Genes and cDNAs encoding plant protein kinases highly homologous to the animal GSK-3/shaggy subfamily were isolated from Arabidopsis thaliana, Brassica napus, Petunia hybrida and Nicotiana tabacum using the P. hybrida PSK6 GSK-3/shaggy related cDNA as a probe. All the derived protein sequences contained the characteristic catalytic domain of GSK-3/shaggy protein kinases. Sequence comparisons within the catalytic domain with other plant GSK-3/shaggy like kinases clearly indicate that the novel sequences form an isolated group of genes termed the PSK6 group. All the proteins within this group possess an amino-terminal extension which contains short amino acid motifs highly conserved between species and possibly implicated in mitochondrial targeting. Northern hybridisation experiments and reverse transcriptase PCR analysis demonstrated that these novel cDNAs are predominantly expressed in developing pollen. The three genes isolated from P. hybrida and A. thaliana show the same genomic organisation into 12 introns and 13 exons. Although the size of the introns varies, their positions are conserved between genes and species. The comparison of these gene structures and the analysis of deduced protein sequences belonging to different plants hold important information to understand the function of individual members. They suggest that some of the characterised sequences represent most likely true orthologues whereas others must be paralogues. They also allow us to discuss the evolution of the plant GSK-3/shaggy like gene family with regard to plant speciation.