Phospholipase-C gamma-1 (PLCgamma-1) is critical in hepatocyte growth factor induced in vitro invasion and migration without affecting the growth of prostate cancer cells.
BACKGROUND
Phospholipase C gamma-1 (PLCgamma-1) is an intracellular signalling molecule regulating a number of biological processes including transporter mechanisms, transcription factors, and scaffolding proteins mediating cytoskeleton and membrane trafficking. Hepatocyte growth factor (HGF) is a pleiotrophic factor and a mediator of metastatic spread. This study sought to determine the effect of HGF on the invasive and migratory potential of prostate cancer cells targeted by a ribozyme transgene to PLCgamma-1.


METHODS
A ribozyme transgene consisting of hammerhead ribozyme and antisense specific to PLCgamma-1 was cloned into a PEF6 expression vector and transfected into PC-3 cells. RT-PCR and Western blotting confirmed knock down of PLCgamma-1. In vitro invasion and a cytodex-2 bead motility assays with in vitro and in vivo growth models were used to assess the impact of PLCgamma-1 manipulation.


RESULTS
PC-3 cells stably transfected with PLCgamma-1 ribozyme transgene (PC-3(DeltaPLC)gamma) manifested a reduction of PLCgamma-1 expression at mRNA/protein levels. HGF/SF increased invasiveness (P < 0.01) and motility (P < 0.0001) of PC-3(WT) and PC-3(PEF6) cells. In contrast, PLCgamma-1 knock down PC-3(DeltaPLC)gamma cells had reduced invasiveness (P < 0.05) and motility (P < 0.01) compared with PC-3(WT) and PC-3(PEF6) cells. Although there was a marginal change of cell growth in vitro, there was no difference in the rate of growth between PC-3(DeltaPLC)gamma, PC-3(WT), and PC-3(PEF6) cells.


CONCLUSIONS
Targeting PLCgamma-1 by way of a hammerhead ribozyme to PLCgamma-1 is an effective method in reducing invasive phenotype of prostate cancer. PLCgamma-1 is a signalling intermediate that has prime influence on HGF induced cellular invasion and migration without affecting the growth of the cells.
Davies
Gaynor
G
Metastasis and Angiogenesis Research Group, Department of Surgery, School of Medicine, Cardiff University, Cardiff, United Kingdom.
Martin
Tracey A
TA
Ye
Lin
L
Lewis-Russell
Jonathon M
JM
Mason
Malcolm D
MD
Jiang
Wen G
WG
D045744:Cell Line, Tumor; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D017228:Hepatocyte Growth Factor / Q000494:pharmacology*; D006801:Humans; D008297:Male; D009361:Neoplasm Invasiveness; D051966:Phospholipase C gamma / Q000502:physiology*; D011471:Prostatic Neoplasms / Q000473:pathology*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014162:Transfection
D017228:Hepatocyte Growth Factor; D051966:Phospholipase C gamma