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[Cancer Research 65, 6551-6556, August 1, 2005]
© 2005 American Association for Cancer Research

Molecular Biology, Pathobiology and Genetics

Mice Lacking the p53/p63 Target Gene Perp Are Resistant to Papilloma Development

Michelle R. Marques1, Jennifer S. Horner1, Rebecca A. Ihrie1, Roderick T. Bronson3 and Laura D. Attardi1,2

1 Department of Radiation Oncology, Division of Radiation and Cancer Biology and 2 Department of Genetics, Stanford University School of Medicine, Stanford, California; and 3 Department of Pathology, Harvard Medical School, Boston, Massachusetts

Requests for reprints: Laura D. Attardi. Department of Radiation Oncology, Stanford University School of Medicine, Stanford, CA 94305. Phone: 650-725-8424; Fax: 650-723-7382; E-mail: attardi{at}stanford.edu.


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
Perp is a target of the p53 tumor suppressor involved in the DNA damage-induced apoptosis pathway. In addition, Perp is a target of the p53-related transcription factor p63 during skin development, where it participates in cell-cell adhesion mediated through desmosomes. Here we test the role of Perp in tumorigenesis in a two-step skin carcinogenesis model system. We find that mice lacking Perp in the skin are resistant to papilloma development, displaying fewer and smaller papillomas than wild-type mice. Proliferation levels, apoptotic indices and differentiation patterns are similar in the skin of treated Perp-deficient and wild-type mice. Instead, impaired adhesion through aberrant desmosome assembly may explain the diminished tumor development in the absence of Perp. These studies indicate that in certain contexts, Perp is required for efficient carcinogenesis and suggest a role for intact cell-cell adhesion in supporting tumor development in these settings.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
The p53 tumor suppressor is the most commonly mutated gene in human cancer, with over half of all human cancers sustaining p53 mutations (1). Moreover, p53 null mice are highly tumor prone, succumbing universally to cancer within 10 months after birth (2). Together, these findings underscore the crucial role p53 plays in preventing cancer. p53 suppresses tumorigenesis by responding to various cellular stresses, such as DNA damage, hypoxia, or hyperproliferation, and inducing cells to undergo cell cycle arrest or apoptosis (3). Activation of either the cell cycle arrest or apoptotic pathway provides a mechanism to limit the propagation of potentially oncogenic cells. p53 is a transcriptional activator that engages these pathways at least in part through the induction of target genes involved in cell cycle arrest or apoptosis (3). Whereas a host of p53 target genes have been identified, the importance of most of these genes in tumor suppression in vivo has not been clearly elucidated.

p53 is a member of a multiprotein family of transcription factors, including the p63 and p73 proteins (4). Whereas p53's main function is in tumor suppression, p63 and p73 play more critical developmental roles, as deduced from the analysis of mice lacking these gene products (5–7). p73 is essential for neurogenesis and pheromone sensing, and p63 is required for the development of a variety of stratified epithelia, including the skin, hair follicles, and oral mucosa. Although these proteins have important developmental functions, their role in cancer has been controversial, as few cases of mutations in either p63 or p73 in cancers have been reported (8, 9). However, some studies indicate that there is interplay between p53 and its family members, suggesting that in some contexts, p73 or p63 might also participate in restricting tumorigenesis. For example, mice expressing p53 tumor mutants have a dominant predisposition to developing metastatic epithelial cancers that are thought to result from the binding and neutralization of p73 and p63 by mutant p53 (10, 11). Similarly, mice with compound mutations in p53 and p63, or p53 and p73, develop tumor types not observed in the single mutants, indicating cooperativity between these mutant alleles (12). These findings suggest that p63 and p73 provide back-up systems that limit tumorigenesis in the absence of p53. Target genes regulated by more than one p53 family member therefore may be very important for tumor suppression.

The Perp gene was identified in a screen for p53 target genes selectively induced during p53-mediated apoptosis rather than during cell cycle arrest (13). Perp overexpression is sufficient to induce cell death, and certain cell types lacking Perp display compromised apoptotic responses upon treatment with DNA damage (13, 14). Together, these findings indicate that Perp is an important component of the p53 apoptotic pathway. However, in addition to this apoptotic function downstream of p53, Perp plays a p53-independent role in development (15). Unlike p53 null mice, which are viable, Perp null mice die post-natally within 10 days after birth from blisters in stratified epithelia, including the skin and oral mucosa. This critical function for Perp in stratified epithelial integrity reflects a role as a downstream target of the p53 family member, p63 (15). Like p63, Perp is highly expressed in stratified epithelia, including the skin, and Perp expression depends on direct activation by p63.

