Skin Development - Genetic Analysis of Cellular Adhesion in the Developing Zebrafish Epidermis

The skin is the outermost organ system, which covers all the internal organs and seals the body from the outside. It prevents the loss of body fluids and provides protection from pathogens. It is composed of the outer epidermis, a stratified epithelium, and the underlying dermis of mesodermal origin. The epidermal integrity is essential for most of the functions that the skin performs. Various cellular junctions form an integral part of this stratified epithelium. The tight junctions are essential for preventing loss of body fluids due to evaporation whereas adherens junctions are essential for maintenance of the epidermal homeostasis. The hemidesmosomes and desmosomes, which mediate the cell-basal membrane and cell-cell adhesion, respectively, are essential to maintain the integrity of the epidermis and its attachment to the underlying basal lamina.

We combine forward and reverse genetic approaches to uncover the genetic pathways and molecular mechanisms involved in assembling cellular junctions such as hemidesmosomes and adherens junctions during epidermal development. We have shown that the function of penner/lethal giant larvae 2 (lgl2) is essential for the formation of hemidesmosomes. Lgl2 mediates the targeting of the hemidesmosomal component Integrin alpha 6 to the membrane. In addition, Lgl2 and E-cadherin act antagonistically to regulate the number of hemidesmosomes.

The Drosophila lgl is a tumour suppressor gene, and mutants display hyperplasia of the brain tissue, which loses its compact structure. In the zebrafish, in absence of pen/lgl2 function the skin cells of the larvae hyper-proliferate, get spindle shaped and become highly motile (see figure below). We are investigating a molecular mechanism responsible for imparting growth advantage and migratory potential to epidermal cells in absence of pen/lgl2 function.

_{The loss of lgl2 function leads to acquisition of migratory behaviour and epidermal cell proliferation. A, B  Immuno-histological staining in WT (A) and pen mutant (B) using anti keratin (Ks pan-8) antibody. Epidermal cells exhibiting keratin staining are never present in fin fold of WT larvae (A). In contrast, mutant larvae show presence of keratin containing cells in fin folds (arrow in B). These cells are spindle shaped and appear to migrate from the lateral epidermis into fin folds. C,D, Assessment of cell proliferation using BrdU incorporation assay.  The number of proliferating cells in ventral epidermis is higher in pen mutant (D) as compared to WT (C). Scale bar in D is equal to 40mm in A-B and 0.2mm in C, D, respectively}

By participating in the large-scale genetic screen done in year 2005-2006, we have isolated 30 mutants with specific skin defects such as blistering of the skin or detachment of epithelial cells. Several of the mutants with blistering phenotypes are reminiscent of a human genetic disorder, Epidermolysis bullosa. In several of these mutants targeting of the adherens junction component E-cadherin or a hemidesmosomal component Integrin alpha 6 to the membrane is altered. We aim to clone these loci to identify additional components that are involved in formation of cellular junctions in the epidermis.

Personnel:

• Dr. Mahendra Sonawane                               Group leader MPG Partner Project, Tata Institute, Mumbai (01/09 - )
  

• Sven Reischauer                                             PhD student (01/04 - 01/10)                                        

Collaborators:

• Dr. Robert Geisler                                           KIT, Karlsruhe, Germany
• Dr. Heinz Schwarz                                           Electron Microscopy Unit, MPI EB       

Key Publications:

Sonawane, M., Carpio, Y., Geisler, R., Schwarz, H., Maischein, H.-M. and Nüsslein-Volhard, C. (2005): Zebrafish penner/lethal giant larvae 2 functions in hemidesmosome formation, maintenance of cellular morphology and growth regulation in the developing basal epidermis. Development 132: 3255-3265