Disease mechanism for retinitis pigmentosa (RP11) caused by missense mutations in the splicing factor gene PRPF31

Wilkie, Susan E. and Vaclavik, Veronika and Wu, Huimin and Bujakowska, Kinga and Chakarova, Christina F. and Bhattacharya, Shomi S. and Warren, Martin J. and Hunt, David M. (2008) Disease mechanism for retinitis pigmentosa (RP11) caused by missense mutations in the splicing factor gene PRPF31. Molecular Vision, 14 (81-83). pp. 683-690. ISSN 1090-0535. (The full text of this publication is not currently available from this repository. You may be able to access a copy if URLs are provided)

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Purpose: Missense mutations in the splicing factor gene PRPF31 cause a dominant form of retinitis pigmentosa (RP11) with reduced penetrance. Missense mutations in PRPF31 have previously been shown to cause reduced protein solubility, suggesting insufficiency of functional protein as the disease mechanism. Here we examine in further detail the effect of the A216P mutation on splicing function. Methods: Splicing activity was assayed using an in vivo assay in transfected mammalian cells with rhodopsin (RHO) and transducin (GNAT1) splicing templates. Pull-down assays were used to study the interaction between PRPF31 and one of its cognate partners in the spliceosome, PRPF6. Results: Splicing of RHO intron 3 and GNAT1 introns 3-5 mini-gene templates was inefficient with both spliced and unspliced products clearly detected. Assays using the RHO minigene template revealed a direct negative effect on splicing efficiency of the mutant. However, no effect of the mutation on splicing efficiency could be detected using the longer GNAT1 minigene template or using a full-length RHO transcript, splicing of which had an efficiency of 100%. No unspliced RHO transcripts could be detected in RNA from human retina. Pull-down assays between PRPF31 and PRPF6 proteins showed a stronger interaction for the mutant than wild type, suggesting a mechanism for the negative effect. Conclusions: Splicing of full-length RHO is more efficient than splicing of the minigene, and assays using a full-length template more accurately mimic splicing in photoreceptors. The RP11 missense mutations exert their pathology mainly via a mechanism based on protein insufficiency due to protein insolubility, but there is also a minor direct negative effect on function.

Item Type: Article
Subjects: Q Science > Q Science (General)
Divisions: Faculties > Science Technology and Medical Studies > School of Biosciences
Depositing User: Louise Dorman
Date Deposited: 19 Mar 2009 16:42
Last Modified: 17 Apr 2014 10:21
Resource URI: https://kar.kent.ac.uk/id/eprint/15329 (The current URI for this page, for reference purposes)
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