Recruitment of a splicing factor to the nuclear lamina for its inactivation

Abstract

Precursor messenger RNA splicing is a highly regulated process, mediated by a complex RNA-protein machinery, the spliceosome, that encompasses several hundred proteins and five small nuclear RNAs in humans. Emerging evidence suggests that the spatial organization of splicing factors and their spatio-temporal dynamics participate in the regulation of splicing. So far, methods to manipulate the spatial distribution of splicing factors in a temporally defined manner in living cells are missing. Here, we describe such an approach that takes advantage of a reversible chemical dimerizer, and outline the requirements for efficient, reversible re-localization of splicing factors to selected sub-nuclear compartments. In a proof-of-principle study, the partial re-localization of the PRPF38A protein to the nuclear lamina in HEK293T cells induced a moderate increase in intron retention. Our approach allows fast and reversible re-localization of splicing factors, has few side effects and can be applied to many splicing factors by fusion of a protein tag through genome engineering. Apart from the systematic analysis of the spatio-temporal aspects of splicing regulation, the approach has a large potential for the fast induction and reversal of splicing switches and can reveal mechanisms of splicing regulation in native nuclear environments.

Evolutionary, structural and functional insights in nuclear organisation and nucleocytoplasmic transport in trypanosomes

“…Beyond structural roles, in mammalian cells, lamins participate in controlling genome architecture, DNA replication and transcription by interactions with chromatin ( via lamin‐associated domains, LADs), chromatin‐modifying enzymes and transcription factors [ 37 , 38 ]. Lamins A/C can shuttle to the nucleoplasm where they bind and regulate euchromatic regions and interact with transcription and cell cycle regulators [ 39 ].…”