cDNA Library Screening Identifies Protein Interactors Potentially Involved in Non-Telomeric Roles of Arabidopsis Telomerase

Abstract

Telomerase-reverse transcriptase (TERT) plays an essential catalytic role in maintaining telomeres. However, in animal systems telomerase plays additional non-telomeric functional roles. We previously screened an Arabidopsis cDNA library for proteins that interact with the C-terminal extension (CTE) TERT domain and identified a nuclear-localized protein that contains an RNA recognition motif (RRM). This RRM-protein forms homodimers in both plants and yeast. Mutation of the gene encoding the RRM-protein had no detectable effect on plant growth and development, nor did it affect telomerase activity or telomere length in vivo, suggesting a non-telomeric role for TERT/RRM-protein complexes. The gene encoding the RRM-protein is highly expressed in leaf and reproductive tissues. We further screened an Arabidopsis cDNA library for proteins that interact with the RRM-protein and identified five interactors. These proteins are involved in numerous non-telomere-associated cellular activities. In plants, the RRM-protein, both alone and in a complex with its interactors, localizes to nuclear speckles. Transcriptional analyses in wild-type and rrm mutant plants, as well as transcriptional co-analyses, suggest that TERT, the RRM-protein, and the RRM-protein interactors may play important roles in non-telomeric cellular functions.

An armadillo-domain protein participates in a telomerase interaction network

“…4). These results also show that the presence of the telobox regulatory motif is not a critical determinant for specific gene expression, confirming results of previous studies (Dokladal et al 2015;Regad et al 1994;Schrumpfova et al 2016;Tremousaygue et al 2003;Zhou et al 2016). Secondly, we investigated genes involved in DNA repair pathways and found significant changes in expression of HR-related genes ( Fig.…”

“…Initially, we tested genes with telobox-containing promoters because binding to such genomic loci was demonstrated for SMH proteins (Schrumpfova et al 2016;Zhou et al 2016), and also because TRF-like family I proteins can bind telomeric repeats in vitro, similarly to SMH proteins (Karamysheva et al 2004;Schrumpfova et al 2004). Increased mRNA levels of ribosomal genes under the control of telobox-containing promoters were also observed in mutant lines with a disrupted RRM gene that encodes another AtTERT-interacting protein (Dokladal et al 2015). We observed significant changes in transcription of some (but not all) ribosomal genes under the control of teloboxcontaining promoters in arm-1 and arm-2 T-DNA insertion lines (Fig.…”

“…We and other groups (Dokladal et al 2015;Schrumpfova et al 2016;Zhou et al 2016) recently suggested that , c) or 7-dayold seedlings (d) using the ΔΔCt method and ubi10 as a reference gene. Relative transcript levels were calculated as a median of transcript levels; two-tailed P values were calculated using an unpaired t-test (*p < 0.05; **p < 0.001; ***p = 0.0001).…”

“…However, data from tert−/− mutants (Amiard et al 2014) showed no changes in transcript levels of ribosomal protein genes. To test a possible involvement of ARM protein in these processes, we measured transcript levels of selected representatives of ribosomal protein genes, that were upregulated in rrm mutants (Dokladal et al 2015), in arm mutant lines (Fig. 3c).…”

“…Our previous search for AtTERT partners identified two proteins from an Arabidopsis cDNA library that interacted with the CTE domain (Lee et al 2012). One of these was an RNA-recognitionmotif (RRM)-containing protein that also interacted with proteins responsible for transcriptional and post-transcriptional regulation (Dokladal et al 2015). The second protein of unknown function was an armadillo/β-catenin-like repeat (ARM, encoded by AT4G33945) protein that interacted with the CTE domain of AtTERT, predominantly in the cytoplasm of BY-2 protoplasts (Lee et al 2012).…”