UG Repeats/TDP-43 Interactions near 5′ Splice Sites Exert Unpredictable Effects on Splicing Modulation

Abstract

TDP-43 is a nuclear protein implicated in the pathogenesis of several neurodegenerative diseases such as amyotrophic lateral sclerosis and frontotemporal lobar degeneration, with broad involvement in numerous stages of RNA processing ranging from transcription to translation. In diseased neurons, TDP-43 mostly aggregates in the cytoplasm, suggesting that a loss of protein function in the nucleus may play an important role in neurodegeneration. A better understanding of TDP-43 general nuclear functions is therefore an essential step to evaluate this possibility. Presently, the TDP-43 best-characterized functional property is its ability to modulate pre-mRNA splicing when binding in proximity of 3′SS acceptor sequences. In this work, using a variety of artificial and natural splicing substrates, we have investigated the effects of TDP-43 binding to UG repeats in the vicinity of 5′SS donor sequences. In general, our results show that UG repeats are not powerful splicing regulatory elements when located near to exonic 5′SS sequences. However, in cases like the BRCA1, ETF1, and RXRG genes, TDP-43 binding to natural UG-repeated sequences can act as either an activator or a suppressor of 5′SS recognition, depending on splice site strength and on the presence of additional splicing regulatory sequences. The results of this analysis suggest that a role of UG repeats/TDP-43 in 5′SS recognition may exists and may become critical in the presence of mutations that weaken the 5′SS. The general rule that can be drawn at the moment is that the importance of UG repeats near 5′ splice sites should always be experimentally validated on a case-by-case basis.