A study of the nuclear trafficking of the splicing factor protein PRPF31 linked to autosomal dominant retinitis pigmentosa (ADRP)

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• 作者：Susan E. Wilkie ; Keith J. Morris ; Shomi S. Bhattacharya ; Martin J. Warren and David M. Hunt
• 刊名：Biochimica et Biophysica Acta (BBA)/Molecular Basis of Disease
• 出版年：2006
• 出版时间：March 2006
• 年：2006
• 卷：1762
• 期：3
• 页码：304-311
• 全文大小：293 K

文摘

In this study the mechanism of nuclear importation of the splicing factor PRPF31 is examined and the impact of two disease-linked mutations, A194E and A216P, assessed. Using pull-down assays with GST-tagged importin proteins, we demonstrate that His-tagged PRPF31 interacts with importin β1 for translocation to the nucleus, with no requirement for importin α1. The A194E and A216P mutations have no affect on this interaction. Fluorescence recovery after photobleaching (FRAP) was used to estimate the rate of movement of EGFP-tagged PRPF31 into the nuclei of live cells. The kinetics indicated a two-component recovery process; a fast component with τ  6 s and a slow component with τ  80 s. The mutations affected neither component. We conclude that the two mutations have no negative effect on interaction with the nuclear importation machinery. Reduced mutant protein solubility resulting in an insufficiency of splicing activity in cells with a very high metabolic demand remains the most likely explanation for the disease pathology in ADRP patients.