Regulation of Neuronal Differentiation by Proteins Associated with Nuclear Bodies

Benjamin Förthmann,Michal K Stachowiak,Michael Hebert,Claudia Grothe,Hella Brinkmann,Peter Claus,Yvonne Schill,Verena Lübben,Yu-Wei Lee,Andreas Ratzka,Jeroen Van Bergeijk,Ferdinando Di Cunto

doi:10.1371/journal.pone.0082871

Abstract

Nuclear bodies are large sub-nuclear structures composed of RNA and protein molecules. The Survival of Motor Neuron (SMN) protein localizes to Cajal bodies (CBs) and nuclear gems. Diminished cellular concentration of SMN is associated with the neurodegenerative disease Spinal Muscular Atrophy (SMA). How nuclear body architecture and its structural components influence neuronal differentiation remains elusive. In this study, we analyzed the effects of SMN and two of its interaction partners in cellular models of neuronal differentiation. The nuclear 23 kDa isoform of Fibroblast Growth Factor – 2 (FGF-223) is one of these interacting proteins – and was previously observed to influence nuclear bodies by destabilizing nuclear gems and mobilizing SMN from Cajal bodies (CBs). Here we demonstrate that FGF-223 blocks SMN-promoted neurite outgrowth, and also show that SMN disrupts FGF-223-dependent transcription. Our results indicate that FGF-223 and SMN form an inactive complex that interferes with neuronal differentiation by mutually antagonizing nuclear functions. Coilin is another nuclear SMN binding partner and a marker protein for Cajal bodies (CBs). In addition, coilin is essential for CB function in maturation of small nuclear ribonucleoprotein particles (snRNPs). The role of coilin outside of Cajal bodies and its putative impacts in tissue differentiation are poorly defined. The present study shows that protein levels of nucleoplasmic coilin outside of CBs decrease during neuronal differentiation. Overexpression of coilin has an inhibitory effect on neurite outgrowth. Furthermore, we find that nucleoplasmic coilin inhibits neurite outgrowth independent of SMN binding revealing a new function for coilin in neuronal differentiation.

Highlights

The nucleus comprises a number of subnuclear structures important for regulation of cellular functions
We found that Survival of Motor Neuron (SMN)-enhanced neurite outgrowth as well as FGF-223-promoted transcription was inhibited by FGF-223/SMN complex formation
Interaction of SMN with FGF-223 negatively regulates neurite outgrowth How does FGF-223 affect the function of SMN in neuronal differentiation? To address this question we analyzed the effects of FGF-223 and SMN in a neurite outgrowth model using PC12 cells

Summary

Introduction

The nucleus comprises a number of subnuclear structures important for regulation of cellular functions. Several proteins are organized in endogenous and physiological aggregates which are called nuclear bodies [1,2]. One of these structures was described as the ‘‘nucleolar accessory body’’ because of its closeness to the nucleolus and later named the Cajal body (CB) [3,4]. FGF-2 is an extracellular factor binding to receptor tyrosine kinase receptors and expressed as different isoforms with intracellular functions [8]. One of these isoforms, nuclear FGF-223 has been shown to bind directly to SMN whereas 18 kDa FGF-2 (FGF-218) does not [9,10]. The FGFR1/FGF-223 complex promotes neuronal differentiation by TH-gene activation in a pathway referred to as integrative nuclear FGFR1 signaling (INFS) [13,14,15]

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Results

Conclusion