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First published online March 21, 2007
doi: 10.1242/10.1242/jcs.005462
Correspondence |
1 Department of Dermatology, Baylor College of Medicine, Houston, TX 77030, USA
2 Department of Molecular and Cellular Biology, Baylor College of Medicine, Houston, TX 77030, USA
* Author for correspondence (e-mail: pkoch{at}bcm.tmc.edu)
The intermediate filament (IF) cytoskeleton of epidermal keratinocytes changes its biochemical composition during the process of terminal differentiation, in which keratinocytes undergo morphological and biochemical changes that ultimately result in the formation of the dead squames that cover the body surface. Epidermal stem cells and proliferating transient amplifying (TA) cells of the basal cell layer express the keratin pair K5/K14. Coinciding with cell cycle exit, these cells cease to express K5/14 and start to synthesize K1/K10. The biological consequence of this change in the keratin expression pattern is the central issue addressed in our previous publication (Chen et al., 2006
) and the above correspondence entitled `The ends of a conundrum?' (Paramio et al., 2007). These authors have published experimental data in support of the hypothesis that K10 expression is causative for the cell cycle exit of differentiating keratinocytes (e.g. Paramio et al., 1999; Paramio et al., 2001; Paramio and Jorcano, 2002; Santos et al., 2002). Furthermore, transgenic experiments by this group suggested that K10 expression suppresses or delays tumor development in mice subjected to a skin carcinogenesis protocol (Santos et al., 1997; Santos et al., 2002). Cell culture experiments subsequently indicated that the K10-mediated inhibition of cell proliferation might be a function of the amino- and carboxyl-terminal end domains of this keratin.
The experiments published so far were done using either cultured cells that normally do not express K10, or transgenic mice that ectopically expressed the full-length K10 protein in the epidermis (including the proliferating compartment) [see references in Koch and Roop (Koch and Roop, 2004)]. We recently decided to test whether K10 end-domain-mediated cell cycle arrest occurs in vivo when these protein domains are expressed in proliferating keratinocytes. To avoid potential toxic effects owing to transgene overexpression, we modified the endogenous K14 gene of mice in such a way that the central K14 rod domain was fused with the K10 head and tail domains, generating a fusion protein termed K1014chim (Chen et al., 2006). The design of this mouse model ensured that the K10 end domains were present in the basal layer of the epidermis and the outer root sheath (ORS) of hair follicles in a 1:1 molar ratio with the K5 end domains. Our results demonstrated that the K5/K1014chim keratin pair formed a functional IF network in keratinocytes. Specifically, we did not observe a reduction in the proliferation of K1014chim keratinocytes when compared with wild-type cells. This was observed in: (1) primary keratinocytes in culture and (2) epidermal keratinocytes in the back skin of newborn mice. Whereas Paramio and colleagues seem to suggest that proliferation (as measured by BrdU incorporation) was affected by K1014chim expression in the ORS of hair follicles, this was in fact not the case. How to explain the obvious difference between prediction (reduced proliferation and hypoplastic epidermis) and observation (normal epidermal development) regarding the phenotype of K1014chim mice? Paramio and colleagues put forward several interesting hypotheses to resolve this apparent contradiction. Nevertheless, in our opinion, toxic expression levels of ectopically expressed K10 might explain the previously observed phenotypes. In this respect it is worth mentioning that Santos et al. reported that the hypoplastic phenotype of mice expressing K10 in the basal layer of the epidermis is dependent on transgene expression levels (Santos et al., 2002; Santos et al., 2003). As indicated by Paramio and colleagues, another factor affecting phenotypic differences between mouse models is the genetic background of the strain used. Although we cannot exclude this possibility, we believe that this is not likely to be the case, since we generated K1014chim mice on two different genetic backgrounds (C57/Bl, Balb/C), neither of which led to the development of a hypoplastic epidermis.
An interesting observation in the K1014chim line was that these animals are more susceptible to benign tumor (papilloma) formation than genetically matched wild-type controls. They developed papillomas in greater numbers and earlier than controls. The conversion rate to malignant tumors, however, was similar in mutants and controls. This observation is also inconsistent with data previously published by the Jorcano laboratory, since mice with forced K10 expression appeared to be more resistant to tumor development (Santos et al., 1997; Santos et al., 2002).
