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Notch and Melanocytes: Diverse Outcomes from a Single Signal

2008; Elsevier BV; Volume: 128; Issue: 11 Linguagem: Inglês

10.1038/jid.2008.289

1523-1747

Masatake Osawa, David E. Fisher,

Hedgehog Signaling Pathway Studies

Notch signaling is an evolutionally conserved pathway that serves as a critical regulator of cell fate. From a series of studies, including a report in this issue, researchers have begun to elucidate the critical functions of Notch signaling in the regulation of melanocyte lineage development. With evidence of a recently identified role for Notch signaling in melanomagenesis, characterization of downstream molecular events may offer potential avenues for the development of novel therapeutic strategies. Notch signaling is an evolutionally conserved pathway that serves as a critical regulator of cell fate. From a series of studies, including a report in this issue, researchers have begun to elucidate the critical functions of Notch signaling in the regulation of melanocyte lineage development. With evidence of a recently identified role for Notch signaling in melanomagenesis, characterization of downstream molecular events may offer potential avenues for the development of novel therapeutic strategies. Tissue organogenesis is governed by a remarkably small set of evolutionally conserved signaling pathways. These signals, which may be termed a "developmental toolkit," include the BMP, FGF, Notch, Hedgehog, and Wnt signaling pathways (Canestro et al., 2007Canestro C. Yokoi H. Postlethwait J.H. Evolutionary developmental biology and genomics.Nat Rev Genet. 2007; 8: 932-942Crossref PubMed Scopus (101) Google Scholar). During development, these toolkit signaling pathways are utilized in a diverse range of discrete cell lineages in a spaciotemporally regulated manner to build a wide variety of morphologically and functionally distinct tissues. One central question in developmental biology is how such a small number of toolkit pathways can generate the huge diversity seen in tissue development. To accomplish this diversity, the signals appear to engage in cross-talk with other signaling pathways, allowing them to be integrated into a network of large signaling complexes, which may generate diversity from individual signaling events. The genome sequencing project has helped to identify each component in the signaling networks. Researchers in the postgenomic era are trying to "connect the dots" to understand how networks interact. Such efforts will lead to a more comprehensive understanding of signaling networks and the maintenance of healthy homeostasis. Because disruption of these networks may be pathogenic, a detailed understanding of these signaling effects may have broad implications, not only for developmental biology but also for the discovery of novel therapeutic strategies. Notch compromises a family of evolutionarily conserved receptors whose activation is mediated by specific cell–cell interaction with cells expressing the ligands Jagged or Delta (Artavanis-Tsakonas et al., 1999Artavanis-Tsakonas S. Rand M.D. Lake R.J. Notch signaling: cell fate control and signal integration in development.Science. 1999; 284: 770-776Crossref PubMed Scopus (4896) Google Scholar). Upon activation, the intracellular domain of the Notch receptor is cleaved and translocated into the nucleus to generate a transactivation complex with the RBP-J transcription factor. The resulting complex initiates transactivation of various target genes, including the hairy/enhancer of split (Hes) transcriptional repressors (Artavanis-Tsakonas et al., 1999Artavanis-Tsakonas S. Rand M.D. Lake R.J. Notch signaling: cell fate control and signal integration in development.Science. 1999; 284: 770-776Crossref PubMed Scopus (4896) Google Scholar). The Hes family repressor represents a bona fide target of Notch signaling and plays a central role downstream in the Notch pathway. Hes family repressors antagonize the expression of a wide variety of activator-type bHLH transcription factors, including Ascl1, Atoh1, and Neurog3, which is critical for maintaining cells in an undifferentiated state (Kageyama et al., 2007Kageyama R. Ohtsuka T. Kobayashi T. The Hes gene family: repressors and oscillators that orchestrate embryogenesis.Development. 