Die Rolle des RNA-bindenden Proteins Rrm4 während des polaren Wachstums von Ustilago maydis

Microtubule-dependent transport of mRNA in form of ribonucleoprotein (RNP) particles is a common mechanism for local restriction of protein synthesis and is essential for the establishment or maintenance of cell polarity during differentiation and development. Pathogenic development of Ustilago maydis depends on the formation of infectious filaments that grow with a defined axis of polarity. The RNA-binding protein Rrm4 mediates microtubule-dependent transport of distinct mRNAs in these filaments and loss of transport correlates with defects in polarity. Although a number of potential target mRNAs could be identified, cellular processes that depend on Rrm4-mediated transport remain largely unknown. Based on differential proteomics we identified seven differentially regulated protein-variants in rrm4Δ filaments involved in cell wall remodelling, translation and mitochondrial function. Consequently, we uncovered that the mRNA encoding the bacterial-type endochitinase Cts1 represents a direct target of Rrm4. In wild type filaments the encoded protein localizes predominantly at the growth cone, whereas it accumulates at both poles in rrm4Δ filaments. Filaments of cts1Δ mutants aggregate in liquid culture suggesting an altered cell surface. By activity measurements we demonstrated that Cts1 is secreted and associates most likely with the cell wall of filaments. Secretion is drastically impaired in filaments lacking Rrm4 or conventional kinesin Kin1 as well as in filaments with disrupted microtubules. Thus, Rrm4-mediated mRNA transport appears to be essential for efficient export of Cts1, uncovering a novel molecular link between mRNA transport and secretion. Additionally we identified three proteins of the mitochondrial inner membrane including the AAA protease Rca1. Preliminary results indicate that efficient mitochondrial import is disturbed in rrm4 strains resulting in increased production of mitochondrial superoxide. To get more insights into the process of this mRNA-transport, we performed localization studies of the poly(A)-binding protein Pab1. Pab1 colocalizes with Rrm4 in vivo in almost all shuttling particles. Interestingly, in rrm4 filaments, Pab1 is no longer present in shuttling particles. Thus, the RNA-binding protein Rrm4 constitutes a key component of microtubule-dependent mRNA transport in the filamentous fungus Ustilago maydis. It seems that active mRNP transport is involved in apical secretion and might be important to support symmetric import of mitochondrial proteins.