A crucial step in the embryonic development of insects is the decision whether the early anlage of the limb bud is evaginating to form the embryonic leg, or whether the anlage is invaginating to form an imaginal disc. In contrast to Drosophila, the beetle Tribolium develops during embryonic development clearly formed appendages from limb buds. The mechanism of apical constriction of cells leading to invagination of tissue has been described in many organisms.  How the evagination (of the single-layered epithelium) of the appendages is directed is not known. The distal outgrowth of the limb is a process combining cell shape changes and cell proliferation. Thereby the proportions in relation to the size are determined. Several important genes of Tribolium leg development have been analysed already and confirm data from Drosophila. The Distal-less gene is responsible for the direction of the proximo-distal outgrowth, whereas the Wnt signaling pathway is regulating the dorso-ventral axis in the appendages. The Sp8 gene is involved in Tribolium as well as in the mouse in the allometric size regulation of the appendages.

We analyse the following questions:

• How is the early limb field allocated in Tribolium and which molecular and cellbiological events initiate the evagination?
• How are cell shape changes and cell proliferation contributing to the outgrowth of the appendages?
• How are the proportions of the limb segments and the limb proper regulated in relation to the body size?

Our experimental approach is:

• direct isolation and analysis of candidate genes in Tribolium, which are part of this processes
• extensive analysis of already isolated genes and genes from the iBeetle RNAi screen
• the analysis of the cell biology of evagination and outgrowth in Tribolium with cell markers and antibodies synthesis of transgenic constructs for misexpression and overexpression studies of selected genes in Tribolium

We expect that the results shed light on the molecular and cellular basis of the development of the appendages in Tribolium and are therefore an important contribution to the understandung of development and evolution of the limb fields in Invertebrates. In the future we will combine our results with the new genes from the iBeetle RNAi screen and will look for the correlation to comparable processes in Vertebrates