A5031110778- RNA Research and Splicing
- RNA and protein synthesis mechanisms
- Bacterial Genetics and Biotechnology
- RNA modifications and cancer
- CRISPR and Genetic Engineering
- Genomics and Chromatin Dynamics
- DNA Repair Mechanisms
- Gene Regulatory Network Analysis
- Fungal and yeast genetics research
- Bacteriophages and microbial interactions
- Plant Disease Resistance and Genetics
- Advanced biosensing and bioanalysis techniques
- DNA and Nucleic Acid Chemistry
- Molecular Biology Techniques and Applications
- RNA Interference and Gene Delivery
- Neurogenetic and Muscular Disorders Research
University of Turku
2012-2024
Aarhus University
2018-2021
University of Wisconsin–Madison
2016
Åbo Akademi University
2012
Multisubunit RNA polymerase (RNAP) is the central information-processing enzyme in all cellular life forms, yet its mechanism of translocation along DNA molecule remains conjectural. Here, we report direct monitoring bacterial RNAP following addition a single nucleotide. Time-resolved measurements demonstrated that delayed relative to nucleotide incorporation and occurs shortly after or concurrently with pyrophosphate release. An investigation equilibrium suggested strength interactions...
Universally conserved factors from NusG family bind at the upstream fork junction of transcription elongation complexes and modulate RNA synthesis in response to translation, processing, folding nascent RNA. Escherichia coli enhances vitro by a poorly understood mechanism. Here we report that E. slows Gre factor-stimulated cleavage RNA, but does not measurably change rates single nucleotide addition translocation non-paused polymerase. We demonstrate inhibiting backtracking. This activity is...
Poly(A) tails of newly synthesized mRNAs have uniform lengths, arising through cooperation between the cleavage and polyadenylation complex (CPAC) poly(A) binding proteins (PABPs). In budding yeast, Saccharomyces cerevisiae , responsible PABP is evolutionarily conserved CCCH zinc finger protein Nab2 that facilitates biogenesis ~60 adenosine mRNA tails. Here, we address molecular basis for such length control. Reconstituting reactions during formation Nab2:poly(A) RNA ribonucleoprotein...
Bacterial RNA polymerase (RNAP) is a validated target for antibacterial drugs. CBR703 series antimicrobials allosterically inhibit transcription by binding to conserved α helix (β′ bridge helix, BH) that interconnects the two largest RNAP subunits. Here we show disruption of BH-β subunit contacts amino-acid substitutions invariably results in accelerated catalysis, slowed-down forward translocation and insensitivity regulatory pauses. partially reverses these effects CBR-resistant RNAPs...
RNA cleavage by bacterial polymerase (RNAP) has been implicated in transcriptional proofreading and reactivation of arrested transcription elongation complexes but its molecular mechanism is less understood than the nucleotide addition, despite both reactions taking place same active site. RNAP from radioresistant bacterium Deinococcus radiodurans characterized highly efficient intrinsic comparison with Escherichia coli RNAP. We find that enhanced activity largely derives amino acid...
Cell-free environments are becoming viable alternatives for implementing biological networks in synthetic biology. The reconstituted cell-free expression system (PURE) allows characterization of genetic under defined conditions but its applicability to native bacterial promoters and endogenous is limited due the poor transcription rate Escherichia coli RNA polymerase this minimal system. We found that addition elongation factors GreA GreB PURE increased rates E. from sigma factor 70 up...
Biogenesis of most eukaryotic mRNAs involves the addition an untemplated polyadenosine (pA) tail by cleavage and polyadenylation machinery. The pA tail, its exact length, impacts mRNA stability, nuclear export, translation. To define how is controlled in S. cerevisiae , we have used vivo assay capable assessing synthesis, analyzed length distributions direct RNA sequencing, reconstituted reactions with purified components. This revealed three control mechanisms for length. First, found that...
All cellular RNA polymerases (RNAP) occasionally backtrack along the template DNA as part of transcriptional proofreading and regulation. Here, we studied mechanism RNAP backtracking by one nucleotide using two complementary approaches that allowed us to precisely measure occupancy lifetime backtracked state. Our data show stability state is critically dependent on closure active site a mobile domain, trigger loop (TL). The measurably decreased substitutions TL residues interact with...
ABSTRACT All cellular RNA polymerases (RNAP) occasionally backtrack along the template DNA as part of transcriptional proofreading and regulation. Here, we studied mechanism RNAP backtracking by one nucleotide using two complementary approaches that allowed us to precisely measure occupancy lifetime backtracked state. Our data show stability state is critically dependent on closure active site a mobile domain, trigger loop (TL). The measurably decreased substitutions TL residues interact...
DNA lesions can severely compromise transcription and block RNA synthesis by polymerase (RNAP), leading to subsequent recruitment of repair factors the stalled complex. Recent structural studies have uncovered molecular interactions several within elongation However, little is known about role key elements RNAP active site in translesion transcription. Here, using recombinantly expressed proteins, vitro transcription, kinetic analyses, vivo cell viability assays, we report that point amino...
Cell-free environments are becoming viable alternatives for implementing biological networks in synthetic biology. The reconstituted cell-free expression system (PURE) allows characterization of genetic under defined conditions but its applicability to native bacterial promoters and endogenous is limited due the poor transcription rate Escherichia coli RNA polymerase this minimal system. We found that addition elongation factors GreA GreB PURE increased rates E. from sigma factor 70 up...
Abstract The polyadenosine tail (pA-tail) regulates mRNA nuclear export, stability, and translatability. Based on reporter constructs, the prevailing model suggests that pA-tail removal mediated by Ccr4-NOT or PAN2/3 deadenylases is required for decapping degradation. Here, we use direct RNA sequencing to track deadenylation decay at steady-state in stress conditions show a global correlation between decay. Interestingly, codon optimality, previously postulated dictate only strongly affects...