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RNA export factor Ddx19 is required for nuclear import of the SRF coactivator MKL1.

ABSTRACT: Controlled transport of macromolecules between the cytoplasm and nucleus is essential for homeostatic regulation of cellular functions. For instance, gene expression entails coordinated nuclear import of transcriptional regulators to activate transcription and nuclear export of the resulting messenger RNAs for cytoplasmic translation. Here we link these two processes by reporting a novel role for the mRNA export factor Ddx19/Dbp5 in nuclear import of MKL1, the signal-responsive transcriptional activator of SRF. We show that Ddx19 is not a general nuclear import factor, and that its specific effect on MKL1 nuclear import is separate from its role in mRNA export. Both helicase and nuclear pore-binding activities of Ddx19 are dispensable for MKL1 nuclear import, but RNA binding is required. Mechanistically, Ddx19 operates by modulating the conformation of MKL1, which affects its interaction with Importin-? for efficient nuclear import. Thus, Ddx19 participates in mRNA export, translation and nuclear import of a key transcriptional regulator.

SUBMITTER: Rajakyla EK

PROVIDER: S-EPMC4309436 | biostudies-literature | 2015 Jan

REPOSITORIES: biostudies-literature

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RNA export factor Ddx19 is required for nuclear import of the SRF coactivator MKL1.

Rajakylä Eeva Kaisa EK Viita Tiina T Kyheröinen Salla S Huet Guillaume G Treisman Richard R Vartiainen Maria K MK

Nature communications 20150114

Controlled transport of macromolecules between the cytoplasm and nucleus is essential for homeostatic regulation of cellular functions. For instance, gene expression entails coordinated nuclear import of transcriptional regulators to activate transcription and nuclear export of the resulting messenger RNAs for cytoplasmic translation. Here we link these two processes by reporting a novel role for the mRNA export factor Ddx19/Dbp5 in nuclear import of MKL1, the signal-responsive transcriptional a ...[more]

PMID: 25585691

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Project description:BACKGROUND: Chikungunya virus (CHIKV) is an alphavirus of the Togaviridae family. After autoproteolytic cleavage, the CHIKV capsid protein (CP) is involved in RNA binding and assembly of the viral particle. The monomeric CP is approximately 30 kDa in size and is small enough for passive transport through nuclear pores. Some alphaviruses are found to harbor nuclear localization signals (NLS) and transport of these proteins between cellular compartments was shown to be energy dependent. The active nuclear import of cytoplasmic proteins is mediated by karyopherins and their export by exportins. As nuclear and cytoplasmic trafficking may play a role in the life cycle of CHIKV, we have sought to identify nuclear localization and nuclear export signals in CHIKV CP in a virus-free system. METHODS: EGFP-fusion proteins of CHIKV CP and mutants thereof were created and used to monitor their intracellular localization. Binding of cellular proteins was confirmed in pull-down assays with purified CP using co-immuoprecipitation. Nuclear localization was demonstrated in a virus-free system using fluorescence microscopy. RESULTS: Here we show that CHIKV CP is a nuclear-cytoplasmic shuttling protein with an active NLS that binds to karyopherin ? (Kar?) for its nuclear translocation. We also found that the Kar?4 C-terminal NLS binding site is sufficient for this interaction. We further demonstrate that CHIKV CP interacts directly with the export receptor CRM1 to transport this viral protein out of the nucleus via a nuclear export signal (NES). The CHIKV CP NES was mapped between amino acids 143 and 155 of CP. Deduced from in silico analyses we found that the NES has a mode of binding similar to the snurportin-1 CRM1 complex. CONCLUSIONS: We were able to show that in a virus-free system that the CHIKV capsid protein contains both, a NLS and a NES, and that it is actively transported between the cytoplasma and the nucleus. We conclude that CHIKV CP has the ability to shuttle via interaction with karyopherins for its nuclear import and, vice versa, by CRM1-dependent nuclear export.

Dataset Information

RNA export factor Ddx19 is required for nuclear import of the SRF coactivator MKL1.

Publications

RNA export factor Ddx19 is required for nuclear import of the SRF coactivator MKL1.

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