Abstract
ABSTRACTCajal bodies are nuclear structures that are involved in biogenesis of snRNPs and snoRNPs, maintenance of telomeres and processing of histone mRNA. Recently, the SUMO isopeptidase USPL1 was identified as a component of Cajal bodies that is essential for cellular growth and Cajal body integrity. However, a cellular function for USPL1 is so far unknown. Here, we use RNAi-mediated knockdown in human cells in combination with biochemical and fluorescence microscopy approaches to investigate the function of USPL1 and its link to Cajal bodies. We demonstrate that levels of snRNAs transcribed by RNA polymerase (RNAP) II are reduced upon knockdown of USPL1 and that downstream processes such as snRNP assembly and pre-mRNA splicing are compromised. Importantly, we find that USPL1 associates directly with U snRNA loci and that it interacts and colocalises with components of the Little Elongation Complex, which is involved in RNAPII-mediated snRNA transcription. Thus, our data indicate that USPL1 plays a key role in RNAPII-mediated snRNA transcription.
Highlights
The eukaryotic cell nucleus contains multiple compartments, or bodies, including the nucleolus, PML bodies, Cajal bodies and splicing speckles
Using a combination of fluorescence microscopy, molecular biology and proteomic approaches, we demonstrate that USPL1 interacts with components of the RNAPII-associated little elongation complex (LEC) and is associated with the U1 and U2 snRNA gene loci
Efficient knockdown of USPL1 by RNAi leads to reduced RNAPIImediated snRNA transcription, diminished production of snRNPs and altered pre-mRNA splicing
Summary
The eukaryotic cell nucleus contains multiple compartments, or bodies, including the nucleolus, PML bodies, Cajal bodies and splicing speckles (reviewed by Dundr and Misteli, 2010; Handwerger and Gall, 2006; Spector, 2001; Spector, 2006; Spector and Lamond, 2011; Zhao et al, 2009). Cajal bodies are dynamic structures that vary in size and number according to the cell cycle stage, differentiation and developmental status of the cell (reviewed by Cioce and Lamond, 2005; Machyna et al, 2013; Morris, 2008). They have been associated with roles in biogenesis of spliceosomal small nuclear (snRNPs) and small nucleolar RNPs (snoRNPs), maintenance of telomeres and histone mRNA processing. Exceptions are U6 and U6atac snRNAs, which are transcribed by RNAPIII and pass through the nucleolus for their maturation process (Ganot et al, 1999; Kunkel et al, 1986; Lange and Gerbi, 2000; Reddy et al, 1987; Tycowski et al, 1998)
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