PgmNr D1020: RPTPs mediate the temporal control of Drosophila airway maturation through an interaction with the non-receptor tyrosine kinase Btk29A and its effector WASH.

Authors:
V. Tsarouhas 1 ; D. Liu 1 ; G. Tsikala 1 ; K. Zinn 2 ; C. Samakovlis 1


Institutes
1) Stockholm University, Stockholm, Sweden; 2) California Institute of Technology, Pasadena, CA, USA.


Keyword: endocytosis and membrane dynamics

Abstract:

The respiratory tubes of mammalian lungs and the Drosophila tracheal system undergo a series of maturation events at the end of embryogenesis. An important transition during Drosophila airway maturation is the clearance of the solid luminal material before the replacement of luminal liquid by gas. We had shown earlier that luminal protein clearance depends on a wave of apical endocytosis but its regulation is largely unknown. Here we show that Drosophila type III receptor tyrosine phosphatases (RPTPs), Ptp4E and Ptp10D restrict tube growth and apical endocytosis during embryonic airway maturation. Live imaging of mutant embryos lacking both RPTP genes revealed an earlier clearance of GFP-tagged luminal proteins compared to wild type embryos. Genetic alterations in endocytosis by the generation of ptp4E10D and rab5 or vps45 triple mutants could ameliorate luminal clearance phenotypes suggesting elevated endocytic levels in ptp4E10D mutants. Quantifications of the intercellular vesicles through an established endocytic Dextran assay in live embryos revealed a direct role of RPTPs in the temporal control of luminal endocytosis. Endo-lysosomal analysis showed an enhanced endocytic trafficking oriented towards the degradation pathway in ptp4E10D mutants.  Following a short scale genetic screen we identified the non-receptor tyrosine kinase Btk29A as a downstream target of RPTPs with ptp4E10D;btk29A triple mutants showing rescue in tube size and premature luminal clearance. Immuno-precipitation experiments from embryo extracts confirmed a physical interaction between Ptp10D and Btk29A and revealed binding of Btk29A to actin nucleator protein WASH. Single mutant analysis showed that both Btk29A and WASH proteins are required for the endocytic clearance of luminal material. Interestingly, loss of wash in ptp4E10D mutants suppressed the tube size defects suggesting an antagonistic role of RPTPs in Btk29A/WASH pathway. In summary, we show that RPTPs and Btk29A have antagonistic functions on WASH mediated actin polymerization to control the timing of luminal protein clearance in Drosophila airways.



Flybase Genetic Index:
1. FlyBase gene symbol: Ptp4E; FBgn: FBgn0004368
2. FlyBase gene symbol: Ptp10D; FBgn: FBgn0004370
3. FlyBase gene symbol: Btk29A; FBgn: FBgn0003502
4. FlyBase gene symbol: wash; FBgn: FBgn0033692
5. FlyBase gene symbol: Rab5; FBgn: FBgn0014010
6. FlyBase gene symbol: Vps45; FBgn: FBgn0261049