Deciphering endosomal sorting mechanisms in epithelial cells

• Applicant: Privatdozent Dr. Veit Riechmann
• Subject Area: Developmental Biology ; Cell Biology
• Term: from 2013 to 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 243729385

Sorting of endocytosed proteins is central for cellular homeostasis. Proteins can be either sorted for degradation in the lysosome or into the different recycling pathways. We know little about the molecular mechanisms controlling sorting, and so far no Rab GTPase is known that orchestrates endosomal sorting. We have identified RabX1 as a central regulator for recycling and degradation of adhesion proteins in the Drosophila follicular epithelium. Our data indicate that RabX1 facilitates endosomal sorting by connecting different Rab domains. Thus, RabX1 provides an excellent starting point to elucidate the mechanisms controlling endosomal sorting. As Rab proteins recruit different effectors and are themselves regulated by activators and inhibitors, we aim to identify such regulators and effectors for RabX1. We will purify activated, inactivated and wild type forms of RabX1 specifically from the follicular epithelium to identify co-precipitating proteins by mass spectrometry. The function of the co-precipitating proteins will be tested in elaborated genetic assays to identify candidates with a function in sorting. The assays are designed to find out if the candidates act as activators, inhibitors or effectors of RabX1. We will combine histological stainings with live imaging to examine how the depletion of candidates affects the dynamics of the RabX1 compartment. Moreover, we will analyse the sorting process in living epithelia of wild type and knockdown flies. Collectively these experiments will elucidate the different pathways by which RabX1 controls sorting and thereby advance our understanding of how a sorting endosome works. We are in an excellent position to carry out the proposed experiments as we acquired a great expertise in analysing vesicle trafficking in the follicular epithelium over the last years.

• DFG Programme: Research Grants