Our results suggest a reduced apoptotic rate in K1014chim keratinocytes as a possible mechanism to explain the increased tumor incidence. Several groups have reported in recent years that keratin IF can affect apoptosis by sequestering of apoptotic effector molecules in the cytoplasm (e.g. Caulin et al., 2000; Inada et al., 2001; Tong and Coulombe, 2006) [see references in Pallari and Eriksson (Pallari and Eriksson, 2006)]. However, the molecular mechanism that leads to a reduced apoptotic response in mutant keratinocytes is currently unknown.
We agree with Paramio and colleagues that this is an exciting time in keratin and IF biology, with the revelation that cellular processes like apoptotic response, proliferation, and metabolic activity (e.g. Kim et al., 2006) can be affected and, in fact, controlled by IF proteins simply by modulating the cytoplasmic and nuclear distribution of signaling molecules. Although we cannot resolve the discrepancies in the phenotypes of the mouse models described by us and the Jorcano group at this time, we believe that these examples demonstrate the need for further studies into the cell biological consequences of keratin expression in keratinocytes.
References
Caulin, C., Ware, C. F., Magin, T. M. and Oshima, R. G. (2000). Keratin-dependent, epithelial resistance to tumor necrosis factor-induced apoptosis. J. Cell Biol. 149, 17-22.
Chen, J., Cheng, X., Merched-Sauvage, M., Caulin, C., Roop, D. R. and Koch, P. J. (2006). An unexpected role for keratin 10 end domains in susceptibility to skin cancer. J. Cell Sci. 119, 5067-5076.
Inada, H., Izawa, I., Nishizawa, M., Fujita, E., Kiyono, T., Takahashi, T., Momoi, T. and Inagaki, M. (2001). Keratin attenuates tumor necrosis factor-induced cytotoxicity through association with TRADD. J. Cell Biol. 155, 415-426.
Kim, S., Wong, P. and Coulombe, P. A. (2006). A keratin cytoskeletal protein regulates protein synthesis and epithelial cell growth. Nature 441, 362-365.[CrossRef][Medline]
Koch, P. J. and Roop, D. R. (2004). The role of keratins in epidermal development and homeostasis - going beyond the obvious. J. Invest. Dermatol. 123, x-xi.[Medline]
Pallari, H. M. and Eriksson, J. E. (2006). Intermediate filaments as signaling platforms. Sci. STKE 2006, e53.
Paramio, J. M. and Jorcano, J. L. (2002). Beyond structure: do intermediate filaments modulate cell signalling? BioEssays 24, 836-844.[CrossRef][Medline]
Paramio, J. M., Casanova, M. L., Segrelles, C., Mittnacht, S., Lane, E. B. and Jorcano, J. L. (1999). Modulation of cell proliferation by cytokeratins K10 and K16. Mol. Cell. Biol. 19, 3086-3094.
Paramio, J. M., Santos, M. and Jorcano, J. L. (2007). The ends of a conundrum. J. Cell Sci. 120, 1145-1147.
Paramio, J. M., Segrelles, C., Ruiz, S. and Jorcano, J. L. (2001). Inhibition of protein kinase B (PKB) and PKCzeta mediates keratin K10-induced cell cycle arrest. Mol. Cell. Biol. 21, 7449-7459.
Santos, M., Ballestin, C., Garcia-Martin, R. and Jorcano, J. L. (1997). Delays in malignant tumor development in transgenic mice by forced epidermal keratin 10 expression in mouse skin carcinomas. Mol. Carcinog. 20, 3-9.[CrossRef][Medline]
Santos, M., Paramio, J. M., Bravo, A., Ramirez, A. and Jorcano, J. L. (2002). The expression of keratin k10 in the basal layer of the epidermis inhibits cell proliferation and prevents skin tumorigenesis. J. Biol. Chem. 277, 19122-19130.
Santos, M., Perez, P., Segrelles, C., Ruiz, S., Jorcano, J. L. and Paramio, J. M. (2003). Impaired NF-kappa B activation and increased production of tumor necrosis factor alpha in transgenic mice expressing keratin K10 in the basal layer of the epidermis. J. Biol. Chem. 278, 13422-13430.
Tong, X. and Coulombe, P. A. (2006). Keratin 17 modulates hair follicle cycling in a TNF{alpha}-dependent fashion. Genes Dev. 20, 1353-1364.
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