Dissection of DEK1 evolution and function in position dependant epidermis cell fate specification.

This project studies the DEK1 protein which is essential for epidermal cell fate specification in most, if not all plants. DEK1 is a 240 kDa protein that is anchored in the plasma membrane by 21 membrane segments with a predicted external loop region and a C-terminal calpain proteinase linked to the membrane domain by the DEK1-Hand. In Arabidopsis, DEK1 is essential for formation of all epidermal cells. The moss Physcomiterella patens contains a single Dek1 gene that has 74% simality to maize DEK1, sugge sting that DEK1 assumed a vital role in plant development over 500 million years ago. Chalmydomonas, a single celled fresh water algae, contains a cytosolic calpain that has high similarity to the calpain of DEK1, suggesting that the DEK1 calpain fused to the membrane part of DEK1 during the transition from single cells to multi-cellular land plants. The project has three main objectives, (I) to identify of the when the transition from cytosolic to membrane bound calpains (DEK1) took place and its funct ional significance during land plant evolution. This task involves (i) cequencing of representative species in the dendrogram between chlamy and Physco, (ii) characterization of plant body organization of species representing the transition from a cytoso lic calpain to DEK1 by microscopy and cell biology tools, (iii) determination of DEK1 function in Physcomiterella patens and (iv) Chalmydomonas. The second objective is to (II) to identify essential functional DEK1 amino acid residues, (i) by sequencin g representative species from Physcomitrella patens to Arabidopsis thaliana (ii) to asses their functional role in genetic complementation tests1, (iii) to assess the functional significance of conserved residues in the loop region using the dominant neg ative effect of over-expressing the DEK1-MEM in Arabidopsis thaliana. The third main objective (iii) is the structural determination of DEK1 using X-rax diifraction, with emphasis on the DEK1 loop and arm regions.