In its capacity as a p63 target, Perp functions in specific cell-cell adhesion complexes known as desmosomes. Desmosomes are essential both for anchoring cells to each other and for conferring strength on a tissue by virtue of contacts to the intermediate filament cytoskeleton, a facet especially important in tissues subject to mechanical stress (16). Perp localizes to desmosomes, and cells in the epidermis of Perp-deficient mice display aberrant assembly of desmosomal complexes both by biochemical assays and electron microscopy, indicating the central role for Perp in desmosomal function (15). Thus, Perp is a key effector of the p63 stratified epithelial development program, mediating a subprogram specifically involved in cell-cell adhesion.

Given that Perp is positioned downstream of both p53 and p63 and participates in both apoptosis and adhesion, loss of Perp may be expected to promote cancer. To test potential tumor suppressor activity of Perp, we used a carcinogenesis model in the skin, a tissue where Perp is known to play an important role. Here, by examining conditional knockout mice in which Perp is ablated in the skin, we define the role for Perp in skin tumorigenesis. Surprisingly, we find that Perp-deficient mice are resistant to cancer, suggesting that intact Perp function is required for efficient tumorigenesis.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Skin carcinogenesis protocol. Male K5-Cre; Perpfl/+ were mated to Perpfl/+ females to generate mice used for this study, including 17 K5-Cre; Perp+/+ mice, 12 K5-Cre; Perpfl/fl mice, and 11 K5-Cre Perpfl/+ mice. Mice were of a mixed 129/Sv/C57BL/6 genetic background and littermates were used for experiments. Each mouse was genotyped by PCR. To detect the Cre transgene, we used the primers 5'-TGGGCGGCATGGTGCAAGTT-3' and 5'-CGGTGCTAACCAGCGTTTTC-3'. To determine the Perp genotype, we used primers located in the first intron, 5'-AGTCTTCAGGGATGACACAGA and 5'-TACGAAACTAGAGCACAGCTA-3', which result in a 326-bp product on the wild-type allele and a 410-bp product on the conditional allele.

The backs of 8-week-old mice were shaved and treated with a single application of 7,12-dimethylbenz(a)anthracene (DMBA, Sigma, St. Louis, MO; 10 µg in 100 µL acetone) followed by twice weekly application of 12-O-tetradecanoylphorbol 13-acetate (TPA, Sigma; 12.5 µg in 100 µL acetone) for 27 weeks. The numbers and sizes of papillomas were recorded once a week. With the exception of two K5-Cre; Perp+/+ mice and six K5-Cre; Perpfl/fl mice that died between weeks 24 and 28, mice were sacrificed 27 weeks after the initiation of TPA treatment (week 28), and skin with and without papillomas was fixed in 10% buffered formalin.

Histology and immunohistochemistry. Paraffin sections from adult mouse skin were prepared for immunohistochemical or H&E staining by standard methods. Immunohistochemistry was done as described (15).

Terminal deoxynucleotidyl transferase–mediated nick-end labeling and Ki67 assays. Terminal deoxynucleotidyl transferase–mediated nick-end labeling (TUNEL) and Ki67 assays were done as described (14, 15). Proliferation was quantified by counting the number of labeled cells as a percentage of total basal cells in each 20x field. Assays were done on treated skin from mice at the end of the study.

Protein preparation and immunoblotting. For skin protein preparation, skin was snap-frozen in liquid nitrogen and homogenized using a chilled mortar and pestle. Triton-soluble and urea-soluble protein fractions were prepared as described (15). Western blotting was done according to standard methods, with 25 µg of protein per lane.

Antibodies. For immunofluorescence and immunohistochemistry, antibodies against Ki67 (PharMingen, San Diego, CA) as well as keratin 14, keratin 1, and loricrin (Covance, Berkeley, CA) were used. For immunoblotting, antibodies against desmoglein 1 (4B2, gift of K. Green, Northwestern), plakoglobin (1407, gift of K. Green), and tubulin (Sigma) were used. Polyclonal antibodies against Perp have been described (15).


