The overall objective of this renewal application is to extend our investigations into the role that Ha-ras gene expression plays during skin tumorigenesis in vivo, and to delineate the various factors involved in regulating the expression of this proto- oncogene in epidermal cells. Experiments are proposed to investigate regulation of Ha-ras gene expression at both the transcriptional and post-transcriptional level. We first propose to employ the heteroduplex mismatching method to characterize the Ha-ras mRNA transcripts previously observed by us to be present at elevated levels in developing papillomas, to determine whether Ha- ras transcription is originating from the normal or point-mutated allele, or both. We will also investigate transcriptional regulation in cis of the mouse c-Ha-ras gene in vitro, by identifying and characterizing cis- regulatory elements of the Ha- ras oncogene, using the transient gene expression assay for chloramphenicol acetyltransferase. In addition to examining factors which regulate expression in cis, we will investigate elements that may regulate expression of the Ha-ras gene in trans: A series of experiments is proposed which will manipulate trans- regulatory elements involved in expression of the mouse Ha-ras promoter in vitro. The glucocorticoids (which have long been known to inhibit skin tumor development) and tumor promoters (which encourage skin tumor development) will be used to characterize trans-acting regulatory factors in vitro. In addition transcriptional studies, we propose to carry out experimental cis- and trans-transcriptional regulation of the mouse c-Ha-ras gene in vivo. Mouse skin cells will be transfected with the normal and point-mutated mouse c-Ha-ras gene under the control of heterologous promoters, and the cells transplanted into SENCAR mice to test for development of papillomas. Another aim of this project is to investigate factors involved in post-transcriptional regulation of the mouse Ha-ras proto-oncogene. We have proposed experiments to compare the stability of Ha-ras transcripts in normal epidermis and in tumors, and to identify structural features of Ha-ras message which may determine its susceptibility to decay in the cytoplasm.

Agency
National Institute of Health (NIH)
Institute
National Cancer Institute (NCI)
Type
Research Project (R01)
Project #
2R01CA040847-04
Application #
3181159
Study Section
Chemical Pathology Study Section (CPA)
Project Start
1985-02-01
Project End
1993-01-31
Budget Start
1988-02-01
Budget End
1989-01-31
Support Year
4
Fiscal Year
1988
Total Cost
Indirect Cost
Name
University of Nebraska Medical Center
Department
Type
Overall Medical
DUNS #
City
Omaha
State
NE
Country
United States
Zip Code
68198
Liu, M; Pelling, J C; Ju, J et al. (1998) Antioxidant action via p53-mediated apoptosis. Cancer Res 58:1723-9
Dhanwada, K R; Dickens, M; Neades, R et al. (1995) Differential effects of UV-B and UV-C components of solar radiation on MAP kinase signal transduction pathways in epidermal keratinocytes. Oncogene 11:1947-53
Betz, N A; Pelling, J C (1995) Ha-ras p21-GTP levels remain constant during primary keratinocyte differentiation. Mol Carcinog 12:66-76
Liu, M; Pelling, J C (1995) UV-B/A irradiation of mouse keratinocytes results in p53-mediated WAF1/CIP1 expression. Oncogene 10:1955-60
Chakravarti, D; Pelling, J C; Cavalieri, E L et al. (1995) Relating aromatic hydrocarbon-induced DNA adducts and c-H-ras mutations in mouse skin papillomas: the role of apurinic sites. Proc Natl Acad Sci U S A 92:10422-6
Liu, M; Dhanwada, K R; Birt, D F et al. (1994) Increase in p53 protein half-life in mouse keratinocytes following UV-B irradiation. Carcinogenesis 15:1089-92
Betz, N A; Pelling, J C (1992) Resistance of adult keratinocytes to differentiation-induced decrease in Ha-ras mRNA levels observed in newborn keratinocytes. Mol Carcinog 6:10-7
Betz, N A; Wolterman, K J; Reiners, J J et al. (1992) DNA-mediated gene transfection into primary cultures of adult mouse keratinocytes. In Vitro Cell Dev Biol 28A:188-92
Neades, R; Betz, N A; Sheng, X Y et al. (1991) Transient expression of the cloned mouse c-Ha-ras 5' upstream region in transfected primary SENCAR mouse keratinocytes demonstrates its power as a promoter element. Mol Carcinog 4:369-75
Fujii, H; Egami, H; Chaney, W et al. (1990) Pancreatic ductal adenocarcinomas induced in Syrian hamsters by N-nitrosobis(2-oxopropyl)amine contain a c-Ki-ras oncogene with a point-mutated codon 12. Mol Carcinog 3:296-301

Showing the most recent 10 out of 17 publications