Integrin linked kinase (ILK) is required for lens epithelial cell survival, proliferation and differentiation.
While the role of growth factors in lens development has been investigated extensively, the role of extracellular matrix signalling is less well understood. The developing lens expresses predominantly laminin-binding integrins (such as α3β1, α6β1), which are cooperatively required in the lens epithelium during development. We investigated the role of ILK, a downstream mediator of integrin signalling in mice conditionally null for Ilk. Mutant lenses showed epithelial thinning at E17.5 with reduced proliferation and epithelial cell number and aberrant fibre differentiation. There was complete loss of the central epithelium from postnatal day (P) 2 due to cell death followed by fibre cell degeneration and death by P10 as well as rupture of the lens capsule between P10 and P21. At E17.5 there was significant inhibition (∼50%) of epithelial cell cycle progression, as shown by BrdU incorporation, cyclin D1/D2 and phospho-histone H3 immunostaining. The epithelial marker, E-cadherin, was decreased progressively from E17.5 to P2, in the central epithelium, but there was no significant change in Pax6 expression. Analyses of ERK and Akt phosphorylation indicated marked depression of MAPK and PI3K-Akt signalling, which correlated with decreased phosphorylation of FRS2α and Shp2, indicating altered activation of FGF receptors. At later postnatal stages there was reduced or delayed expression of fibre cell markers (β-crystallin and p57(kip2)). Loss of Ilk also affected deposition of extracellular matrix, with marked retention of collagen IV within differentiating fibre cells. By quantitative RT-PCR array there was significantly decreased expression of 19 genes associated with focal adhesions, actin filament stability and MAPK and PI3K/Akt signalling. Overall, these data indicate that ILK is required for complete activation of signalling cascades downstream of the FGF receptor in lens epithelium and fibre cells during development and thus is involved in epithelial proliferation, survival and subsequent fibre differentiation.
Ocular Development Laboratory, Anatomy and Neuroscience, University of Melbourne, Victoria 3010, Australia.
Zhi Ling
ZL
Teo
Ocular Development Laboratory, Anatomy and Neuroscience, University of Melbourne, Victoria 3010, Australia.
Lachlan
L
McQueen-Miscamble
Ocular Development Laboratory, Anatomy and Neuroscience, University of Melbourne, Victoria 3010, Australia.
Kirsty
K
Turner
Ocular Development Laboratory, Anatomy and Neuroscience, University of Melbourne, Victoria 3010, Australia.
Gemma
G
Martinez
Zoology, Miami University, Oxford, OH 45056, USA.
Bhavani
B
Madakashira
British Columbia Cancer Research Centre, Vancouver, B.C., Canada.
Shoukat
S
Dedhar
Zoology, Miami University, Oxford, OH 45056, USA.
Michael L
ML
Robinson
Ocular Development Laboratory, Anatomy and Neuroscience, University of Melbourne, Victoria 3010, Australia. Electronic address: r.deiongh@unimelb.edu.au.
Robb U
RU
de Iongh
D000818:Animals; D015153:Blotting, Western; D015820:Cadherins / Q000378:metabolism; D002454:Cell Differentiation / Q000502:physiology*; D049109:Cell Proliferation; D002470:Cell Survival / Q000502:physiology*; D019938:Cyclin D1 / Q000378:metabolism; D056742:Cyclin D2 / Q000378:metabolism; D004847:Epithelial Cells / Q000166:cytology* / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D019084:Fluorescent Antibody Technique, Indirect; D007908:Lens, Crystalline / Q000196:embryology* / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000502:physiology*; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction; D000099200:Scaffold Protein ILK
D015820:Cadherins; D019938:Cyclin D1; D056742:Cyclin D2; D048049:Extracellular Signal-Regulated MAP Kinases; D017346:Protein Serine-Threonine Kinases; D051057:Proto-Oncogene Proteins c-akt; D000099200:Scaffold Protein ILK; C530105:Ccnd1 protein, mouse; C486196:Ccnd2 protein, mouse
EMT; MAPK signalling; PI3K signalling; apoptosis; cell cycle; integrin linked kinase; lens development; signalling