch between the neural stem cell fate and the terminally differentiated neuronal fate.

Neural development in larvae

Extensive studies have been done for analyzing the neuronal stem cell regulation using postembryonic stage. There seem to be many types of NBs in the larval brain: abdominal and thoracic NBs in the ventral nerve cord (VNC) and type I, type II, mushroom body (MB) and optic lobe NBs in the brain lobes (Homen & Knoblich 2012) (Fig. 3C). Among the NBs in larval brain, type I and II NBs have been well characterized by their cell cycle and differentiation. Especially, type I NBs show similar characteristic properties with the embryonic NBs and divide once to generate two GCs that differentiate into neuron and glia (Fig. 3D). Many studies indicate that Pros is expressing and acts both in type I NBs and their GMC cells, and a high level of Pros has been observed in GCs (Colonques et al. 2011). In the embryonic stage, however, Pros is expressed only in NBs and GMC. These discrepancies suggest that there is distinct activity of Pros in the larval NBs.

The expression level of dap seems to have an important role for the G1 arrest at the GCs (Colonques et al. 2011). Dap is expressing not only at GCs, but NBs and GMCs of type I NBs; however, the expression level in the GCs seems to be upregulated by Pros. Detailed analysis showed that Pros inhibits the expression of deadpan (dpn), which encodes bHLH transcriptional factor. Dpn has been shown to inhibit the expression of dap, therefore Pros might use the double negative regulation to inhibit cell cycle progression at the G1 phase (Fig. 3E).

Photoreceptor cells development

The compound eye of Drosophila has been used as a model system for studying cell cycle regulation during developmental processes (Wolff & Ready 1993). The compound eye consists of about 800 units (ommatidia) arranged in a regular hexagonal array. Each ommatidium has eight photoreceptor neurons (R1–R8), together with four cone cells (which secrete the lens) and additional pigment cells. The eye develops from the undifferentiated single layered epithelial sheet of the eye imaginal disc. During the middle of the third instar larva, specification of the eye begins at the posterior of the eye disc and progresses anteriorly (Wolff & Ready 1991) (Fig. 4). This wave progression, called morphogenetic furrow (MF), seems to take about 2 days from posterior to anterior of the eye disc (Fig. 4). Anterior to the MF, undifferentiated cells randomly proliferate, while on the MF, all the cells synchronize at the G1 phase for about 5–6 h. Once the MF progresses, cells can be divided into two groups: one group is in “preclusters” that initiate the omatidial recruitment, and the other is undifferentiated cells. These undifferentiated cells undergo one final and simultaneous round of mitosis right after the MF, producing the rest of the cells in the ommatidia. This synchronous cell division is called the second mitotic wave (SMW) (Wolff & Ready 1991, 1993) (Fig. 4).

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Figure 4. Cell cycle regulation during development of the eye imaginal disc in Drosophila third instar larvae. (Left in figure) eye imaginal disc stained by anti-Elav (pan-neural marker; green) and Cut (cone cells marker; magenta). (Right in figure) A schematic view around the morphogenetic furrow (MF) (from white square with dotted line in left figure). Ahead of the MF, cells are randomly dividing. Just ahead of the MF, cells become synchronized in mitosis by synchronous expressing string/cdc25. On the MF, all the cells are held in G1. Behind the MF, cells either directly integrated into the pre-cluster formation (differentiating state) or enter a final synchronous S phase (second mitotic wave: SMW), which is followed by subsequent differentiation. Yellow cells in pre-cluster formation represent developing R8 cells. Green cells represent Elav-positive photoreceptor cells.

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The initiation of photoreceptor cells development starts with specification of R8 cells, those of which are expressing the transcription factors Atonal (Ato) and Senseless (Sens) (Frankfort & Mardon 2002). On the way from precluster to the R8 cells specification, there are a number of factors, that are required for the transcriptional regulation of Ato and Sens. Among them, Egfr and Notch signaling have an important role for the initial events of cluster formation: Egfr signaling is required for the proper spacing of each precluster cell, while Notch signaling seems to act in lateral inhibition of R8 specification.

