EMBO Conference on Protein Synthesis and Translational Control

DIERK NIESSING
205
Poster Abstracts
An Extended RNA-binding Surface of She2p Oligomers is Required for mRNP
Assembly and Localization
Dierk Niessing 1, Marisa Müller 1, Ralf-Peter Jansen 2
1 Helmholtz Zentrum München, Germany
2 University of Tübingen, Germany
In eukaryotic cells, dozens to hundreds of different mRNAs are localized by specialized
motor-dependent transport complexes. We addressed the open question how the
RNA-binding protein She2p of a yeast mRNA-transport complex specifically binds to several
transcripts and how nuclear mRNA binding influences cytoplasmic transport-complex function.
Previous results suggested that She2p is dimeric in solution. In this study, we determined that
She2p forms tetrameric complexes that are required for mRNA binding in vitro and transcript
localization in vivo. We further observed that She2p has an only moderately higher affinity for
localizing transcripts when compared to unrelated stem-loop RNAs. However, for specific
binding to localizing transcripts She2p requires an extended RNA-binding surface, whereas for
unspecific RNA binding fewer surface features are sufficient. Mutation of the extended She2p
surface selectively impairs in-vitro binding to localizing RNAs, cytoplasmic mRNP assembly in
vivo, and its subsequent mRNA localization. The lack of a strong difference in affinity to
mediate selectivity might be due to She2p’s requirement to recognize dozens of target RNAs in
the nucleus with similar, yet distinct RNA-folding features. However in the cytoplasm, RNAs
bound by She2p via its extended RNA-binding surface assemble with She3p into stable, highly
specific transport complexes. In summary, our study shows that specificity at early, nuclear
steps of complex assembly is not primarily mediated by higher affinity. In addition, we provide
evidence for a close mechanistic dependence of cytoplasmic mRNP assembly on the spatially
separated, preceding nuclear mRNA binding events by She2p.