2007; 134: 1243-1251Crossref PubMed Scopus (490) Google Scholar). In addition to this canonical pathway, an RBP-J-independent noncanonical pathway and a Hes-independent signaling pathway also play important roles downstream of Notch signaling, although the exact molecular events mediating these noncanonical pathways are less fully characterized. Despite a considerable understanding of signal transduction by Notch, its outcomes are complex. One important feature of Notch signaling is that the outcome of this pathway is highly dependent on cell context. This could be partly explained by its ability to cross-talk with diverse signaling pathways (Hurlbut et al., 2007Hurlbut G.D. Kankel M.W. Lake R.J. Artavanis-Tsakonas S. Crossing paths with Notch in the hyper-network.Curr Opin Cell Biol. 2007; 19: 166-175Crossref PubMed Scopus (199) Google Scholar). In fact, it has been demonstrated that Notch signaling may indeed undergo cross-talk with nearly all components of the developmental toolkit (Hurlbut et al., 2007Hurlbut G.D. Kankel M.W. Lake R.J. Artavanis-Tsakonas S. Crossing paths with Notch in the hyper-network.Curr Opin Cell Biol. 2007; 19: 166-175Crossref PubMed Scopus (199) Google Scholar). Accumulating evidence indicates pleiotropic roles for Notch signaling in diverse cellular processes, including cell cycle arrest regulation, apoptosis/survival, differentiation, and stem cell maintenance. Moreover, recent studies have revealed cross-talk between Notch signaling and the hypoxia response (Gustafsson et al., 2005Gustafsson M.V. Zheng X. Pereira T. Gradin K. Jin S. Lundkvist J. et al.Hypoxia requires notch signaling to maintain the undifferentiated cell state.Dev Cell. 2005; 9: 617-628Abstract Full Text Full Text PDF PubMed Scopus (897) Google Scholar). Given that many diverse cellular processes may need to be accompanied by regulation of cellular metabolism (or, conversely, that metabolic changes may require compensatory changes in diverse cellular pathways), it may be notably efficient that Notch signaling simultaneously regulates these distinct cellular events. An obvious question arises: how can Notch signaling mechanistically accomplish these versatile functions? One fresh insight has been obtained from a recent analysis of epidermal development (Moriyama et al., 2008Moriyama M. Durham A.D. Moriyama H. Hasegawa K. Nishikawa S. Radtke F. et al.Multiple roles of Notch signaling in the regulation of epidermal development.Dev Cell. 2008; 14: 594-604Abstract Full Text Full Text PDF PubMed Scopus (118) Google Scholar). In epidermal development, Notch signaling induces granular cell differentiation from spinous cells but simultaneously prevents premature differentiation of spinous cells (Moriyama et al., 2008Moriyama M. Durham A.D. Moriyama H. Hasegawa K. Nishikawa S. Radtke F. et al.Multiple roles of Notch signaling in the regulation of epidermal development.Dev Cell. 2008; 14: 594-604Abstract Full Text Full Text PDF PubMed Scopus (118) Google Scholar). In this setting, Notch signaling through the RBP-J transactivator directly promotes expression of the Ascl2 bHLH transcription factor, which is responsible for induction of granular differentiation. On the other hand, Notch signaling through the Hes1 repressor simultaneously suppresses Ascl2 expression by interfering with the RBP-J-mediated transactivation function, which is essential for maintaining the undifferentiated status of spinous cells (Moriyama et al., 2008Moriyama M. Durham A.D. Moriyama H. Hasegawa K. Nishikawa S. Radtke F. et al.Multiple roles of Notch signaling in the regulation of epidermal development.Dev Cell. 2008; 14: 594-604Abstract Full Text Full Text PDF PubMed Scopus (118) Google Scholar). These findings indicate that, in addition to an ability to cross-talk with various signaling pathways, the simultaneous existence of both a transcriptional activator and a repressor downstream of Notch signaling may play an important role in the versatile functions of Notch signaling. More detailed studies may help to determine how Notch signaling controls these outcomes. During development, melanoblasts–the precursors of pigmented melanocytes–arise in the neural crest and migrate through the epidermis to newly forming hair follicles. Once in the follicle, they eventually segregate into two populations: melanocyte stem cells, which colonize the lower permanent portion of the hair follicle (the bulge region) and maintain the hair pigmentary unit for subsequent hair cycles, and differentiated melanocytes in the