MAPK/ERK1/2 and PI3-kinase signalling pathways are required for vitreous-induced lens fibre cell differentiation

Gata3 is a DNA-binding transcription factor involved in cellular differentiation in a variety of tissues including inner ear, hair follicle, kidney, mammary gland and T-cells. In a previous study in 2009, Maeda et al . ( Dev. Dyn. 238 , 2280–2291; doi:10.1002/dvdy.22035 ) found that Gata3 mutants could be rescued from midgestational lethality by the expression of a Gata3 transgene in sympathoadrenal neuroendocrine cells. The rescued embryos clearly showed multiple defects in lens fibre cell differentiation. To determine whether these defects were truly due to the loss of Gata3 expression in the lens, we generated a lens-specific Gata3 loss-of-function model. Analogous to the previous findings, our Gata3 null embryos showed abnormal regulation of cell cycle exit during lens fibre cell differentiation, marked by reduction in the expression of the cyclin-dependent kinase inhibitors Cdkn1b/p27 and Cdkn1c/p57, and the retention of nuclei accompanied by downregulation of Dnase IIβ. Comparisons of transcriptomes between control and mutated lenses by RNA-Seq revealed dysregulation of lens-specific crystallin genes and intermediate filament protein Bfsp2. Both Cdkn1b/p27 and Cdkn1c/p57 loci are occupied in vivo by Gata3, as well as Prox1 and c-Jun, in lens chromatin. Collectively, our studies suggest that Gata3 regulates lens differentiation through the direct regulation of the Cdkn1b/p27and Cdkn1c/p57 expression, and the direct/or indirect transcriptional control of Bfsp2 and Dnase IIβ.

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Vimentin and CP49/filensin form distinct networks in the lens which are independently modulated during lens fibre cell differentiation

Journal of Cell Science ◽

10.1242/jcs.108.4.1397 ◽

1995 ◽

Vol 108 (4) ◽

pp. 1397-1406 ◽

Cited By ~ 2

Author(s):

A. Sandilands ◽

A.R. Prescott ◽

J.M. Carter ◽

A.M. Hutcheson ◽

R.A. Quinlan ◽

...

Keyword(s):

Cell Differentiation ◽

Epithelial Cells ◽

Confocal Microscopy ◽

Intermediate Filament ◽

Nuclear Lamina ◽

Intermediate Filament Protein ◽

Intermediate Filament Proteins ◽

Fibre Cell ◽

Lens Fibre ◽

Lens Fibre Cell

The cells of the eye lens contain the type III intermediate filament protein vimentin, as well as two other intermediate filament proteins, CP49 and filensin. These two proteins appear to be unique to the differentiated lens fibre cell. Immunoblotting and confocal microscopy were used to describe changes which occur in these three intermediate filament proteins and the networks they form during fibre cell differentiation and maturation. The vimentin network was present in both epithelial cells and some fibre cells. Fibre cells were vimentin positive up to a specific point 2–3 mm in from the lens capsule where the vimentin signal was drastically reduced. The CP49/filensin network was not present in the undifferentiated epithelial cells but emerged in the differentiating fibre cells. This latter network exhibited a principally plasma membrane localization in younger fibre cells but became more cytoplasmic in older fibre cells. This change also occurred at a distinct point in fibre cell differentiation, much earlier than the observed loss of the vimentin network. The subcellular changes in the distributions of these cytoskeletal networks were correlated to the loss of the fibre cell nucleus, another feature of fibre cell differentiation. No correlation was found to changes in the vimentin network but nuclear loss did coincide with changes in the CP49/filensin network. Concomitant with nuclear pyknosis, there were also changes in the nuclear lamina as well as infringement of the nuclear compartment by CP49, as shown by confocal microscopy. This study demonstrates vimentin and the CP49/filensin network to be independent in the lens but both networks undergo dramatic changes in subcellular distribution during the differentiation/maturation of the fibre cell. Only changes in the CP49/filensin network can be correlated to nuclear loss. Thus in the lens, unlike mammalian erythropoiesis which is also characterized by nuclear loss, the vimentin network does not appear linked to nuclear retention.

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Lens fibre cell differentiation and organelle loss: many paths lead to clarity

Philosophical Transactions of the Royal Society B Biological Sciences ◽

10.1098/rstb.2010.0324 ◽

2011 ◽

Vol 366 (1568) ◽

pp. 1219-1233 ◽

Cited By ~ 94

Author(s):

Michael A. Wride

Keyword(s):

Cell Differentiation ◽

Proteolytic Enzymes ◽

Specific Gene ◽

Fibre Cell ◽

Free Zone ◽

Lens Fibre ◽

Evolutionary Context ◽

Lens Fibre Cell ◽

Ubiquitin Proteasome ◽

Interacting Networks

The programmed removal of organelles from differentiating lens fibre cells contributes towards lens transparency through formation of an organelle-free zone (OFZ). Disruptions in OFZ formation are accompanied by the persistence of organelles in lens fibre cells and can contribute towards cataract. A great deal of work has gone into elucidating the nature of the mechanisms and signalling pathways involved. It is apparent that multiple, parallel and redundant pathways are involved in this process and that these pathways form interacting networks. Furthermore, it is possible that the pathways can functionally compensate for each other, for example in mouse knockout studies. This makes sense given the importance of lens clarity in an evolutionary context. Apoptosis signalling and proteolytic pathways have been implicated in both lens fibre cell differentiation and organelle loss, including the Bcl-2 and inhibitor of apoptosis families, tumour necrosis factors, p53 and its regulators (such as Mdm2) and proteolytic enzymes, including caspases, cathepsins, calpains and the ubiquitin–proteasome pathway. Ongoing approaches being used to dissect the molecular pathways involved, such as transgenics, lens-specific gene deletion and zebrafish mutants, are discussed here. Finally, some of the remaining unresolved issues and potential areas for future studies are highlighted.

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