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Specific inactivation of Perp in skin. As constitutive Perp-deficient mice display post-natal lethality, we took advantage of Perp conditional knockout mice that we generated to establish the role of Perp in skin carcinogenesis (Fig. 1A). To conditionally ablate Perp in the skin, we bred floxed Perp mice (denoted Perpfl/fl) to Keratin 5-Cre transgenic mice expressing Cre specifically in stratified epithelia, including the skin (17). Approximately 40% of the K5-Cre, Perpfl/fl mice survived, and were therefore used in skin carcinogenesis studies. Although variable Cre expression in some K5-Cre-expressing tissues could explain this partially penetrant lethality, our detailed analysis shows that deletion is 100% efficient in the skin, both by Western blot and by immunohistochemistry. (Fig. 1B-D). These findings indicate that the surviving K5-Cre; Perpfl/fl adults provide a useful model in which to examine the effect of Perp deficiency on skin cancer development.



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  		Figure 1. K5-Cre transgenic system to inactivate Perp in the skin. A, targeting scheme used to generate conditional (floxed) Perp mice. The generation of embryonic stem cells with the targeted 3-lox Perp allele has been described (14). Mice carrying the 3-lox Perp allele were bred to CMV-Cre transgenic mice (34) to obtain Perp conditional mice. B, Western blot analysis to examine Perp levels in skin from K5-Cre transgenic; Perpfl/fl and control nontransgenic; Perpfl/fl mice. fl, floxed allele. Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) serves as a loading control. C-D, immunohistochemistry to examine Perp protein levels and localization in K5-Cre Perp+/+ and K5-Cre; Perpfl/fl mouse skin. For clarity, epidermis from DMBA/TPA-treated mice was examined and hence appears thickened relative to untreated adult skin.

 
Perp-deficient mice are resistant to skin papillomagenesis. To determine the role of Perp in skin papilloma development, we subjected cohorts of K5-Cre; Perpfl/fl and K5-Cre; Perp+/+ control adult mice to a well-established two-step skin carcinogenesis protocol. In this protocol, tumor initiation results from application of the carcinogen DMBA, which induces activating ras mutations, and tumor promotion ensues after multiple weeks of treatment with the phorbol ester TPA (18, 19). Mice were exposed to one treatment with DMBA followed by 27 weeks of biweekly TPA treatment, and both numbers and sizes of developing papillomas were monitored during this period. Papillomas began to appear in both groups at about the same time, ~8 to 9 weeks after treatment initiation. In K5-Cre; Perp+/+ mice, the numerous papillomas that appeared grew progressively larger throughout the treatment period (Fig. 2A and C). In striking contrast, K5-Cre; Perpfl/fl mice were highly resistant to papillomagenesis, typically displaying only a minuscule number of tumors, that were diminished in size (Fig. 2B-C). At the time of death, the average number of papillomas in the Perp-deficient mice was ~4-fold less than the wild-type mice (Fig. 2D). The difference in papilloma numbers, as well as size, between the wild-type and the Perp-deficient mice at the end of the study suggests that Perp is important for efficient papilloma development (Fig. 2A-D). Interestingly, mice heterozygous for the Perp conditional allele developed the same number and size of papillomas as wild-type mice, indicating that one copy of Perp is sufficient to facilitate papilloma development (Fig. 2D). However, except for the size differences, the gross histologic appearance of the papillomas was not obviously different between the K5-Cre; Perp+/+ and K5-Cre; Perpfl/fl mice (Fig. 2E and F). In addition, none of the papillomas in these cohorts of mice progressed to malignant carcinomas. As our mice are on a mixed 129/Sv;C57BL/6 background, this may relate to the previously described resistance of C57BL/6 mice to malignant conversion in this model system (20).



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  		Figure 2. Perpfl/fl mice are resistant to papilloma development. A, columns, average numbers and sizes of papillomas >2 mm in K5-Cre; Perp+/+ mice as a function of time of TPA treatment. Color scale indicates the sizes of papillomas. B, columns, average numbers and sizes of papillomas >2 mm in K5-Cre; Perpfl/fl mice as a function of time of TPA treatment. Color scale indicates the sizes of papillomas. C, representative images of K5-Cre; Perp+/+ and K5-Cre; Perpfl/fl mice showing the dramatic difference in papilloma numbers and sizes seen in the absence of Perp at week 28. D, columns, average number of papillomas >2 mm, at the time of death (between weeks 24 and 28), in K5-Cre transgenic mice that are either Perp+/+, Perpfl/+, or Perpfl/fl; bars, ±SE. E, H&E staining of a portion of a typical papilloma in a K5-Cre; Perp+/+ mouse. F, H&E staining of typical papilloma in a K5-Cre; Perpfl/fl mouse.

