Intracellular membrane trafficking is crucial for eukaryotic cells. Versatile membranous carriers are endosomes that participate, for example, in the recycling of membrane proteins. In this project we study two additional functions of dynamically shuttling endosomes in Ustilago maydis (i) transport of mRNAs during hyphal growth and (ii) unconventional secretion during yeast growth. Endosomal mRNA transport is mediated by the key RNA-binding protein Rrm4 that is linked via the lipid anchor protein Upa1 to the surface of endosomes. A detailed structure/function analysis revealed that Rrm4 uses two unconventional MLLE domains for interaction with specific peptide motifs in Upa1. In this project, (i) we will solve the co-structure of the protein complex to unambiguously define the structural requirements for the attachment of mRNAs to endosomes. Furthermore, we will study additional functionally important components that participate in assembly and transport of mRNAs on endosomes, processes that were recently also shown to operate in neurons. (ii) Analysing unconventional secretion of Cts1 in the yeast form revealed that it is cell cycle-dependent. It occurs by a lock-type mechanism via a defined zone of unconventional secretion. This zone connects mother and daughter cell prior to cell division and is delimited by two consecutively formed septa. Formation of the secondary septum depends on motile early endosomes and is crucial for unconventional secretion. In addition, a novel essential factor for unconventional secretion, Jps1, was identified in a genetic screen. The protein co-localizes with chitinase Cts1 in the zone of unconventional secretion and binds phosphatidylinositol phosphates suggesting membrane association. We hypothesize that Jps1 acts as an anchoring factor. Now, we aim on further characterizing the role of Jps1 and its membrane association for unconventional secretion using a comprehensive structure/function analysis combined with identification of interaction partners specifically in the zone of unconventional secretion.
Project leaders: Dr. Kerstin Schipper, , and Prof. Dr. Michael Feldbrügge, ,
Researchers: M. Sc. Senthil Devan,
M. Sc. Ashiq Ayub,