In Vitro RNase and Nucleic Acid Binding Activities Implicate Coilin in U snRNA Processing

Hanna J Broome,Michael D Hebert,Alessandro Marcello

doi:10.1371/journal.pone.0036300

Hanna J Broome, Michael D Hebert + Show 1 more

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https://doi.org/10.1371/journal.pone.0036300

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Journal: PloS one	Publication Date: Apr 27, 2012
Citations: 54	License type: CC BY 4.0

Affiliation: University of Mississippi Medical Center

Abstract

Coilin is known as the marker protein for Cajal bodies (CBs), subnuclear domains important for the biogenesis of small nuclear ribonucleoproteins (snRNPs) which function in pre-mRNA splicing. CBs associate non-randomly with U1 and U2 gene loci, which produce the small nuclear RNA (snRNA) component of the respective snRNP. Despite recognition as the CB marker protein, coilin is primarily nucleoplasmic, and the function of this fraction is not fully characterized. Here we show that coilin binds double stranded DNA and has RNase activity in vitro. U1 and U2 snRNAs undergo a processing event of the primary transcript prior to incorporation in the snRNP. We find that coilin displays RNase activity within the CU region of the U2 snRNA primary transcript in vitro, and that coilin knockdown results in accumulation of the 3′ pre-processed U1 and U2 snRNA. These findings present new characteristics of coilin in vitro, and suggest additional functions of the protein in vivo.

Highlights

The Cajal body (CB) is a subnuclear structure especially prominent in cells with relatively high transcription demands, such as neuronal and cancer cells
CBs contain the survival of motor neuron (SMN) protein, which plays an essential role in the assembly of Sm proteins onto small nuclear RNA during spliceosomal small nuclear ribonucleoprotein biogenesis [1,2]
Since the majority of coilin is not located in the CB, but is found in the nucleoplasm [39], effort should be made to determine if nucleoplasmic coilin possess activities related to or independent from CB function

Summary

Introduction

The Cajal body (CB) is a subnuclear structure especially prominent in cells with relatively high transcription demands, such as neuronal and cancer cells. CBs contain the survival of motor neuron (SMN) protein, which plays an essential role in the assembly of Sm proteins onto small nuclear RNA (snRNA) during spliceosomal small nuclear ribonucleoprotein (snRNP) biogenesis [1,2]. It is thought that this happens co-transcriptionally via interactions between the phosphorylated C-terminal domain (CTD) of RNA polymerase II and the Integrator complex [5,6,7]. This processing event results in the formation of a presnRNA that is later trimmed to the fully processed U2 snRNA during reactions occurring in the cytoplasm, followed by import to the nucleus and CB for final snRNP maturation steps. Studies using artificial tandem arrays have shown a clear requirement for active U2 snRNA transcription in the association of CBs with these arrays [9,10]

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