hair matrix (the bulb region), responsible for pigmentation of the growing hairs. In non-hairy regions, melanocytes reside deep in the epidermis (near the dermal/epidermal junction), where they respond to differentiation/pigmentation cues from surrounding keratinocytes or other environmental signals. It has been widely accepted that significant homeostatic regulation of melanocytes may occur through cell–cell interactions between keratinocytes and melanocytes (Hirobe, 2005Hirobe T. Role of keratinocyte-derived factors involved in regulating the proliferation and differentiation of mammalian epidermal melanocytes.Pigment Cell Res. 2005; 18: 2-12Crossref PubMed Scopus (234) Google Scholar), whereas the underlying molecular mechanisms are incompletely understood. A series of recent studies elucidated Notch signaling as a key component among keratinocyte–melanocyte interactions (Kumano et al., 2008Kumano K. Masuda S. Sata M. Saito T. Lee S.Y. Sakata-Yanagimoto M. et al.Both Notch1 and Notch2 contribute to the regulation of melanocyte homeostasis.Pigment Cell Melanoma Res. 2008; 21: 70-78Crossref PubMed Scopus (64) Google Scholar; Moriyama et al., 2006Moriyama M. Osawa M. Mak S.S. Ohtsuka T. Yamamoto N. Han H. et al.Notch signaling via Hes1 transcription factor maintains survival of melanoblasts and melanocyte stem cells.J Cell Biol. 2006; 173: 333-339Crossref PubMed Scopus (193) Google Scholar; Schouwey et al., 2007Schouwey K. Delmas V. Larue L. Zimber-Strobl U. Strobl L.J. Radtke F. et al.Notch1 and Notch2 receptors influence progressive hair graying in a dose-dependent manner.Dev Dyn. 2007; 236: 282-289Crossref PubMed Scopus (97) Google Scholar). Now Aubin-Houzelstein et al. (Aubin-Houzelstein et al., 2008Aubin-Houzelstein G. Djian-Zaouche J. Bernex F. Gadin S. Delmas V. Larue L. et al.Melanoblasts' proper location and timed differentiation depend on Notch/RBP-J signaling in postnatal hair follicles.J Invest Dermatol. 2008; 128: 2686-2695Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar, this issue) extend these studies and add several new implications for the function of Notch signaling in regulation of the melanocyte lineage within the hair follicle. Notch signaling plays an indispensable role in the maintenance of melanoblasts and melanocyte stem cells of the epidermis (Moriyama et al., 2006Moriyama M. Osawa M. Mak S.S. Ohtsuka T. Yamamoto N. Han H. et al.Notch signaling via Hes1 transcription factor maintains survival of melanoblasts and melanocyte stem cells.J Cell Biol. 2006; 173: 333-339Crossref PubMed Scopus (193) Google Scholar; Schouwey et al., 2007Schouwey K. Delmas V. Larue L. Zimber-Strobl U. Strobl L.J. Radtke F. et al.Notch1 and Notch2 receptors influence progressive hair graying in a dose-dependent manner.Dev Dyn. 2007; 236: 282-289Crossref PubMed Scopus (97) Google Scholar). Genetic ablation of Notch signaling in the mouse results in a dramatic reduction of embryonic melanoblasts (via apoptosis), which manifests itself as dilution of initial hair pigmentation (Moriyama et al., 2006Moriyama M. Osawa M. Mak S.S. Ohtsuka T. Yamamoto N. Han H. et al.Notch signaling via Hes1 transcription factor maintains survival of melanoblasts and melanocyte stem cells.J Cell Biol. 2006; 173: 333-339Crossref PubMed Scopus (193) Google Scholar; Schouwey et al., 2007Schouwey K. Delmas V. Larue L. Zimber-Strobl U. Strobl L.J. Radtke F. et al.Notch1 and Notch2 receptors influence progressive hair graying in a dose-dependent manner.Dev Dyn. 2007; 236: 282-289Crossref PubMed Scopus (97) Google Scholar). These animals also exhibit premature hair graying in subsequent hair cycles, suggesting an important role for Notch signaling in the maintenance of melanocyte stem cells (Moriyama et al., 2006Moriyama M. Osawa M. Mak S.S. Ohtsuka T. Yamamoto N. Han H. et al.Notch signaling via Hes1 transcription factor maintains survival of melanoblasts and melanocyte stem cells.J Cell Biol. 2006; 173: 333-339Crossref PubMed Scopus (193) Google Scholar; Schouwey et al., 2007Schouwey K. Delmas V. Larue L. Zimber-Strobl U. Strobl L.J. Radtke F. et al.Notch1 and Notch2 receptors influence progressive hair graying in a dose-dependent manner.Dev Dyn. 2007; 236: 282-289Crossref PubMed Scopus (97) Google Scholar). The latter function of Notch signaling has recently been further supported by the pharmacologic inhibition of Notch signaling in adult mice (Kumano et al., 2008Kumano K. Masuda