Developmental Genetics

Phosphoregulation during morphogenensis in the fly (Drosophila melanogaster)
 

Regulation of the single MAST Kinase homolog in Drosophila encoded by the drop out gene

Background and previous work

MAST (microtubule associated Ser/Thr) kinases belong to the superfamily of the AGC (Protein Kinase A, Protein Kinase G, Protein Kinase C) kinases. The human MAST kinase family comprises four homologues, called MAST1-4, while the Drosophila genome encodes a single MAST kinase, called Drop out(Dop). MAST share a highly conserved domain structure consisting of a DUF1908 (domain of unknown function) domain, a kinase domain and a PDZ (PSD95, Dlg, ZO-1) domain. Human MAST kinases are implicated in several diseases, like inflammatory bowel disease, rabies and breast cancer. MAST kinases are widely expressed, but their regulation is poorly understood. Some interaction partners are known, for example tumor suppressor PTEN (phosphatase and tensin homolog deleted on chromosome 10) binds to PDZ domain of MAST, and the E3 ubiquitin ligase TRAF6, which plays a role in the NF-κB pathway. Their involvement in distinct molecular pathways and in human diseases raises the question about the regulatory mechanisms controlling MAST activity in different cell biological contexts. To investigate regulatory mechanisms of MAST kinases, D. melanogaster offers a perfect model system, because a range of mutations have been created in the dop gene, which seriously affect early embryogenesis and other developmental processes. The most severe phenotype in dop mutant embryos represents a complete block of cell formation at the transition from syncytial blastoderm to cellular blastoderm embryo. Nuclei drop out of the embryo cortical cytoplasm, membrane proteins are mislocalised and furrow canal formation is blocked, leading to lethality. This project focuses on the regulation of Dop activity through its subcellular localization, potentially by action of highly conserved protein domains, and its regulation through post-translational modifications, including phosphorylation and ubiquitinylation.

Specific aims

Our study on the regulation of Drosophila MAST kinase, Dop include different approaches. To this end, the conserved protein domains will be analyzed regarding regulation, but also possible interaction partners of these domains, especially with the DUF1908 and PDZ domain. Another aim is the identification and functional analysis of post-translational modifications.

Working program

·         functional genetic studies on DUF1908 and PDZ domains

·         identification of interaction partners of DUF and PDZ domains (collaboration with P5)

·         analysis of posttranslational modifications and their function (collaboration with P1)

Graphical abstract