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Updated: Jul 13, 2026
In vitro Organoid Culture of Primary Mouse Colon Tumors
Published on: May 17, 2013
Sp1 regulates osteopontin expression in SW480 human colon adenocarcinoma cells
Yoji Takami1, Michael B Russell, Chengjiang Gao
1Chiba University, Chiba City, Japan.
Background:
Osteopontin (OPN) mediates cancer metastasis. Mechanisms regulating OPN expression in human colorectal cancer are unknown. Using SW480 colon adenocarcinoma cells, we hypothesized that transcription determines OPN expression.
Methods:
SW480 constitutively express OPN. Transient transfection and deletion analysis of human OPN promoter (full-length 2.1 kb)-luciferase constructs identified cis-regulatory regions. Gelshift and chromatin immunoprecipitation (ChIP) assays identified the trans-regulatory nuclear protein. Using in vitro adhesion, migration, and invasion studies, siRNA was used to determine the functional effect of decreased nuclear protein expression.
Results:
A cis-regulatory promoter region, nt-80 to nt-108, upregulated OPN transcription. Gelshift assays demonstrated specific binding of nuclear proteins. Competition with unlabeled mutant oligonucleotides indicated that the region, nt-94 to nt-104 (TGGGCTGGGC), was essential for protein binding in gelshift assays. Confirmatory ChIP assays showed the corresponding nuclear protein to be Sp1. Sp1 expression was ablated with siRNA (si-Sp1), resulting in decreased OPN-dependent adhesion, migration, and invasion by 50%, 70%, and 65%, respectively. Exogenous addition of OPN to si-Sp1 cells restored adhesion, migration, and invasion indices.
Conclusions:
In SW480 human colon cancer cells, we conclude that Sp1 mediated expression of the tumor metastasis protein, OPN, regulates in vitro functional correlates of tumor metastasis.
