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Key message Understanding the molecular network, including protein-protein interactions, of VRS5 provide new routes towards the identification of other key regulators of plant architecture in barley. The TCP transcriptional regulator TEOSINTE BRANCHED 1 (TB1) is a key regulator of plant architecture. In barley, an important cereal crop, HvTB1 (also referred to as VULGARE SIX-ROWED spike (VRS) 5), inhibits the outgrowth of side shoots, or tillers, and grains. Despite its key role in barley development, there is limited knowledge on the molecular network that is utilized by VRS5. In this work, we performed protein–protein interaction studies of VRS5. Our analysis shows that VRS5 potentially interacts with a diverse set of proteins, including other class II TCP’s, NF-Y TF, but also chromatin remodelers. Zooming in on the interaction capacity of VRS5 with other TCP TFs shows that VRS5 preferably interacts with other class II TCP TFs in the TB1 clade. Induced mutagenesis through CRISPR–Cas of one of the putative VRS5 interactors, HvTB2 (also referred to as COMPOSITUM 1 and BRANCHED AND INDETERMINATE SPIKELET 1), resulted in plants that have lost their characteristic unbranched spike architecture. More specifically,  hvtb2  mutants exhibited branches arising at the main spike, suggesting that  HvTB2  acts as inhibitor of branching. Our protein–protein interaction studies of VRS5 resulted in the identification of  HvTB2  as putative interactor of VRS5, another key regulator of spike architecture in barley. The study presented here provides a first step to underpin the protein–protein interactome of VRS5 and to identify other, yet unknown, key regulators of barley plant architecture.

The TCP transcription factor HvTB2 heterodimerizes with VRS5 and controls spike architecture in barley