An Integrative Approach to Gain Insights into the Cellular Function of Human Ataxin-2

Abstract

Spinocerebellar ataxia type 2 (SCA2) is a hereditary neurodegenerative disorder caused by a trinucleotide expansion in the SCA2 gene, encoding a polyglutamine stretch in the gene product ataxin-2 (ATX2), whose cellular function is unknown. However, ATX2 interacts with A2BP1, a protein containing an RNA-recognition motif, and the existence of an interaction motif for the C-terminal domain of the poly(A)-binding protein (PABC) as well as an Lsm (Like Sm) domain in ATX2 suggest that ATX2 like its yeast homolog Pbp1 might be involved in RNA metabolism. Here, we show that, similar to Pbp1, ATX2 suppresses the petite (pet K ) phenotype of Dmrs2 yeast strains lacking mitochondrial group II introns. This finding points to a close functional relationship between the two homologs. To gain insight into potential functions of ATX2, we also generated a comprehensive protein interaction network for Pbp1 from publicly available databases, which implicates Pbp1 in diverse RNA-processing pathways. The functional relationship of ATX2 and Pbp1 is further corroborated by the experimental confirmation of the predicted interaction of ATX2 with the cytoplasmic poly(A)-binding protein 1 (PABP) using yeast-2-hybrid analysis as well as co-immunoprecipitation experiments. Immunofluorescence studies revealed that ATX2 and PABP co-localize in mammalian cells, remarkably, even under conditions in which PABP accumulates in distinct cytoplasmic foci representing sites of mRNA triage.

Ataxin-2 Interacts with the DEAD/H-Box RNA Helicase DDX6 and Interferes with P-Bodies and Stress Granules

“…Along this line, it has been reported that ATXN2 contains an Like Sm (LSm) domain that might be capable of RNA binding as observed for other LSm domain proteins (Neuwald and Koonin, 1998;Achsel et al, 2001;Albrecht et al, 2004). Further evidence for a likely role in RNA metabolism was provided by demonstrating that ATXN2 assembles with polysomes and interacts with the cytoplasmic poly(A)-binding protein 1 (PABP-C1) that functions in translation initiation and mRNA decay regulation (Ralser et al, 2005a;Satterfield and Pallanck, 2006). Proteomics-based results demonstrated that PABP and T-plastin, which itself associates with ATXN2, are present together in large protein complexes mostly consisting of proteins involved in RNA processing (Ho et al, 2002;Blagoev et al, 2003).…”

“…Thus, ATXN2 seems to be required for the assembly of SGs. Furthermore, we included in the course of the abovedescribed experiments the PABP protein as an additional component of SGs that is of particular interest, because of its interaction with ATXN2 (Ralser et al, 2005a). Strikingly, we observed that the PABP immunoreactivity was dramatically increased in all cells with diminished ATXN2 level ( Figure 8A).…”

“…Because ATXN2 and Pbp1 are functionally related as shown previously (Ralser et al, 2005a), we were questioning whether additional protein interactions identified in the yeast Pbp1 interaction network can be assigned to the human system. Here, we addressed the question whether the interaction between Pbp1 and Dhh1, which itself is a component of P-bodies and plays a major role in mRNA turnover and degradation (Coller et al, 2001;Fischer and Weis, 2002;Tseng-Rogenski et al, 2003), is evolutionary conserved in humans. …”

“…In Caenorhabditis elegans, the ATXN2 homologue is found in complexes with PAB1 and may be implicated in translational regulation (Ciosk et al, 2004). Additionally, the yeast homologue of ATXN2, named Pbp1 (Pab1-binding protein 1, also known as MRS16) is involved in important RNA-processing pathways such as RNA editing, pre-mRNA splicing, mRNA export, and degradation, which is clearly demonstrated by its comprehensive protein interaction network (Ralser et al, 2005a). Because ATXN2 and Pbp1 are functionally related as shown previously (Ralser et al, 2005a), we were questioning whether additional protein interactions identified in the yeast Pbp1 interaction network can be assigned to the human system.…”

“…The activity of the reporter genes was analyzed by monitoring the growth of the respective transformants after incubation of plates for 3 d at 30°C. The activity of ␤-galactosidase was determined as described previously (Ralser et al, 2005a). …”