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Specific down-regulation of annexin II expression in human cells interferes with cell proliferation
Y Chiang1, A Rizzino, Z A Sibenaller
1Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha 68198-4525, USA.
Abstract:
The protein-tyrosine kinase substrate annexin II is a growth regulated gene whose expression is increased in several human cancers. While the precise function of this protein is not understood, annexin II is proposed to be involved in multiple physiological activities, including DNA synthesis and cell proliferation. Targeted disruption of the annexin II gene affects calcium signaling, tyrosine phosphorylation and apoptosis, indicating the important physiological role of this protein. We used a transient co-transfection assay to regulate annexin II expression in human HeLa, 293 and 293T cells, and measured the effects of annexin II down regulation on DNA synthesis and proliferation. Transfection of cells with an antisense annexin II vector results in inhibition of cell division and proliferation, with concomitant reduction in annexin II message and protein levels. Cellular DNA synthesis is significantly reduced in antisense transfected cells. Replication extracts made from antisense transfected cells have significantly reduced efficiency to support SV40 in vitro DNA replication, while the extracts made from sense transfected cells are fully capable of replication. Our results indicate an important role of annexin II in cellular DNA synthesis and cell proliferation.
Insights
Annexin II, a protein involved in cell growth, is crucial for DNA synthesis and cell proliferation. Downregulating annexin II expression inhibits cell division and DNA replication, highlighting its vital role.
Area of Science:
- Molecular Biology
- Cell Biology
- Cancer Research
Background:
- Annexin II is a growth-regulated gene with increased expression in human cancers.
- Its precise function is unclear, but it's implicated in DNA synthesis and cell proliferation.
- Gene disruption affects calcium signaling, tyrosine phosphorylation, and apoptosis, indicating a significant physiological role.
Purpose of the Study:
- To investigate the role of annexin II in DNA synthesis and cell proliferation.
- To determine the effects of annexin II downregulation on cellular processes.
Main Methods:
- Transient co-transfection assays were used to regulate annexin II expression in HeLa, 293, and 293T cells.
- Antisense annexin II vectors were employed to downregulate gene expression.
- Cellular DNA synthesis and proliferation rates were measured.
- In vitro DNA replication assays using replication extracts were performed.
Main Results:
- Antisense transfection significantly inhibited cell division and proliferation.
- Annexin II message and protein levels were reduced following antisense transfection.
- Cellular DNA synthesis was significantly decreased in transfected cells.
- Replication extracts from antisense-transfected cells showed reduced efficiency in supporting SV40 DNA replication.
Conclusions:
- Annexin II plays a critical role in cellular DNA synthesis.
- Annexin II is essential for cell proliferation.
- Downregulation of annexin II negatively impacts DNA replication and cell division.