GLYCOGEN SYNTHASE KINASE3/Shaggy-like kinases (GSKs) represent a highly conserved group of proteins found in all eukaryotes. In plants they are encoded by multigene families and integrate signaling of brassinosteroids, auxin and abscisic acid in wide range of physiological and developmental processes with a strong impact on plant responses to environmental and biotic factors. Based on comprehensively studied structures of 10 Arabidopsis thaliana GSK genes and encoded proteins we report identification and phylogenetic reconstruction of 7 transcriptionally active GSK genes in barley. We re-evaluated annotation of the GSK genes in the current barley genome (Hv_IBSC_PGSB_v2) and provided data that a single gene annotated in the previous barley genome ensemble should be retained in the current one. The novel structure of another GSK, predicted in Hv_IBSC_PGSB_v2 to encode both GSK and amine oxidase domains, was proposed and experimentally confirmed based on the syntenic region in Brachypodium distachyon. The genes were assigned to 4 groups based on their encoded amino acid sequences and protein kinase domains. The analysis confirmed high level of conservation of functional protein domains and motifs among plant GSKs and the identified barley orthologs. Each of the seven identified HvGSK genes was expressed indicating semi-constitutive regulation in all tested organs and developmental stages. Regulation patterns of GSKs from the indicated groups showed a shift in organ-preferential expression in A. thaliana and barley illustrating diversification of biological roles of individual HvGSKs in different plant species.