Completing the TRB family: newly characterized members show ancient evolutionary origins and distinct localization, yet similar interactions

Abstract

Telomere repeat binding proteins (TRBs) belong to a family of proteins possessing a Myb-like domain which binds to telomeric repeats. Three members of this family (TRB1, TRB2, TRB3) from Arabidopsis thaliana have already been described as associated with terminal telomeric repeats (telomeres) or short interstitial telomeric repeats in gene promoters (telo-boxes). They are also known to interact with several protein complexes: telomerase, Polycomb repressive complex 2 (PRC2) E(z) subunits and the PEAT complex (PWOs-EPCRs-ARIDs-TRBs). Here we characterize two novel members of the TRB family (TRB4 and TRB5). Our wide phylogenetic analyses have shown that TRB proteins evolved in the plant kingdom after the transition to a terrestrial habitat in Streptophyta, and consequently TRBs diversified in seed plants. TRB4-5 share common TRB motifs while differing in several others and seem to have an earlier phylogenetic origin than TRB1-3. Their common Myb-like domains bind long arrays of telomeric repeats in vitro, and we have determined the minimal recognition motif of all TRBs as one telo-box. Our data indicate that despite the distinct localization patterns of TRB1-3 and TRB4-5 in situ, all members of TRB family mutually interact and also bind to telomerase/PRC2/PEAT complexes. Additionally, we have detected novel interactions between TRB4-5 and EMF2 and VRN2, which are Su(z)12 subunits of PRC2.

Distinct clades of TELOMERE REPEAT BINDING transcriptional regulators interplay to regulate plant development

“…To test this, we carried out yeasttwo-hybrid (Y2H) experiments using each protein either as bait or as prey. Our assay confirmed known interactions among clade I and, in agreement with (Kusová et al, 2023)…”

“…The three proteins localize throughout euchromatin, sometimes as small speckles, but are depleted from the DAPI-bright heterochromatic chromocenters (Figure 4D). TRB4 and TRB5 as well as TRB1 are also detected in the nucleolus as was already observed after transient expression of TRBs in tobacco leaves (Zhou et al, 2016;Kusová et al, 2023). To obtain a precise view on the genomic distribution of a clade II TRB proteins, we carried out ChIP-seq targeting TRB4-GFP in 7-day old plantlets and identified more than 5000 TRB4 peaks robustly detected in two independent biological replicates (Supplementary Figure 4A).…”

“…In addition to TRB1, TRB2 and TRB3, data reanalysis identified two poorly characterized members of the TRB family, namely TRB4 and TRB5, which are significantly enriched (Fold Change (FC) of 2 and 7.3 respectively) in the telomere pull-down compared to the shuffled DNA control (Figure 1A). Previous studies suggested that TRB4 and TRB5 belong to a separate clade in the TRB phylogeny (Kotlinski et al, 2017;Kusová et al, 2023). To time the appearance of this clade, we selected 24 species to represent the diversity of the green land plant lineage and interrogated several databases using Arabidopsis TRB1 to TRB5 as query.…”

“…Given the ability of TRB1 and TRB4 proteins to from hetero-dimers ( (Kusová et al, 2023), Figure 3A-B) and the faculty of TRB1 and TRB4 to bind similar consensus sequences, we searched for potential TRB1 and TRB4 co-occurrence. In addition to TRB1-and TRB4-specific target sites, TRB4 shares more than half of its binding sites with TRB1 (Figure 5E -F).…”

Cytokinins – regulators of de novo shoot organogenesis

“…In this inducible state, WUS can be activated by type-B ARRs, for instance ARR10, directly targeting WUS (Zubo et al, 2017). The deposition of H3K27me3 by PRC2 (Zhou et al, 2018a) may be coordinated also by Telomeric repeat binding (TRB) proteins (Schrumpfováet al, 2016;Kusováet al, 2023). TRB proteins also recruit H3K4 demethylase JMJ14 implicated in the regulation of WUS expression (Li et al, 2011;Wang et al, 2023b).…”