New papers from Professor Baralle's lab

Please, have a look at the 3 new papers from Professor Baralle's lab:

Dhir, A., Buratti, E. Alternative splicing: role of pseudoexons in human disease and potential therapeutic strategies. 2010. FEBS J. 277, 841-855


Abstract: What makes a nucleotide sequence an exon (or an intron) is a question that still lacks a satisfactory answer. Indeed, most eukaryotic genes are full of sequences that look like perfect exons, but which are nonetheless ignored by the splicing machinery (hence the name ‘pseudoexons’). The existence of these pseudoexons has been known since the earliest days of splicing research, but until recently the tendency has been to view them as an interesting, but rather rare, curiosity. In recent years, however, the importance of pseudoexons in regulating splicing processes has been steadily revalued. Even more importantly, clinically oriented screening studies that search for splicing mutations are beginning to uncover a situation where aberrant pseudoexon inclusion as a cause of human disease is more frequent than previously thought. Here we aim to provide a review of the mechanisms that lead to pseudoexon activation in human genes and how the various cis- and trans-acting cellular factors regulate their inclusion. Moreover, we list the potential therapeutic approaches that are being tested with the aim of inhibiting their inclusion in the final mRNA molecules.

Dhir, A., Buratti, E., van Santen, M.A., Lührmann, R., Baralle, F.E. The intronic splicing code: multiple factors involved in ATM pseudoexon definition. 2010. EMBO J. 29, 749-760


Abstract: Abundance of pseudo splice sites in introns can potentially give rise to innumerable pseudoexons, outnumbering the real ones. Nonetheless, these are efficiently ignored by the splicing machinery, a process yet to be understood completely. Although numerous 5’ splice site-like sequences functioning as splicing silencers have been found to be enriched in predicted human pseudoexons, the lack of active pseudoexons pose a fundamental challenge to how these U1snRNP-binding sites function in splicing inhibition. Here, we address this issue by focusing on a previously described pathological ATM pseudoexon whose inhibition is mediated by U1snRNP binding at intronic splicing processing element (ISPE), composed of a consensus donor splice site. Spliceosomal complex assembly demonstrates inefficient A complex formation when ISPE is intact, implying U1snRNP-mediated unproductive U2snRNP recruitment. Furthermore, interaction of SF2/ASF with its motif seems to be dependent on RNA structure and U1snRNP interaction. Our results suggest a complex combinatorial interplay of RNA structure and trans-acting factors in determining the splicing outcome and contribute to understanding the intronic splicing code for the ATM pseudoexon.

Haque, A., Buratti, E., Baralle, F.E. Functional properties and evolutionary splicing constraints on a composite exonic regulatory element of splicing in CFTR exon 12. 2010. Nucleic Acids Res. 38(2), 647-659


Abstract: In general, splicing regulatory elements are defined as Enhancers or Silencers depending on their positive or negative effect upon exon inclusion. Often, these sequences are usually present separate from each other in exonic/intronic sequences. The Composite Exonic Splicing Regulatory Elements (CERES) represent an extreme physical overlap of enhancer/silencer activity. As a result, when CERES elements are mutated the consequences on the splicing process are difficult to predict. Here, we show that the functional activity of the CERES2 sequence in CFTR exon 12 is regulated by the binding, in very close proximity to each other, of several SR and hnRNP proteins. Moreover, our results show that practically the entire exon 12 sequence context participate in its definition. The consequences of this situation can be observed at the evolutionary level by comparing changes in conservation of different splicing elements in different species. In conclusion, our study highlights how it is increasingly difficult to define many exonic sequences by simply breaking them down in isolated enhancer/silencer or even neutral elements. The real picture is close to one of continuous competition between positive and negative factors where affinity for the target sequences and other dynamic factors decide the inclusion or exclusion of the exon.