In these two decades, the bulk of investigation has been done to reveal the mechanism of G1 arrest at MF. The regulatory mechanism against CycE, cycA and E2F seems to play a key role for the G1 arrest on the MF, and consequence inducing SMW. During eye morphogenesis secretion molecules, Hedgehog (Hh) and Decapentapledgic (Dpp), are expressed around the furrow (Firth & Baker 2005). When the furrow approached, these secretion factors caused synchronized mitosis through the cdc25/string at the posterior domain of MF, and all the cells enter the G1 phase at the furrow (Escudero & Freeman 2007). In this time Dpp signaling downregulates cycE expression and E2F activity. Notch signaling also seems to be involved in the G1 arrest at MF by de-repressing the inhibition of dE2F1 with RBF (Baonza & Freeman 2005). There seems to be an interplay between Notch and Hh/Dpp; Notch ligand Delta (Dl) is induced by hh/dpp around the furrow. Robust G1 arrest seems to be guaranteed by Hh activity secreted after the furrow by upregulating dap expression. Combinatorial action of these factors might contribute as all the cells at the furrow are synchronized at G1 phase.

Synchronous G1 arrest at the MF might contribute to the proper photoreceptors development. This notion was clarified by analyzing the function of roughex (rux) during eye development. In rux mutation ectopic S phase entry can be observed at the MF, and as a consequence, proper differentiation was prohibited and caused abnormal eye morphologies (Thomas et al. 1994). In rux clone cycA expression level was increased, which promotes S phase entry at the MF (Thomas et al. 1994, 1997). The inhibitory activity of rux against cycA expression seems to be independent with Dpp/Hh signaling, implying that cycA expression is not dependent on cycE and dE2F1 (Escudero & Freeman 2007). Interestingly, recent study revealed that the rux/cycA activity might also be required for the inhibition of cell cycle re-entry for R8 photoreceptor neurons (Ruggiero et al. 2012).

Ebi inhibits aberrant growth signaling

To identify regulatory factors that contribute to the G1 phase regulation at the MF by rux, a genetic modifier screen was performed using rux mutant, and ebi was identified as an enhancer of rux (Dong et al. 1999). Ebi is a Drosophila homologue of trunsducin β-like protein 1 and its related protein (TBL1/TBR1), which are F-box/WD-40-containing proteins (Dong et al. 1999; Perissi et al. 2004). Extensive studies on the function of ebi have shown that ebi has multiple functions during development. For example, Ebi acts as an E3 ubiquitin ligase that targets Tramtrack88, a repressor of neuronal development (Dong et al. 1999). It also functions as a transcriptional co-repressor by forming a complex with many types of transcription factors, such as the ecdysone receptor, Suppressor of Hairless protein, or activator protein-1 (AP-1) (Tsuda et al. 2002, 2006; Lim et al. 2012). These multiple functions of ebi appear to be evolutionarily conserved; TBL1 seems to have a function as an ubiquitin-ligase, in addition to the co-repressor complex activity (Matsuzawa & Reed 2001; Perissi et al. 2004).

Previously, ebi was also identified as an enhancer of abnormal eye morphology caused by the ectopic expression of dE2f1 (Boulton et al. 2000). Notably, in that study it has been shown that ebi mutant embryo increased S phase entry after the 16th cell division cycles (Boulton et al. 2000). This supports the idea that ebi may act at G1-S in many situations. In terms of cell cycle regulation, recent biochemical analysis revealed that Ebi is forming a complex with RBF1 and inhibits the expression of target genes Rbf/dE2F1 pathway (Lim et al. 2013). However, Ebi did not seem to associate with L(3)mbt, suggesting that Ebi/RBF/E2F is a distinct complex to the dREAM/MMB complex acting in the G1 phase.

Interestingly, expression study suggests that Ebi seems to have multiple roles in the regulation of cell cycle progression (Lim et al. 2013). Reducing ebi activity showed downregulation of cell cycle related genes, such as Rbf and rux. Inhibition of Psc, one of the major components of PcG, alleviates the reduction of Rbf and rux expression in ebi mutant (Lim et al. 2013) (Fig. 5A). Two recent studies showed that ebi seems to have an antagonistic role against PcG silencing activity (Strubbe et al. 2011; Lim et al. 2013). Given the fact that Rbf and rux are directly silenced by PcG, ebi might regulate expression of those cell cycle inhibitors by modulating activity of PcG (Fig. 5B). All of these results suggest that ebi might have multiple functions against RBF/E2F, and contribute to the cell cycle regulation at the G1-S transition. Notably, it has been shown that the expression level of ebi seems to be regulated by the activity of E2F, suggesting that there might be a regulatory loop between ebi and Rbf/E2F in the cell cycle regulation (Buttita et al. 2010).

