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Induction of cyclin D1 by simian virus 40 small tumor antigen
1Department of Medicine, Lurie Cancer Center, Northwestern University, Chicago, IL 60611, USA.
Abstract:
Cell-cycle progression is mediated by a co-ordinated interaction between cyclin-dependent kinases and their target proteins including the pRB and E2F/DP-1 complexes. Immunoneutralization and antisense experiments have established that the abundance of cyclin D1, a regulatory subunit of the cyclin-dependent kinases, may be rate-limiting for G1 phase progression of the cell cycle. Simian virus 40 (SV40) small tumor (t) antigen is capable of promoting G1 phase progression and augments substantially the efficiency of SV40 transformation through several distinct domains. In these studies, small t antigen stimulated cyclin D1 promoter activity 7-fold, primarily through an AP-1 binding site at -954 with additional contributions from a CRE site at -57. The cyclin D1 AP-1 and CRE sites were sufficient for activation by small t antigen when linked to an heterologous promoter. Point mutations of small t antigen between residues 97-103 that reduced PP2A binding were partially defective in the induction of the cyclin D1 promoter. These mutations also reduced activation of MEK1 and two distinct members of the mitogen-activated protein kinase family, the ERKs (extracellular signal regulated kinases) and the SAPKs (stress-activated protein kinases), in transfected cells. Dominant negative mutants of either MEK1, ERK or SEK1, reduced small t-dependent induction of the cyclin D1 promoter. SV40 small t induction of the cyclin D1 promoter involves both the ERK and SAPK pathways that together may contribute to the proliferative and transformation enhancing activity of small t antigen.
Insights
Simian virus 40 (SV40) small t antigen promotes cell cycle G1 progression by activating the cyclin D1 promoter. This involves the AP-1 and CRE sites, and impacts ERK and SAPK pathways, enhancing cell proliferation and transformation.
Area of Science:
- Molecular Biology
- Cell Biology
- Virology
Background:
- Cell-cycle progression relies on cyclin-dependent kinases and their targets like pRB and E2F/DP-1.
- Cyclin D1 abundance is critical for G1 phase progression.
- Simian virus 40 (SV40) small t antigen promotes G1 progression and SV40 transformation.
Purpose of the Study:
- To investigate the molecular mechanisms by which SV40 small t antigen stimulates cyclin D1 promoter activity.
- To identify the specific regulatory elements and signaling pathways involved in small t antigen-mediated cyclin D1 induction.
- To understand the contribution of these pathways to the proliferative and transformation-enhancing effects of SV40 small t antigen.
Main Methods:
- Reporter assays to measure cyclin D1 promoter activity.
- Site-directed mutagenesis to probe the function of specific SV40 small t antigen domains and promoter elements (AP-1, CRE).
- Expression of dominant-negative mutants of MEK1, ERK, and SEK1 to assess pathway involvement.
Main Results:
- SV40 small t antigen stimulated cyclin D1 promoter activity 7-fold, primarily via an AP-1 site (-954) and a CRE site (-57).
- Mutations in small t antigen affecting PP2A binding impaired cyclin D1 promoter induction and activation of MEK1, ERK, and SAPK pathways.
- Dominant-negative mutants of MEK1, ERK, or SEK1 reduced small t-dependent cyclin D1 promoter induction.
Conclusions:
- SV40 small t antigen induces cyclin D1 promoter activity through specific cis-regulatory elements (AP-1, CRE).
- The ERK and SAPK signaling pathways are crucial for small t antigen-mediated induction of cyclin D1.
- These pathways likely contribute to the ability of SV40 small t antigen to enhance cell proliferation and transformation.