Heterogeneous nuclear ribonucleoproteins (hnRNPs) represent one of the most abundant classes of RNPs. They are involved in many crucial aspects of the control of gene expression. In particular, hnRNP type I (PTB) is known as a negative regulator of RNA processing, in the role of repressor of tissue-specific exons. The research plan focuses on the study of the role of hnRNP PTB, nPTB and Raver ribonucleproteins in the regulation of gene expression in muscular tissue. This program represents the natural follow up of a long sought and well documented interest of the research unit towards PTB and Raver proteins, which interact with PTB as co-repressors.
We plan to investigate their roles in the regulating alternative splicing of muscle-specific transcripts. The embrionic murine cell line C2C12 will be used as a model system for skeletal muscle differentiation. Various splicing variants of the cited hnRNPs will be characterized and quantified, at different differentiation stages of C2C12 cells in colture. We will then investigate on their role as repressors of tissue specific exons in transcripts from myogenic factors, such as MEF2 and from PKM2 and AXNA7 genes. Alternative exons of PKM2 and AXNA7 are in fact preferentially expressed in skeletal muscle. The involvement of PTB proteins in MEF2, PKM2 and AXNA7 genes alternative splicing will be tested by gene silencing (i.e.RNA interference) assays. Finally, we plan to complete the analysis of Raver genes controlling regions, by characterizing Raver 2 gene promoter.