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Figure 5. ebi regulates the expression of rbf and rux through inhibition of Psc. (A) Semi-quantitative polymerase chain reaction (PCR) of mRNA from S2 cells treated with vector, ebi RNAi, and/or Psc RNAi. The primers used in this analysis are indicated. (Modified from Lim et al. 2013). (B) Model of Ebi in the regulatory networks.

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Although the mutation of ebi did not show strong cell cycle defects during eye development, a recent study found that ebi might be acting at G1-S transition under the excess growth stimulation (Lim et al. 2013). Ebi showed strong genetic interaction with Ellipse (Elp), a gain-of-function mutation of Egfr (Baker & Rubin 1989). Egfr signaling itself might be involved in the precluster spacing; however, Elp shows an unbalanced number of cells between the S phase entering cells and precluster forming cells at the SMW (Fig. 6). In this case, precluster formation was severely inhibited, while S phase entering cells at the SMW were increased (Fig. 6). Increasing Notch signaling might be involved in the ectopic induction of S phase in Elp. Notch activation can induce SMW by supporting expression of cycA, and recent studies revealed that Dl is prolonged around the MF in Elp mutation (Baonza & Freeman 2005; Lim et al. 2013).

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Figure 6. ebi strongly enhances pattern formation, cell division, and cell death phenotypes of Ellipse. (A–D) Scanning electron micrographs of the adult compound eye. (E–H) BrdU incorporation (white arrows) and immunohistochemical staining using anti-Elav (black arrows). White arrowheads indicate the position of morphogenetic furrow (MF), and white arrows indicate the position of the second mitotic wave. (I–L) Schematic models of cell cycle regulation. Dif, differentiated cells. (A, E, I) Wild type. (B, F, J) ElpB1/ElpB1. (C, G, K) ElpB1/ElpB1; gl-p21/gl-p21. Human p21 was expressed after the MF by glass-promoter (gl) in Elp mutant. (D, H, L) ElpB1/ElpB1; ebi4/+. (Modified from Lim et al. 2013).

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Reducing ebi strongly enhanced Elp phenotype; photoreceptor cells development was severely prohibited and most of the cells entered the S phase after the MF (Lim et al. 2013) (Fig. 6). The results support the idea that there might be a link between cell cycle progression and differentiation, and the activity of ebi prohibits the excess growth signaling, which perturbs the proper balance.

G1 arrest and sensory cells survival

In Elp, excess apoptosis was also observed after the MF, therefore, Elp showed small eye phenotype (Baker & Rubin 1992) (Fig. 6). Ectopic induction of S-phase after the MF might contribute to the cell death induction, since overexpression of human p21, which prevented S phase entry, completely inhibited this apoptotic induction (Lim et al. 2013) (Fig. 6). To support this idea, Elp mutation with ebi heterozygote, in which most cells were entering S phase after the MF, showed massive apoptosis and severe eye defects (Fig. 6). These results suggest that aberrant progression from the G1 to S phase might induce apoptotic cell death in sensory cells.

Ectopic induction of apoptosis along with aberrant S phase entry can also be observed in mammalian sensory system. Cochlea consists of approximately 12 000 outer hair cells (OHCs), 3500 inner hair cells (IHCs) and support cells, which are well arrayed along the basement membrane of cochlea. The precursor cells of hair cells and their support cells are developed from ear vesicle, which is derived from placode at embryonic stage 4 (E4) (Chen & Segil 1999). Those precursor cells are randomly dividing until embryonic E14 stage, when the expression level of CKI, Kip1 (p27) increases in the primordium of hair cells and support cells (Fig. 7A). As a consequence, all the primordia are arrested at G1 phase. There seems to be two groups in this G1-arrested primordium. One group of cells keeps expressing Kip1, and they become support cells for the hair cells (Fig. 7A). Another group of cells reduces Kip1 expression and starts expressing Ink4d (p19) and MyoVIIA (a marker for the hair cells) and become hair cells (Fig. 7A). In this process, inhibition of S phase re-entry by Ink4d (p19) has an important role for the proper differentiation, since knockout mouse of Ink4d caused S phase re-entry at the hair cells, and those cells are removed afterward by apoptosis (Chen et al. 2003). Thus, Ink4d-KO mouse showed progressive hearing loss phenotype. Laine et al. showed that the co-deletion of Ink4d and Cip1 (p21), accelerated the S phase re-entry, suggesting that Ink4d and Cip1 might have a redundant activity for the G1 arrest of final differentiating hair cells (Laine et al. 2007). Interestingly, the cells entering S phase in this double KO mouse showed increasing DNA damage response (Fig. 7B). This suggests that checkpoint mechanism inhibiting DNA damage response, might contribute to the survival of sensory cells.

