Analysis of germline GLI1 variation implicates hedgehog signalling in the regulation of intestinal inflammatory pathways.
BACKGROUND
Ulcerative colitis (UC) and Crohn's disease (CD) are polygenic chronic inflammatory bowel diseases (IBD) of high prevalence that are associated with considerable morbidity. The hedgehog (HH) signalling pathway, which includes the transcription factor glioma-associated oncogene homolog 1 (GLI1), plays vital roles in gastrointestinal tract development, homeostasis, and malignancy. We identified a germline variation in GLI1 (within the IBD2 linkage region, 12q13) in patients with IBD. Since this IBD-associated variant encodes a GLI1 protein with reduced function and our expression studies demonstrated down-regulation of the HH response in IBD, we tested whether mice with reduced Gli1 activity demonstrate increased susceptibility to chemically induced colitis.


RESULTS
Using a gene-wide haplotype-tagging approach, germline GLI1 variation was examined in three independent populations of IBD patients and healthy controls from Northern Europe (Scotland, England, and Sweden) totalling over 5,000 individuals. On log-likelihood analysis, GLI1 was associated with IBD, predominantly UC, in Scotland and England (p < 0.0001). A nonsynonymous SNP (rs2228226C-->G), in exon 12 of GLI1 (Q1100E) was strongly implicated, with pooled odds ratio of 1.194 (confidence interval = 1.09-1.31, p = 0.0002). GLI1 variants were tested in vitro for transcriptional activity in luciferase assays. Q1100E falls within a conserved motif near the C terminus of GLI1; the variant GLI protein exhibited reduced transactivation function in vitro. In complementary expression studies, we noted the colonic HH response, including GLI1, patched (PTCH), and hedgehog-interacting protein (HHIP), to be down-regulated in patients with UC. Finally, Gli1(+/lacZ) mice were tested for susceptibility to dextran sodium sulphate (DSS)-induced colitis. Clinical response, histology, and expression of inflammatory cytokines and chemokines were recorded. Gli1(+/lacZ) mice rapidly developed severe intestinal inflammation, with considerable morbidity and mortality compared with wild type. Local myeloid cells were shown to be direct targets of HH signals and cytokine expression studies revealed robust up-regulation of IL-12, IL-17, and IL-23 in this model.


CONCLUSIONS
HH signalling through GLI1 is required for appropriate modulation of the intestinal response to acute inflammatory challenge. Reduced GLI1 function predisposes to a heightened myeloid response to inflammatory stimuli, potentially leading to IBD.
Lees
Charlie W
CW
Gastrointestinal Unit, Molecular Medicine Centre, University of Edinburgh, Edinburgh, United Kingdom.
Zacharias
William J
WJ
Tremelling
Mark
M
Noble
Colin L
CL
Nimmo
Elaine R
ER
Tenesa
Albert
A
Cornelius
Jennine
J
Torkvist
Leif
L
Kao
John
J
Farrington
Susan
S
Drummond
Hazel E
HE
Ho
Gwo-Tzer
GT
Arnott
Ian D R
ID
Appelman
Henry D
HD
Diehl
Lauri
L
Campbell
Harry
H
Dunlop
Malcolm G
MG
Parkes
Miles
M
Howie
Sarah E M
SE
Gumucio
Deborah L
DL
Satsangi
Jack
J
D000328:Adult; D000818:Animals; D004739:England; D005260:Female; D020869:Gene Expression Profiling; D020022:Genetic Predisposition to Disease; D005820:Genetic Testing; D018095:Germ-Line Mutation; D053823:Hedgehog Proteins / Q000235:genetics / Q000502:physiology*; D006801:Humans; D007249:Inflammation / Q000235:genetics; D015212:Inflammatory Bowel Diseases / Q000235:genetics*; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008875:Middle Aged; D020411:Oligonucleotide Array Sequence Analysis; D020641:Polymorphism, Single Nucleotide; D012606:Scotland; D015398:Signal Transduction / Q000235:genetics* / Q000276:immunology; D013548:Sweden; D014157:Transcription Factors / Q000235:genetics*; D000071676:Zinc Finger Protein GLI1
C400551:GLI1 protein, human; D053823:Hedgehog Proteins; D014157:Transcription Factors; D000071676:Zinc Finger Protein GLI1