 
Perp-deficient skin displays normal proliferation, apoptosis, and differentiation profiles. The dramatic inhibition in papilloma development observed in the K5-Cre; Perpfl/fl mice relative to K5-Cre; Perp+/+ mice could be accounted for by decreased proliferation or enhanced apoptosis in the absence of Perp. To determine if there were inherent differences in the properties of keratinocytes in the wild-type and Perp-deficient mice that could result in differences in the propensity to papillomagenesis, we examined levels of cell division and apoptosis in treated skin from mice of both genotypes.

To establish proliferation indices, we measured the percentages of Ki67-positive basal cells in the skin of treated Perp-deficient and wild-type mice (Fig. 3A-D). We found that the fraction of Ki67-positive cells in the Perp-deficient skin was indistinguishable from that observed in wild-type skin, indicating that a difference in the proliferative capacity of the cells of the treated skin does not account for the relative resistance of Perp-deficient mice to skin carcinogenesis (Fig. 3E). In addition, no clear difference in proliferation levels was observed in the developing papillomas of K5-Cre; Perpfl/fl mice relative to K5-Cre; Perp+/+ mice (data not shown).



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  		Figure 3. Proliferation and apoptosis levels are comparable in the epidermis of treated K5-Cre; Perp+/+ and K5-Cre; Perpfl/fl mice. A-B, proliferation index was determined by Ki67 staining of skin from K5-Cre; Perp+/+ and K5-Cre; Perpfl/fl mice after 27 weeks of TPA treatment. C-D, 4',6-diamidino-2-phenylindole (DAPI) labels all nuclei of the epidermis. E, columns, average percentage of basal cells of the epidermis displaying Ki67 positivity, in treated K5-Cre; Perp+/+ and K5-Cre; Perpfl/fl mice; bars, ±SE. F-G, apoptosis was assessed through TUNEL staining of skin from K5-Cre; Perp+/+ and K5-Cre; Perpfl/fl mice after 27 weeks of TPA treatment. Arrows, TUNEL-positive cells.

 
Given Perp's established role in apoptosis (14), we sought to determine if apoptosis levels were altered in the absence of Perp. TUNEL staining was done on K5-Cre; Perp+/+ and K5-Cre; Perpfl/fl mouse skin (Fig. 3F and G). TUNEL staining failed to reveal significant numbers of apoptotic cells in treated skin derived from mice of either genotype. Moreover, apoptotic levels were also minimal in papillomas derived from both cohorts of mice (data not shown). These findings indicate that augmented apoptosis does not explain the inability of papillomas to form efficiently in the absence of Perp.

One additional possibility is that papillomagenesis is impeded in the absence of Perp because of aberrant differentiation of keratinocytes in the skin, as has been observed with certain mouse models examined in the two-step protocol (21). To address this issue, the differentiation program in treated skin from K5-Cre; Perp+/+ and K5-Cre; Perpfl/fl mice was examined (Fig. 4). Staining for K14, K1, and loricrin, markers of the basal, spinous, and granular layers of the epidermis, respectively (22, 23), was done. All of these markers were detected irrespective of Perp genotype, indicating that the differentiation process occurs normally and suggesting that the disruption of differentiation is not the basis for the inhibited papillomagenesis in K5-Cre; Perpfl/fl mice (Fig. 4). Taken together, these findings suggest that neither the cell number nor the differentiation state in the treated skin vary with Perp status but rather some other property of the skin may differ. Given the previously described role for Perp as a crucial desmosomal protein, we hypothesized that adhesion differences might explain the differences in papilloma development observed in the two cohorts.



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  		Figure 4. The epidermal differentiation program occurs normally in the absence of Perp. Immunohistochemical analysis was used to examine the terminal differentiation program in the skin of K5-Cre; Perp+/+ and K5-Cre; Perpfl/fl mice treated for 27 weeks with TPA. A-B, keratin 14, typically a marker of the basal cell population in the skin, shows widespread expression in treated skin of mice of both genotypes. C-D, keratin 1 is a marker of the spinous layer of the skin. E-F, loricrin is a marker of the granular layer of the epidermis.

 
Significant adhesion defects are observed in Perp-deficient skin. Perp–/– mice are prone to the development of blisters, or separation between epithelial layers, as well as separation between individual epithelial cells in the skin due to disrupted desmosome function (15). To test the hypothesis that compromised desmosomal function may form the basis for the decreased susceptibility to papilloma development in K5-Cre; Perpfl/fl mice, we examined adhesion in the skin of these mice. Skin from untreated newborn K5-Cre; Perpfl/fl mice showed a propensity to develop blisters, similar to that observed in Perp constitutive null animals, indicating that clear adhesion defects are observed when Perp is deleted selectively in the skin (Fig. 5A-B). To specifically determine if adhesion is disrupted in the adult mice subjected to the carcinogen protocol, we examined the treated skin by histologic analysis. Interestingly, whereas the keratinocytes in the K5-Cre; Perp+/+ mice showed normal attachment to each other, cells in the K5-Cre; Perpfl/fl mouse skin displayed clear signs of separation (Fig. 5C-F, arrows). These results suggest that adhesion is impaired in the skin of Perp-deficient mice.



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  		Figure 5. Desmosome function is perturbed in the skin of K5-Cre; Perpfl/fl mice. A, H&E staining of newborn K5-Cre; Perp+/+ skin. B, H&E staining of newborn K5-Cre; Perpfl/fl skin. A blister formed by separation of the basal and suprabasal layers of the epidermis (arrow). C, H&E staining of K5-Cre; Perp+/+ mouse skin treated with TPA for 27 weeks. D, H&E staining of K5-Cre; Perpfl/fl mouse skin treated with TPA for 27 weeks. Note spaces between epithelial cells (arrows). E, higher magnification view of treated K5-Cre; Perp+/+ mouse skin. F, higher magnification view of treated K5-Cre; Perpfl/fl mouse skin showing separation between epidermal cells. G, Western blot analysis examining Triton X-100 solubility profiles of desmosomal proteins in K5-Cre; Perpfl/fl and control (K5-Cre; Perp+/+ and Perpfl/fl) mouse skin. Tubulin and K14 serve as loading controls for the Triton X-100–soluble and –insoluble fractions, respectively. H, Western blot analysis showing that total protein levels of desmoglein 1 and plakoglobin are similar regardless of genotype.

 
To show specifically that desmosome function is responsible for the observed alterations in adhesion, we used an assay that we established previously as a measure of desmosome integrity in studies of Perp–/– mice (15). When desmosomes undergo proper assembly, desmosomal components are largely insoluble in the detergent Triton X-100. If the assembly or stability of the desmosome is abnormal, however, then desmosomal proteins can be solubilized by Triton X-100 to a greater extent than usual. In skin from newborn Perp–/– mice, the desmosomal proteins desmoglein 1 and plakoglobin display increased solubility relative to skin from wild-type mice. This biochemical property is accompanied by clear defects in desmosome structure at the electron microscopy level (15, 24). We analyzed the solubility of these central desmosomal proteins in untreated skin of K5-Cre; Perp+/+ mice and K5-Cre; Perpfl/fl adult mice. Both desmosomal constituents examined, desmoglein 1 and plakoglobin, were more Triton X-100 soluble in the K5-Cre; Perpfl/fl mouse skin than in control skin, indicating that in the absence of Perp in the skin, normal assembly of mature desmosomes is compromised (Fig. 5G), consistent with our previous observations in the skin of newborn constitutive Perp–/– mice. Total levels of these desmosome proteins do not vary, indicating that the observed effects relate specifically to solubility differences (Fig. 5H). Together, our results suggest that in the absence of Perp, skin tumorigenesis is impeded and that proper desmosome function is important for tumorigenesis.


   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
Here, we test the role of the p53/p63 target gene Perp in cancer development using a two-step skin carcinogenesis model. Based on the central roles for Perp in both apoptosis and adhesion, we hypothesized that Perp-deficient mice would display an enhanced predisposition to cancer. Surprisingly, however, we observed a striking resistance to tumorigenesis in mice lacking Perp in the skin, with diminished numbers and sizes of skin papillomas. In a variety of mouse models, similar resistance to papilloma development is clearly correlated with inherent changes in proliferation, apoptosis, or differentiation (21, 25, 26). In contrast, in the treated skin of mice lacking Perp, we observed no such alterations. Instead, we observed defects in adhesion, specifically in desmosome assembly, in the skin of Perp-deficient mice. Our findings indicate that Perp is important for skin tumorigenesis in this system and suggest a requirement for intact desmosome function during this process.

