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Title: Regulation of dual specificity phosphatase 1 gene expression by glucocorticoids
Author: Silva Martins, Joana Rita
ISNI:       0000 0004 2686 4739
Awarding Body: Imperial College London
Current Institution: Imperial College London
Date of Award: 2010
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Dual specificity phosphatase 1 (DUSP1) limits the expression of pro-inflammatory gene products by dephosphorylating and inactivating mitogen-activated protein kinases (MAPKs), in particular p38 MAPK and cJun N-terminal kinase. In many mouse and human cell types, glucocorticoids (GCs) increase the expression of DUSP1. The resulting inhibition of intracellular signalling pathways is thought to contribute to the powerful anti-inflammatory effects of GCs. The purpose of this project was to define mechanisms by which GCs control expression of mouse and human DUSP1 genes. Orthologous genes of distantly related species often respond to GCs via regulatory elements that are evolutionarily conserved in sequence and position. Therefore interspecies sequence comparison was used as one tool to identify GC-responsive regions (GRRs) of mouse and human DUSP1 genes. Functional assays of GRRs were carried out by transient and stable transfection of mouse and human cells. Interactions of these cis-acting elements with transcription factors (including the GC receptor itself, GR) were assessed by in vitro DNA-binding assays and chromatin immunoprecipitation. A number of conserved, putative GRRs were identified up to 29 kb upstream of the DUSP1 transcription start site (TSS). Surprisingly, some of these were found to be differently utilised by mouse and human DUSP1 genes, in spite of their sequence similarity. Strongly GC-responsive elements were found 4.6 and 1.3 kb upstream of the human DUSP1 TSS. The corresponding regions of the mouse DUSP1 gene were unresponsive or very weakly responsive to GC. Instead the mouse gene appeared to be regulated via an element 29 kb upstream of the TSS, the corresponding human element being unresponsive. GC responsiveness correlated with the ability to recruit GR in vivo, but could not be explained on the basis of differences in the sequences of GR binding sites. These observations suggest that recruitment of GR to DUSP1 loci and transcriptional activation in response to GC are critically dependent on “accessory” transcription factors. GC responses of mammalian DUSP1 genes appear to have evolved via gain and/or loss of binding sites for accessory factors, whilst maintaining overall output. GC regulation of DUSP1 genes involves atypical GR binding sites and, unusually, does not appear to require dimerisation of GR.
Supervisor: Clark, Andy Sponsor: Arthritis Research Campaign
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral