We are interested in the early regulatory events underlying the switch between epithelial cell growth and differentiation, under normal conditions and during neoplastic development. In particular, we recently found that a specific tyrosine kinase, fyn, is activated at early times of mouse keratinocyte differentiation and is required for specific aspects of this process to occur. We will expand on these initial findings, by testing the following hypothesis: 1) fyn may play an essential function in early cell/substrate and cell/cell adhesion events, and/or ensuing changes associated with the onset of keratinocyte differentiation. We will test this hypothesis by comparing wild type versus fyn deficient keratinocytes, under well defined culture conditions, as well as in vivo, under basal conditions and by wound healing experiments. 2) Other src-family members may contribute to keratinocyte growth/differentiation control. We will test the hypothesis that fyn function in keratinocytes can be compensated, in part, by other src-related kinases expressed in these cells. For this, we will compare keratinocytes with single versus double knockout mutations, i.e. derived from (fyn-/-, yes-/-), (fyn-/-, src-/-) and (yes-/-, src-/-) mice, under both in vitro and in vivo conditions. 3) Increased fyn kinase activity may exert on its own specific effects on keratinocyte growth/differentiation control, in the absence of the other changes associated with induction of the differentiation process. We will test this hypothesis by expressing either a wild type or constitutively active Fyn kinase in primary keratinocytes, by transient transfection of plasmid vector DNAs or infection with recombinant adenoviruses. Growth/differentiation parameters of these cells will be evaluated and compared with those of cells expressing the related Src and Yes kinases. 4) One or more domains of the Fyn protein may be more specifically connected with its function in keratinocytes. For this purpose, we will express variously mutated Fyn proteins in primary keratinocytes, by either transient transfection or recombinant adenovirus infection. These mutants will be compared with the intact Fyn protein for their ability (a) to rescue the abnormal phenotype of fyn deficient keratinocytes; (b) to modulate on their own specific aspects of keratinocyte growth and differentiation. Significant mechanistic insights will be gained by the identification of the specific region(s) of Fyn more closely connected with its function in keratinocytes, and by the knowledge of the protein(s) with which these region(s) interact.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
5R01CA073796-03
Application #
6124544
Study Section
General Medicine A Subcommittee 2 (GMA)
Program Officer
Spalholz, Barbara A
Project Start
1997-12-08
Project End
2001-11-30
Budget Start
1999-12-01
Budget End
2000-11-30
Support Year
3
Fiscal Year
2000
Total Cost
$336,707
Indirect Cost
Name
Massachusetts General Hospital
Department
Type
DUNS #
City
Boston
State
MA
Country
United States
Zip Code
02199
Zhu, Zehua; Todorova, Kristina; Lee, Kevin K et al. (2014) Small GTPase RhoE/Rnd3 is a critical regulator of Notch1 signaling. Cancer Res 74:2082-93
Mandinova, Anna; Kolev, Vihren; Neel, Victor et al. (2009) A positive FGFR3/FOXN1 feedback loop underlies benign skin keratosis versus squamous cell carcinoma formation in humans. J Clin Invest 119:3127-37
Mandinova, Anna; Lefort, Karine; Tommasi di Vignano, Alice et al. (2008) The FoxO3a gene is a key negative target of canonical Notch signalling in the keratinocyte UVB response. EMBO J 27:1243-54
Lefort, Karine; Mandinova, Anna; Ostano, Paola et al. (2007) Notch1 is a p53 target gene involved in human keratinocyte tumor suppression through negative regulation of ROCK1/2 and MRCKalpha kinases. Genes Dev 21:562-77
Nguyen, Bach-Cuc; Lefort, Karine; Mandinova, Anna et al. (2006) Cross-regulation between Notch and p63 in keratinocyte commitment to differentiation. Genes Dev 20:1028-42
Devgan, Vikram; Nguyen, Bach-Cuc; Oh, Heysun et al. (2006) p21WAF1/Cip1 suppresses keratinocyte differentiation independently of the cell cycle through transcriptional up-regulation of the IGF-I gene. J Biol Chem 281:30463-70
Grossi, Maddalena; Hiou-Feige, Agnes; Tommasi Di Vignano, Alice et al. (2005) Negative control of keratinocyte differentiation by Rho/CRIK signaling coupled with up-regulation of KyoT1/2 (FHL1) expression. Proc Natl Acad Sci U S A 102:11313-8
Mammucari, Cristina; Tommasi di Vignano, Alice; Sharov, Andrey A et al. (2005) Integration of Notch 1 and calcineurin/NFAT signaling pathways in keratinocyte growth and differentiation control. Dev Cell 8:665-76
Devgan, Vikram; Mammucari, Cristina; Millar, Sarah E et al. (2005) p21WAF1/Cip1 is a negative transcriptional regulator of Wnt4 expression downstream of Notch1 activation. Genes Dev 19:1485-95
Wang, Jian; Devgan, Vikram; Corrado, Marcella et al. (2005) Glucocorticoid-induced tumor necrosis factor receptor is a p21Cip1/WAF1 transcriptional target conferring resistance of keratinocytes to UV light-induced apoptosis. J Biol Chem 280:37725-31

Showing the most recent 10 out of 25 publications