Functional Regulation of DAPK1 Serine 308 Phosphorylaton in Drosophila

• Main Authors: CHEN, SZU-WEN, 陳似玟
• Other Authors: CHA, TAI-LUNG
• Format: Others
• Language: zh-TW
• Published: 2019
• Online Access: http://ndltd.ncl.edu.tw/handle/z2v949

碩士 === 國防醫學院 === 生物化學研究所 === 107 === DAPK (Death Associated Protein Kinase) is a Ca2+/calmodulin-regulated serine/threonine kinase. DAPK can suppress carcinogenic factors (such as c-myc or E2F) and also affects tumor metastasis. It is also considered to be an important tumor suppressor. Phosphorylation on different amino acid residues in DAPK causes its structural and functional changes. Previous studies showed that autophosphorylation on Serine 308 results in decreasing kinase activity of DAPK. Our lab has discovered that phosphorylation of S308 site causes mitochondrial morphological changes in culture cells. However, the detail molecular mechanism is still unclear. In addition, it is not known whether there are other kinases capable of phosphorylating DAPK S308 site. Drosophila muscle tissue has been widely used in studying mitochondrial dynamics. To investigate the effect of DAPKS308 phosphorylation on mitochondria, we established transgenic flies bearing different DAPKS308 point mutations (Wild Type; S308A: blocks phosphorylation; S308D: mimics phosphorylation). When overexpressing different forms of DAPK in muscle tissue, we found that the transgenic animals exhibit different extent of mortality (S308A>WT>S308D). These results are consistent with previous experiments conducted in culture cells. Furthermore, we found that the kinase activity of DAPK nicely correlates with mitochondrial elongation in fly muscle tissue. These results suggest that DAPK kinase activity might regulate mitochondrial fusion/fission in vivo. Intriguingly, we still observed the S308 being phosphorylated in muscles and eyes when overexpressing kinase dead mutant DAPKK42A (loss of autophosphorylation). This indicates that other unknown kinases are able to phosphorylate S308 in Drosophila. Therefore, we conducted a kinome screening to look for kinases that might phosphorylate DAPK S308 residue in vivo. Currently, our preliminary screening has discovered several candidate S308 kinases which are needed for further validation in the future.