RNA-binding protein PSPC1 promotes the differentiation-dependent nuclear export of adipocyte RNAs
Jiexin Wang, Prashant Rajbhandari, Andrey Damianov, Areum Han, Tamer Sallam, Hironori Waki, Claudio J. Villanueva, Stephen D. Lee, Ronni Nielsen, Susanne Mandrup, Karen Reue, Stephen G. Young, Julian Whitelegge, Enrique Saez, Douglas L. Black, Peter Tontonoz
Jiexin Wang, Prashant Rajbhandari, Andrey Damianov, Areum Han, Tamer Sallam, Hironori Waki, Claudio J. Villanueva, Stephen D. Lee, Ronni Nielsen, Susanne Mandrup, Karen Reue, Stephen G. Young, Julian Whitelegge, Enrique Saez, Douglas L. Black, Peter Tontonoz
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Research Article Metabolism

RNA-binding protein PSPC1 promotes the differentiation-dependent nuclear export of adipocyte RNAs

Abstract

A highly orchestrated gene expression program establishes the properties that define mature adipocytes, but the contribution of posttranscriptional factors to the adipocyte phenotype is poorly understood. Here we have shown that the RNA-binding protein PSPC1, a component of the paraspeckle complex, promotes adipogenesis in vitro and is important for mature adipocyte function in vivo. Cross-linking and immunoprecipitation followed by RNA sequencing revealed that PSPC1 binds to intronic and 3′-untranslated regions of a number of adipocyte RNAs, including the RNA encoding the transcriptional regulator EBF1. Purification of the paraspeckle complex from adipocytes further showed that PSPC1 associates with the RNA export factor DDX3X in a differentiation-dependent manner. Remarkably, PSPC1 relocates from the nucleus to the cytoplasm during differentiation, coinciding with enhanced export of adipogenic RNAs. Mice lacking PSPC1 in fat displayed reduced lipid storage and adipose tissue mass and were resistant to diet-induced obesity and insulin resistance due to a compensatory increase in energy expenditure. These findings highlight a role for PSPC1-dependent RNA maturation in the posttranscriptional control of adipose development and function.

Authors

Jiexin Wang, Prashant Rajbhandari, Andrey Damianov, Areum Han, Tamer Sallam, Hironori Waki, Claudio J. Villanueva, Stephen D. Lee, Ronni Nielsen, Susanne Mandrup, Karen Reue, Stephen G. Young, Julian Whitelegge, Enrique Saez, Douglas L. Black, Peter Tontonoz

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Figure 1

PSPC1 expression promotes adipogenesis.

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PSPC1 expression promotes adipogenesis. (A) Real-time PCR analysis of Ps...
(A) Real-time PCR analysis of Pspc1 mRNA during the differentiation of 10T1/2 and 3T3-L1 preadipocytes. Cells were stimulated to differentiate with dexamethasone (1 μM), IBMX (0.5 mM), insulin (5 μg/ml), and GW1929 (20 nM) for 2 days followed by insulin and GW1929 for 5 days. Comparison was made against D0 by 1-way ANOVA. Results represent 4 independent experiments. Unless mentioned otherwise, mRNA expression in this and all subsequent figures was normalized to 36B4 control. (B) Immunoblot analysis of PSPC1 protein during differentiation of 10T1/2 and 3T3-L1 preadipocytes. Cells were treated as in A. Results are representative of 3 independent experiments. (C) Real-time PCR analysis of Pspc1, Pparg, and adiponectin mRNA in epididymal white adipose tissue from control ob/ob mice and WT C57BL/6 mice fed a high-fat diet (HFD) for 12 weeks. n = 8–10 per group. Statistical analysis was performed using Student’s t test. (D) Immunoblot analysis of PSPC1 protein expression in retrovirally derived 10T1/2 stable cell lines expressing vector or PSPC1. (E) Oil Red O staining of 10T1/2 stable cells described in D. Cells were stimulated to differentiate with DMI + 20 nM GW for 8 days. (F) Real-time PCR analysis of adipogenic gene expression in 10T1/2 stable cells described in D on different days during differentiation. Cells were stimulated to differentiate as in E. Comparison was made against vector control by Student’s t test. Results represent 3 independent experiments. (G) Immunoblot analysis of PSPC1 protein expression in retrovirally derived 10T1/2 stable cell lines expressing PSPC1 shRNAs (shPspc1, sh#1, and sh#2) or lacZ shRNA (shLacZ) control. (H) Oil Red O staining of the 10T1/2 stable cells described in G. Cells were stimulated to differentiate with DMI + 20 nM GW for 7 days. (I) Real-time PCR analysis of adipogenic gene expression in 10T1/2 stable cells described in G on different days during differentiation. Cells were stimulated to differentiate as in H. Comparison was made against shLacZ control by 1-way ANOVA. Results are representative of 3 independent experiments. Error bars represent mean + SEM. *P < 0.05, **P < 0.01.