The Function of Heat Shock Protein 27 in Interleukin-1 Signalling
Author(s)
Brennan, Bryony Rachel
Type
Thesis
Abstract
The p38 mitogen activated protein kinase (MAPK) pathway enhances the expression of proinflammatory
response genes, including cyclooxgenase (COX)-2 and interleukin (IL)-6 in
response to stimuli including IL-1. Heat shock protein 27 (HSP27) is a major protein
phosphorylated by MAPK activated protein kinase 2 (MK2), a downstream kinase of p38
MAPK. However, the role of HSP27 in inflammation is unclear. Previous work has shown the
requirement of HSP27 for stabilisation of COX-2 and IL-6 mRNAs following IL-1 treatment of
HeLa cells. To examine the role of HSP27 in IL-1 signalling I depleted the protein from HeLa
cells, human alveolar epithelial (A549) cells and human dermal fibroblasts (HDFs) by RNA
interference (RNAi) using multiple short interfering RNAs (siRNAs).
HSP27 suppression in HeLa cells inhibited IL-1 induced activation of MK2, p38 MAPK and c-Jun N-terminal kinase (JNK), but not extracellular signal-regulated kinase (ERK). Upstream
MAPK kinase (MKK) 3, 4, 6 and 7 and TGF-beta-activated kinase 1 (TAK1) were also
inhibited. HSP27 depletion did not disrupt the constitutive interaction found between TAK1 and
its binding protein TAB1. HSP27 appeared to impinge on signalling at a point lying between
cell surface receptors and TAK1 or at TAK1 itself as activation of JNK and p38 MAPK by
tumour necrosis factor (TNF) was inhibited by HSP27 knockdown.
A transfection procedure was developed for transfection of siRNA into HDF cells. In HDFs and
A549 cell types HSP27 was required for COX-2 protein expression and the full activation of
p38 MAPK. However, while TAK1 was shown to be responsible for sustained activation of p38
MAPK, TAK1 activity was not affected by HSP27 depletion in HDFs. MKK3 (rather than
MKK6) appeared to be the major HDF activator of p38 MAPK in response to IL-1. My results
demonstrate that HSP27 has a novel role in augmenting IL-1 signalling in epithelial cell lines
and fibroblasts.
response genes, including cyclooxgenase (COX)-2 and interleukin (IL)-6 in
response to stimuli including IL-1. Heat shock protein 27 (HSP27) is a major protein
phosphorylated by MAPK activated protein kinase 2 (MK2), a downstream kinase of p38
MAPK. However, the role of HSP27 in inflammation is unclear. Previous work has shown the
requirement of HSP27 for stabilisation of COX-2 and IL-6 mRNAs following IL-1 treatment of
HeLa cells. To examine the role of HSP27 in IL-1 signalling I depleted the protein from HeLa
cells, human alveolar epithelial (A549) cells and human dermal fibroblasts (HDFs) by RNA
interference (RNAi) using multiple short interfering RNAs (siRNAs).
HSP27 suppression in HeLa cells inhibited IL-1 induced activation of MK2, p38 MAPK and c-Jun N-terminal kinase (JNK), but not extracellular signal-regulated kinase (ERK). Upstream
MAPK kinase (MKK) 3, 4, 6 and 7 and TGF-beta-activated kinase 1 (TAK1) were also
inhibited. HSP27 depletion did not disrupt the constitutive interaction found between TAK1 and
its binding protein TAB1. HSP27 appeared to impinge on signalling at a point lying between
cell surface receptors and TAK1 or at TAK1 itself as activation of JNK and p38 MAPK by
tumour necrosis factor (TNF) was inhibited by HSP27 knockdown.
A transfection procedure was developed for transfection of siRNA into HDF cells. In HDFs and
A549 cell types HSP27 was required for COX-2 protein expression and the full activation of
p38 MAPK. However, while TAK1 was shown to be responsible for sustained activation of p38
MAPK, TAK1 activity was not affected by HSP27 depletion in HDFs. MKK3 (rather than
MKK6) appeared to be the major HDF activator of p38 MAPK in response to IL-1. My results
demonstrate that HSP27 has a novel role in augmenting IL-1 signalling in epithelial cell lines
and fibroblasts.
Date Issued
2009-01
Date Awarded
2009-07
Copyright Statement
Attribution NoDerivatives 4.0 International Licence (CC BY-ND)
Advisor
Dean, Jon
Saklatvala, Jerry
Creator
Brennan, Bryony Rachel
Publisher Department
The Kennedy Institute of Rheumatology
Publisher Institution
Imperial College London
Qualification Level
Doctoral
Qualification Name
Doctor of Philosophy (PhD)