Nuclear quality control of pre-mRNA splicing

• Applicant: Professorin Dr. Heike Krebber
• Subject Area: General Genetics and Functional Genome Biology ; Cell Biology
• Term: from 2018 to 2024
• Project identifier: Deutsche Forschungsgemeinschaft (DFG) - Project number 394490813

Pre-mRNAs are generated and processed in the nucleus before they are transported to the cytoplasm for translation. During the life cycle of an mRNA many proteins associate and dissociate from the ribonucleoparticle (RNP) that have various functions. The shuttling serine/arginine (SR)-rich proteins Gbp2 and Hrb1 are recruited to the pre-mRNA during late phases of splicing and function as adapter proteins for the export receptor heterodimer Mex67-Mtr2, that allows transit of the mRNP through the nuclear pore complex (NPC) into the cytoplasm. Importantly, our work has shown that the SR-proteins function as quality control factors and prevent leakage of false or premature transcripts to the cytoplasm. Their exact function in RNA surveillance, however, is unclear and requires further studies. In this project we will study the mechanisms of quality control and determine the individual functions of Gbp2 and Hrb1 in this process through a combination of genetic and biochemical studies. With high throughput methods (e.g. mass spectrometry) we will identify the protein complexes in which Gbp2 in comparison to Hrb1 are mainly found. Additionally, we will use mutants of mRNA degradation factors, e.g. components of the TRAMP-complex that marks false mRNAs for degradation or nuclear exosome mutants and mRNA export mutants to enrich the SR-proteins in these complexes not only to study their interactions but also their potential modifications, which might provide switches for degradation or export. In the degradation mutants will also analyze to which mRNAs Gbp2 and Hrb1 bind by RIP-experiments and subsequent RNA-Seq analyses. In this way we will characterize their binding to defective mRNAs. In addition to that we will determine the responsible protein sequence for RNA-binding. Finally, we will investigate the last control step at the NPC that is mediated by Mlp1 and Mlp2. These studies will identify individual functions of Gbp2 and Hrb1 in nuclear quality control and illuminate the underlying mechanisms.

• DFG Programme: Research Grants