Dresden 2014 – wissenschaftliches Programm
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BP: Fachverband Biologische Physik
BP 1: Molecular Motors
BP 1.9: Vortrag
Montag, 31. März 2014, 12:00–12:15, HÜL 386
Single-molecule studies of RNA polymerase I elongation — •Ana Lisica1, Marcus Jahnel1, Christoph Engel2, Patrick Cramer2, and Stephan W. Grill1 — 1MPI-CBG and MPI-PKS, Dresden, Germany — 2Gene Centre Munich, LMU, Munich, Germany
Eukaryotic RNA polymerases (Pol I, Pol II and Pol III) have a highly conserved core and active center region. This implies that the mechanism of RNA polymerization is similar in all three polymerases. Additionally, Pol I and Pol III have specific surface subunits. Furthermore, the strength of the intrinsic activity of transcript cleavage differs highly: Pol I and Pol III possess a strong RNA cleavage mechanism, while Pol II has a weak one and needs a transcription factor (TFIIS) to cleave the RNA. To observe fine differences in transcription dynamics that stem from structural differences between these machines, we are using high-resolution dual-trap optical tweezers to analyze the mechanochemical details of the Pol I elongation in comparison to Pol II. Here we present the first single-molecule optical tweezers traces of Pol I transcribing a bare DNA template. A comparison with the Pol II dynamics revealed that Pol I is faster, with significantly higher overall and pause-free velocities, it pauses less often than Pol II, exhibits shorter pauses and can transcribe against higher opposing forces. Surprisingly, we find that the intrinsic transcript cleavage ability of Pol I is functional only with backtracked RNAs shorter than about 18 nt. Together, our results contribute to the understanding of unique micromechanical function and cellular role of this essential enzyme.