S. Sata M. Saito T. Lee S.Y. Sakata-Yanagimoto M. et al.Both Notch1 and Notch2 contribute to the regulation of melanocyte homeostasis.Pigment Cell Melanoma Res. 2008; 21: 70-78Crossref PubMed Scopus (64) Google Scholar). In this issue, Aubin-Houzelstein et al. describe their use of an elegant in vivo marking technique to examine how loss of Notch signaling affects the behavior of follicular melanocytes/melanoblasts. This permitted the investigators to identify several previously unrecognized abnormalities within the follicular melanocyte lineage, including premature differentiation and mislocalization of melanoblasts and abnormal differentiation of hair matrix melanocytes. These findings indicate that, in addition to maintenance of melanoblasts and melanocyte stem cells, Notch signaling plays roles in maintaining the immature status of melanoblasts, promoting proper localization of melanoblasts, and inducing proper melanocyte differentiation in the hair matrix (Aubin-Houzelstein et al., 2008Aubin-Houzelstein G. Djian-Zaouche J. Bernex F. Gadin S. Delmas V. Larue L. et al.Melanoblasts' proper location and timed differentiation depend on Notch/RBP-J signaling in postnatal hair follicles.J Invest Dermatol. 2008; 128: 2686-2695Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar). The premature differentiation phenotype of Notch-deficient melanoblasts is reminiscent of Mitfvit/vit mutant mice, a mild hypomorphic mutation in the gene encoding the Mitf transcription factor (Nishimura et al., 2005Nishimura E.K. Granter S.R. Fisher D.E. Mechanisms of hair graying: incomplete melanocyte stem cell maintenance in the niche.Science. 2005; 307: 720-724Crossref PubMed Scopus (575) Google Scholar). These similarities suggest a link between Notch signaling and Mitf in the regulation of melanoblasts or melanocyte stem cells, although a direct mechanistic link has not yet been identified and the phenotypes could alternatively reflect common downstream effects from interfering with these different factors. Several interesting questions remain unsolved. Why did only a proportion of melanoblasts (∼13%) undergo differentiation, leaving many melanoblasts (∼77%) immature? This incomplete penetrance could reflect the presence of discrete melanocytic subpopulations that were heretofore unrecognized (unless the effect is due to varied completeness of allele knockout). Second, why did Notch-deficient melanocytes differentiate normally in the initial hairs but not in the second hair cycle? Given the dramatic reduction of embryonic melanoblasts, it may be useful toconsider the possibility that certain abnormal phenotypes in the postnatal hair follicles might be related to secondary or unexpected consequences of embryonic melanoblast reduction. Alternatively, it is possible that signaling requirements for the first cycle vary in certain mechanistic ways relative to subsequent cycles. Regardless, the interesting studies of Aubin-Houzelstein et al., 2008Aubin-Houzelstein G. Djian-Zaouche J. Bernex F. Gadin S. Delmas V. Larue L. et al.Melanoblasts' proper location and timed differentiation depend on Notch/RBP-J signaling in postnatal hair follicles.J Invest Dermatol. 2008; 128: 2686-2695Abstract Full Text Full Text PDF PubMed Scopus (59) Google Scholar should trigger additional experiments utilizing postnatal inducible loss-of-function strategies, such as the use of Tyr-CreER transgenic mice (Bosenberg et al., 2006Bosenberg M. Muthusamy V. Curley D.P. Wang Z. Hobbs C. Nelson B. et al.Characterization of melanocyte-specific inducible Cre recombinase transgenic mice.Genesis. 2006; 44: 262-267Crossref PubMed Scopus (120) Google Scholar; Yajima et al., 2006Yajima I. Belloir E. Bourgeois Y. Kumasaka M. Delmas V. Larue L. Spatiotemporal gene control by the Cre-ERT2 system in melanocytes.Genesis. 