siRNA were transfected into Cervical epithelial carcinoma HeLa cells using Lullaby siRNA transfection reagent.
this article demonstrate the high efficiency of Lullaby, siRNA transfection reagent from OZ Biosciences to induce gene silencing in HeLa cell lines.article reference: J Cell Sci. 2013 Jun 1;126(Pt 11):2381-91.
ERK2 but not ERK1 mediates HGF-induced motility in non-small cell lung carcinoma cell lines.Radtke S, Milanovic M, Rossé C, De Rycker M, Lachmann S, Hibbert A, Kermorgant S, Parker PJ.
Abstract
Aberrant signalling
of receptor tyrosine kinases (RTKs), such as c-Met, the receptor for
hepatocyte growth factor (HGF), has been implicated in the oncogenesis
of various tumours including non-small cell lung carcinoma (NSCLC).
Through its pro-migratory properties, c-Met has been implicated
specifically in the process of tumour metastasis, demanding a better
understanding of the underlying signalling pathways. Various players
downstream of c-Met have been well characterised, including the
extracellular-signal-regulated kinases (ERKs) 1 and 2. In a small
interfering RNA (siRNA)-based high-throughput wound healing screen
performed in A549 lung carcinoma cells, we identified ERK2 but not ERK1
as a strong mediator of HGF-induced motility. This finding was confirmed
in several NSCLC cell lines as well as in HeLa cells. One known
substrate for ERK kinases in cell migration, the focal adhesion protein
paxillin, was also one of the hits identified in the screen. We
demonstrate that HGF stimulation results in a time-dependent
phosphorylation of paxillin on serine 126, a process that can be blocked
by inhibition of the ERK1/2 upstream kinase mitogen-activated protein
kinase/ERK kinase 1 (MEK1) or inhibition of glycogen synthase kinase 3
(GSK3). Further, we show that paxillin turnover at focal adhesions is
increased upon stimulation by HGF, an effect that is dependent on serine
residues 126 (GSK3 site) and 130 (ERK site) within paxillin. In line
with the isoform-specific requirement of ERK2 for HGF-mediated migration
in lung tumour cell models, ERK2 but not ERK1 is shown to be
responsible for paxillin serine 126 phosphorylation and its increased
turnover at focal adhesions.
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