Crystal structure of a SFPQ/PSPC1 heterodimer provides insights into preferential heterodimerization of human DBHS family proteins

Jie Huang,G Patricia Casas Garcia,Matthew A Perugini,Archa H Fox,Charles S Bond,Mihwa Lee

doi:10.1074/jbc.ra117.001451

Jie Huang, G Patricia Casas Garcia + Show 4 more

Open Access

https://doi.org/10.1074/jbc.ra117.001451

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Abstract

Members of the Drosophila behavior human splicing (DBHS) protein family are nuclear proteins implicated in many layers of nuclear functions, including RNA biogenesis as well as DNA repair. Definitive of the DBHS protein family, the conserved DBHS domain provides a dimerization platform that is critical for the structural integrity and function of these proteins. The three human DBHS proteins, splicing factor proline- and glutamine-rich (SFPQ), paraspeckle component 1 (PSPC1), and non-POU domain-containing octamer-binding protein (NONO), form either homo- or heterodimers; however, the relative affinity and mechanistic details of preferential heterodimerization are yet to be deciphered. Here we report the crystal structure of a SFPQ/PSPC1 heterodimer to 2.3-Å resolution and analyzed the subtle structural differences between the SFPQ/PSPC1 heterodimer and the previously characterized SFPQ homodimer. Analytical ultracentrifugation to estimate the dimerization equilibrium of the SFPQ-containing dimers revealed that the SFPQ-containing dimers dissociate at low micromolar concentrations and that the heterodimers have higher affinities than the homodimer. Moreover, we observed that the apparent dissociation constant for the SFPQ/PSPC1 heterodimer was over 6-fold lower than that of the SFPQ/NONO heterodimer. We propose that these differences in dimerization affinity may represent a potential mechanism by which PSPC1 at a lower relative cellular abundance can outcompete NONO to heterodimerize with SFPQ.

Highlights

Members of the Drosophila behavior human splicing (DBHS) protein family are nuclear proteins implicated in many layers of nuclear functions, including RNA biogenesis as well as DNA repair
Defined by the highly conserved DBHS region that consists of two tandem RNA recognition motifs (RRMs), a NonA/paraspeckle domain (NOPS) and a C-terminal coiled-coil, the members of this protein family have been reported as multifunctional proteins, playing important roles in RNA biogenesis and transport, and subnuclear body formation via direction interaction with structural noncoding RNA as well as DNA damage repair [1]
Using the truncated constructs containing the dimerization domain, we have shown that the three human DBHS proteins can form homodimers (SFPQ [5], non-POU domain–containing octamer-binding protein (NONO) [10], and paraspeckle component 1 (PSPC1) (PDB code 5IFN) as well as heterodimers (NONO/PSPC1 [7] and SFPQ/NONO [5] in vitro

Summary

Introduction

Members of the Drosophila behavior human splicing (DBHS) protein family are nuclear proteins implicated in many layers of nuclear functions, including RNA biogenesis as well as DNA repair. The three human DBHS proteins, splicing factor proline- and glutamine-rich (SFPQ), paraspeckle component 1 (PSPC1), and non-POU domain–containing octamer-binding protein (NONO), form either homo- or heterodimers; the relative affinity and mechanistic details of preferential heterodimerization are yet to be deciphered. Tel.: 61-3-9479-3248; NONO, non-POU domain–containing octamer-binding protein; NOPS domain, NonA/paraspeckle domain; PSPC1, paraspeckle protein component 1; RRM, RNA recognition motif; SFPQ, splicing factor proline- and gluaspects of nuclear functions [1]. Defined by the highly conserved DBHS region that consists of two tandem RNA recognition motifs (RRMs), a NonA/paraspeckle domain (NOPS) and a C-terminal coiled-coil, the members of this protein family have been reported as multifunctional proteins, playing important roles in RNA biogenesis and transport, and subnuclear body (paraspeckle) formation via direction interaction with structural noncoding RNA as well as DNA damage repair [1]. DBHS proteins are obligatory dimers; the three human DBHS proteins form either homodimer or heterodimers among the three proteins in vitro [3,4,5] and all three possible heterodimers (SFPQ/NONO, SFPQ/ PSPC1, and PSPC1/NONO) have been shown in mouse Sertoli cells [6]

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