2006; 44: 34-43Crossref PubMed Scopus (65) Google Scholar), to define precise functions of Notch signaling at discrete developmental stages in the melanocyte lineage.Notch signaling plays multiple roles in the melanocyte lineage. Notch signaling plays multiple roles in the melanocyte lineage. Notch signaling is an essential component of epidermal–melanocyte interactions. Why do epidermal melanocytes require Notch signaling? One possible clue is provided by Schouwey et al., 2007Schouwey K. Delmas V. Larue L. Zimber-Strobl U. Strobl L.J. Radtke F. et al.Notch1 and Notch2 receptors influence progressive hair graying in a dose-dependent manner.Dev Dyn. 2007; 236: 282-289Crossref PubMed Scopus (97) Google Scholar, who demonstrate that epidermal melanocytes are eliminated in the melanocyte-specific Notch1 and Notch2 double-deficient mice, whereas dermal and choroidal melanocytes are left intact. This result suggests that Notch signaling is not required for nonepidermal melanocytes. Given the common origin of epidermal and nonepidermal melanocytes, one possible explanation for this is that Notch signaling may be required to adapt melanoblasts to the epidermal environment. In this regard, it may be noteworthy that the epidermis is thought to be more consistently hypoxic compared with the dermis. Considering the potential cross-talk between Notch signaling and the hypoxia response, it is possible that Notch signaling may provide hypoxia adaptation selectively to epidermal melanoblasts. Melanocytes afford an advantageous model in which to define survival pathway interactions, because genetic alterations controlling melanocyte viability are easily identifiable by coat color phenotypes that do not affect the viability of the animal. Hence, by integrating combinatory gene knockout approaches and phenotype-based screening systems, melanocytes can provide an invaluable tool to identify molecules that interact with Notch signaling. This approach has already been validated. The premature-hair-graying phenotype of Bcl2 knockout mice is rescued in Bcl2−/−Bim−/− double-knockout mice (Bouillet et al., 2001Bouillet P. Cory S. Zhang L.C. Strasser A. Adams J.M. Degenerative disorders caused by Bcl-2 deficiency prevented by loss of its BH3-only antagonist Bim.Dev Cell. 2001; 1: 645-653Abstract Full Text Full Text PDF PubMed Scopus (236) Google Scholar), revealing a striking molecular link between Bcl2 and Bim. Relative to Notch signaling, Kumano et al. (Kumano et al., 2008Kumano K. Masuda S. Sata M. Saito T. Lee S.Y. Sakata-Yanagimoto M. et al.Both Notch1 and Notch2 contribute to the regulation of melanocyte homeostasis.Pigment Cell Melanoma Res. 2008; 21: 70-78Crossref PubMed Scopus (64) Google Scholar) reported premature hair graying in Notch1−/+; Notch2−/+;Wv/+ mice, indicating a possibility of cross-talk between Notch and c-Kit signaling in regulation of the melanocyte lineage. Because melanomas emerge primarily within epidermal melanoblasts or melanocytes, it is reasonable to speculate that both normal and malignant melanoblasts may share key pathways that regulate biological homeostasis and maintenance. These similarities include the epithelial–mesenchymal transition and an invasive/migratory capacity that is central to both normal and malignant cells of the melanocyte lineage. Interestingly, it was recently demonstrated that, as in normal melanoblast development, Notch signaling participates in melanoma progression (Balint et al., 2005Balint K. Xiao M. Pinnix C.C. Soma A. Veres I. Juhasz I. et al.Activation of Notch1 signaling is required for beta-catenin-mediated human primary melanoma progression.J Clin Invest. 2005; 115: 3166-3176Crossref PubMed Scopus (283) Google Scholar; Liu et al., 2006Liu Z.J. Xiao M. Balint K. Smalley K.S. Brafford P. Qiu R. et al.Notch1 signaling promotes primary melanoma progression by activating mitogen-activated protein kinase/phosphatidylinositol 3-kinase-Akt pathways and up-regulating N-cadherin expression.Cancer Res. 2006; 66: 4182-4190Crossref PubMed Scopus (231) Google Scholar). Thus, the elucidation of molecular mechanisms downstream of Notch is important not only for melanoblast development but also for understanding the molecular mechanisms of melanomagenesis. Returning to the original question of how small numbers of signals produce great biological diversity, the fact that Notch signaling is now implicated in multiple distinct melanocyte pathways should provide both a clue and an opportunity. Because the melanocyte system permits genetic alteration without loss of mouse viability, the pigmentary phenotype can be deeply probed in order to identify the "dots" that connect Notch to its interacting signaling pathways. These efforts should ultimately reveal a fascinating network, which may provide answers pertinent for other developmental lineages, as well as clues for novel therapeutics aimed at melanocytic malignancy. M.O. states no conflicts of interest. D.E.F. declares industry consulting relationships with Novartis, Magen Biosciences, and SourceMDx.