Ricerc@Sapienza

Spinal muscular atrophy (SMA) is a neurodegenerative disease caused by a deficiency of the Survival Motor Neuron (SMN) protein. SMN, the causative factor of SMA, has a crucial role in snRNPs biogenesis as cells defective for SMN result in aberrantly spliced transcripts. SMN is strickly linked to TGS1, an S-adenosyl-L-methionine dependent methyltransferase responsible for snRNAs cap trimethylation. Our recent work shows a physical and genetic interaction between TGS1 and all the subunits of Drosophila Smn complex. Loss of dTgs1 results in neurodegenerative phenotypes similar to those caused by Smn loss in fly eye imaginal discs, indicating that both dTgs1 and dSmn are required for viability of retinal progenitor cells (1). Loss of TGS1 leads to accumulation of immature snRNA with extended 3' tail both in Drosophila melanogaster and HeLa cells systems. In line with the accumulation of defective snRNAs, sequencing analyses performed on TGS1 and SMN mutant cells reveal a partial overlapping of transcriptome alterations such as unspliced and readthrough transcripts, probably due to incorrect transcription termination. We aim at understanding how 3' extended snRNAs and defective transcripts contribute to neurodegeneration in SMA. Integrating Illumina RNA sequencing with Oxford nanopore technologies, we plan to perform an analysis on Drosophila neuronal tissues like brains and eye imaginal discs as well as HeLa cells in order to gain insight into transcriptome alterations linked to loss of TGS1 and SMN. The power of using long read nanopore sequencing allows the reconstruction of full-length RNA molecules so as to circumvent the ambiguity generated by imprecise attribution of RNA-seq reads to a specific isoform.