Related Experiment Videos
Regulation of growth factor induced gene expression by calcium signalling: integrated mRNA and protein expression
R E Jenkins1, S R Hawley, W Promwikorn
1Department of Human Anatomy & Cell Biology, New Medical School, University of Liverpool, UK.
Abstract:
There is considerable indirect evidence that growth factor induced changes in the intracellular concentration of calcium play an important role in the regulation of the mammalian cell cycle. However, the precise mechanism by which this may be achieved remains unclear. Here we show that SKF-96365, an inhibitor of growth factor induced capacitative calcium entry (CCE), inhibits cell cycle progression by preventing entry into S phase. SKF-96365 changes the temporal profile of growth factor induced calcium signalling and recent studies have shown that alterations in the temporal and spatial patterns of calcium signalling can differentially regulate gene expression. We have therefore sought to examine the effect of inhibition of CCE on growth factor induced gene expression during G1. To achieve this we have initiated a combined transcriptomic and proteomic approach to measure CCE regulated gene expression using cDNA arrays and two-dimensional polyacrylamide gel electrophoresis, respectively. The initial results of this on-going analysis are reported here. They reveal that inhibition of CCE influences the expression of 29 genes at the mRNA level and 22 genes at the protein level. We report the identification of the mRNAs whose expression is altered by inhibition of CCE and describe the potential functional significance of some of these changes. The value of integrating a transcriptomic and two-dimensional gel electrophoresis based proteomic approach to studies of gene expression is discussed.
Insights
SKF-96365, an inhibitor of capacitative calcium entry (CCE), halts mammalian cell cycle progression by blocking entry into S phase. This study reveals CCE inhibition alters gene expression at both mRNA and protein levels during G1.
Area of Science:
- Cell Biology
- Molecular Biology
- Biochemistry
Background:
- Growth factor-induced calcium signaling is crucial for mammalian cell cycle regulation.
- The precise mechanisms by which calcium influences the cell cycle remain largely undefined.
- Capacitative calcium entry (CCE) is a key pathway for calcium signaling.
Purpose of the Study:
- To investigate the role of CCE in regulating cell cycle progression.
- To determine the impact of inhibiting CCE on growth factor-induced gene expression during the G1 phase.
- To identify specific genes affected by CCE inhibition at the transcriptomic and proteomic levels.
Main Methods:
- Utilized SKF-96365, a CCE inhibitor, to block calcium entry.
- Employed a combined transcriptomic (cDNA arrays) and proteomic (2D-PAGE) approach.
- Analyzed changes in gene expression at both mRNA and protein levels during the G1 phase.
Main Results:
- SKF-96365 treatment inhibited cell cycle progression by preventing entry into S phase.
- Inhibition of CCE altered the temporal dynamics of growth factor-induced calcium signaling.
- CCE inhibition affected the expression of 29 genes at the mRNA level and 22 genes at the protein level.
- Identified specific mRNAs with altered expression and discussed their potential functional significance.
Conclusions:
- Capacitative calcium entry plays a significant role in regulating cell cycle progression, specifically entry into S phase.
- Altering calcium signaling dynamics through CCE inhibition impacts global gene expression.
- Integrating transcriptomic and proteomic analyses provides a comprehensive understanding of CCE-regulated gene expression.