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Figure 7. G1 phase regulation tightly linked to the progressive deafness formation. (A) A development of hair cells in the cochlea. At the final stage of development, Ink4 and MyoVIIa positive cells become inner hair cells (IHCs) and outer hair cells (OHCs). (B) Ink4 and Cip1 double mutation showed progressive apoptosis at cochlea hair cells and caused progressive deafness. (C) Model of modifications in DNA-damage response when co-repressor function is removed. (Modified from Scafoglio et al. 2013).

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Recently, it has been shown that the activity of TBL1 containing co-repressor complex has a regulatory function under DNA damage response (Scafoglio et al. 2013). TBL1 was identified in the complex with checkpoint kinase 2 (Chk2), which is a major component of checkpoint mechanism under DNA damage response. Inhibition of the co-repressor activity enhanced DNA damage response and upregulated apoptosis related genes through AP-1 transcription factor; it implicated that co-repressor complex is acting as a “brake” in the DNA damage response (Fig. 7C). Interestingly, TBL1 was identified as a causative factor for a human age-related hearing disorder, called ocular albinism with late-onset sensorineural deafness (OASD) (Bassi et al. 1999; Dong et al. 1999). In the patients of OASD, there is a small deletion in the TBL1 locus which caused C-terminal truncation (Bassi et al. 1999). This suggests that a member of the co-repressor complex, TBL1, might be required for the survival of sensory cells. Recent study suggests that the co-repressor activity in the sensory cells survival might be evolutionarily conserved; ebi mutation caused age-dependent retinal degeneration phenotype in Drosophila (Lim et al. 2012). Introducing a similar mutation of TBL1 observed in OASD patient into Ebi (C-terminal truncation) caused age-dependent retinal degeneration, and Ebi cDNA completely rescued the phenotype. These results suggest that Ebi and TBL1 share the mechanism, which is required for the sensory cells survival (Lim et al. 2012).

Conclusions and future perspectives

Different types of regulation at the G1 phase seem to be involved in the diverged neuronal development in Drosophila. Disintegration of the G1 phase regulation often caused aberrant S phase entering and lead to apoptotic induction. A co-repressor molecule, ebi, seems to be acting in these events; ebi is involved in the regulation of the G1-S transition and inhibits apoptosis induction in sensory neurons. Therefore, deepening of our understanding of the regulation system for this co-repressor molecule in the process of development might lead to the comprehensive understanding of the precise developmental processes involved. Interestingly, many studies have revealed that ebi is acting under many types of cellular signaling, such as Egfr, Notch, and Wingless (Dong et al. 1999; Tsuda et al. 2002, 2006; Li & Wang 2008). However, the precise relationship between ebi and those signaling pathways remain to be clarified. Biochemical analysis of the regulatory system of Ebi activity under those signaling pathways might provide us with a comprehensive view of how extra cellular signaling contributes to the epigenetic regulation of the target genes expression.

Given the fact that Ebi and TBL1 is an evolutionally conserved molecule, the study about ebi function in the photoreceptor survival will extend the therapeutic view of sensory disorders, such as hearing loss.

Acknowledgments

  1. Top of page
  2. Abstract
  3. Introduction
  4. Acknowledgments
  5. References

We acknowledge all members of our laboratory for helpful discussions. This work was supported by a Grant-in-Aid for Scientific Research on Priority Areas from the Ministry of Education, Culture, Sports, Science and Technology of Japan awarded to YM. L. and L.T.

References

  1. Top of page
  2. Abstract
  3. Introduction
  4. Acknowledgments
  5. References
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