Structural and functional analysis of the differential effects of c-Jun and v-Jun on prolactin gene expression
Kathryn N Farrow1, Andrew P Bradford, John J Tentler
1Department of Medicine, University of Colorado Health Sciences Center, Aurora 80045, USA.
Abstract:
The protooncogene c-Jun and its oncogenic isoform v-Jun are members of the activator protein 1 family of transcription factors that have been shown to have differential transcriptional effects that are both promoter specific and cell type specific. Previously, we have demonstrated that whereas c-Jun inhibits pituitary-specific rat prolactin (rPRL) promoter activity, expression of v-Jun stimulates the rPRL promoter in GH4 pituitary cells. In this report, we have conducted an extensive structure-function analysis of c-Jun vs. v-Jun to determine which regions of these proteins are responsible for their differential transcriptional effects in this pituitary model system. We show that isoform-specific responses are mediated by complex interactions between the delta-domain, serine 243, and the amino-terminal transcriptional activation domains. Thus, in contrast to previous reports, no single domain is responsible for the differential transcriptional activities of c-Jun and v-Jun. Mutation of c-Jun serine 243 to phenylalanine and replacement of the c-Jun amino terminus with the corresponding region from v-Jun, thereby removing the delta-domain, are necessary and sufficient to confer a functional switch from the c-Jun-inhibitory to the v-Jun-activating phenotype. Thus, we propose that isoform-specific subdomains in c-Jun and v-Jun dictate discrete interactions with distinct protein partners, which underlie the differential Jun-dependent transcriptional responses of the rPRL promoter.
Insights
The protooncogene c-Jun and its oncogenic form v-Jun have different effects on the rat prolactin promoter. Specific protein regions and interactions, not a single domain, control these distinct transcriptional activities.
Area of Science:
- Molecular Biology
- Oncogenes
- Gene Regulation
Background:
- Proto-oncogene c-Jun and oncogenic isoform v-Jun are Activator Protein 1 transcription factors.
- These Jun isoforms exhibit promoter and cell-type specific transcriptional effects.
- Previously, c-Jun inhibited, while v-Jun stimulated, rat prolactin (rPRL) promoter activity in GH4 pituitary cells.
Purpose of the Study:
- To perform a structure-function analysis of c-Jun and v-Jun.
- To identify regions responsible for their differential transcriptional effects on the rPRL promoter in pituitary cells.
Main Methods:
- Extensive structure-function analysis of c-Jun and v-Jun proteins.
- Mutational analysis, including specific amino acid substitutions and domain replacements.
- Assays to evaluate transcriptional activity on the rat prolactin promoter in GH4 pituitary cells.
Main Results:
- Differential transcriptional responses are mediated by interactions involving the delta-domain, serine 243, and N-terminal activation domains.
- No single domain accounts for the distinct activities of c-Jun and v-Jun.
- Mutating serine 243 to phenylalanine and replacing the N-terminus of c-Jun with that of v-Jun switched function from inhibition to activation.
Conclusions:
- Isoform-specific subdomains within c-Jun and v-Jun dictate interactions with distinct protein partners.
- These interactions underlie the differential Jun-dependent transcriptional regulation of the rPRL promoter.
- The findings challenge previous notions of single-domain responsibility for differential Jun activity.
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