Given Perp's central function in adhesion in the skin, it is likely that its ability to facilitate skin papilloma development is related to this activity. It is well-established that the development of cancer requires the dysregulation of adhesion (27). In particular, changes in adhesion have been shown to play an important part in later stages of cancer, where disrupted cell-cell contacts contribute to the processes of invasion and metastasis (28, 29). Our findings, in contrast, suggest that proper cell-cell adhesion may be required, at least in some contexts, for the establishment and growth of tumors. The resistance of Perp-deficient cells to tumor development may reflect an intrinsic alteration in the behavior of these cells resulting from adhesion defects, such as enhanced motility or altered cell-cell signaling, which may somehow impede tumor development. Alternatively, it may be that efficient tumor morphogenesis and formation of the three-dimensional papilloma structure relies on appropriate cell-cell adhesive contacts, and when these are perturbed, tumor formation is inhibited. The importance of external cues, specifically cell-extracellular matrix (ECM) contacts between epithelial cells and the basement membrane, in tumorigenesis has been suggested previously through a similar skin carcinogenesis study in mice lacking focal adhesion kinase (30, 31). Keratinocytes in Fak+/– mice have impaired integrin-mediated signaling, which results in an inhibition of papilloma development similar to that observed here. Similarly, expression of {alpha}6ß4-integrin and its ligand laminin 5 have been shown to be critical for squamous cell tumorigenesis (32). Together with our study, these results implicate both proper cell-ECM and cell-cell adhesion in supporting tumorigenesis.

Our findings do not exclude the possibility that adhesion through desmosomes may inhibit tumorigenesis in certain instances. In fact, numerous studies have indicated that both desmosomal and adherens junction components are down-regulated during tumorigenesis and that compromised adhesion may promote tumor metastasis (28, 29). For example, inactivation of the transmembrane component of the adherens junction, E-cadherin, through mutation, promoter methylation, or transcriptional repression, is associated with the progression from adenoma to carcinoma and with the acquisition of metastatic potential (28, 33). An interesting possibility is that the specific function of adhesion complexes may depend on the particular stage of cancer examined. For example, whereas the presence of Perp allows early skin carcinogenesis through promoting papilloma development, it may play an anticancer role in late-stage cancer by facilitating adhesion, thereby blocking cell detachment and metastasis. This idea is consistent with reports that Perp is down-regulated in metastatic compared with primary melanomas (34, 35). Future experiments will determine if Perp loss promotes tumorigenesis at later stages of carcinogenesis.

The limited papilloma development in Perp-deficient mice is reminiscent of that seen in p53–/– mice, which display a resistance to developing papillomas compared with wild-type or p53 heterozygous mice (18). The reason for this paradoxical inhibition of tumorigenesis has never been understood. Because Perp is a target of p53, it is tempting to speculate that the cause of the inhibited papillomagenesis observed in the p53-deficient mouse skin could relate to that seen in Perp-deficient mouse skin. However, Perp expression in stratified epithelia such as skin does not depend on p53 and instead relies on p63 (15). Thus, in the absence of p53, Perp expression remains intact, suggesting that the resistance to tumorigenesis in the p53–/– mice has a different basis. It has not yet been possible to examine p63-deficient mice in this two-step protocol because of their neonatal lethal phenotype. Our data suggest the possibility that p63 loss might also inhibit papilloma development in this model, an idea that potentially can be tested in future studies with conditional p63 knockout mice.

The observation that compromised desmosome function impairs tumorigenesis has therapeutic implications. Our findings suggest that treatment of cancers of specific types or at particular stages with anti-desmosome molecules may inhibit tumor development. Perp, as a tetraspan membrane protein exposed at the plasma membrane, represents an accessible target. It will be of great interest to determine in the future if inhibiting the function of molecules such as Perp in developing tumors impedes the tumorigenic process.


   Acknowledgments
 
Grant support: National Cancer Institute grant CA93665-01 and Damon Runyon Cancer Research Foundation (L.D. Attardi).

The costs of publication of this article were defrayed in part by the payment of page charges. This article must therefore be hereby marked advertisement in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.

We thank Anthony Oro, Julien Sage, Steven Artandi, and Paul Khavari for critical reading of the article and Kathleen Green for providing antibodies.

Received 2/ 2/05. Revised 5/ 5/05. Accepted 5/13/05.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

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