Project description:Early eukaryotic ribosome biogenesis involves large multi-protein complexes, which co-transcriptionally associate with pre-ribosomal RNA to form the small subunit processome. The precise mechanisms by which two of the largest multi-protein complexes – UtpA and UtpB – interact with nascent pre-ribosomal RNA have so far been poorly understood. We have combined biochemical and structural biology approaches with ensembles of RNA-protein cross-linking to elucidate the essential function of both complexes. Here we show that UtpA contains a large composite RNA binding site and captures the 5´ end of pre-ribosomal RNA. UtpB forms an extended structure that binds early pre-ribosomal intermediates in close proximity to key architectural sites such as an RNA duplex formed by the 5´ ETS and U3 snoRNA as well as the 3´ boundary of 18S rRNA. Both complexes therefore act as vital RNA chaperones to initiate eukaryotic ribosome assembly.
Project description:Farnesol is a quorum-sensing sesquiterpene alcohol that, via regulation of specific signalling and transcription components, inhibits filamentous growth in Candida. We have found that farnesol also inhibits translation at the initiation step. In contrast to fusel alcohols, that target the eukaryotic initiation factor 2B (eIF2B), farnesol affects the interaction of the mRNA with the small ribosomal subunit leading to reduced levels of the 48S pre-ribosomal complex. To establish the impact of farnesol and how effects on transcription and translation might be co-ordinated, we identified farnesol-dependent changes at both the transcript level and at the level of polysome association using next generation sequencing approaches.
Project description:Each mutant was cultured in duplicate, and from each culture two fractions were sequenced: Total fragmented RNA, and small ribosomal subunit footprints.
Project description:The human ribosomal protein S3 (RPS3), a component of the small 40S ribosomal subunit, is mainly involved in ribosomal maturation and initiation of translation through association with initiation factors. In this study, we firstly identified that RPS3 played an important role in HCC progression. We performed RNA sequencing to profile gene expression patterns before and after RPS3 knockdown.
Project description:The RNA helicase Has1 is involved in the biogenesis of both small and large ribosomal subunit. How it performs these separate roles is not fully understood. We analysed a detailed composition of RNA-protein complexes purified via Has1.
Project description:In eukaryotic cells, ribosomal RNA is transcribed by RNA polymerase I (Pol I). To initiate transcription, Pol I requires the assembly of a multi-subunit pre-initiation complex (PIC). In yeast, the minimal PIC includes Pol I, the transcription factor Rrn3 and Core Factor (CF) composed of subunits Rrn6, Rrn7 and Rrn11. We have determined the cryo-EM structure of a 18-subunit yeast Pol I complex together with CF, Rrn3 and transcription scaffold. To aid in modeling the structure based on the cryo-EM map, we performed crosslinking mass spectrometry.
