Comparative analysis of somitogenesis related genes of the hairy/Enhancer of split class in Fugu and zebrafish

BACKGROUND: Members of a class of bHLH transcription factors, namely the hairy (h), Enhancer of split (E(spl)) and hairy-related with YRPW motif (hey) (h/E(spl)/hey) genes are involved in vertebrate somitogenesis and some of them show cycling expression. By sequence comparison, identified orthologues of cycling somitogenesis genes from higher vertebrates do not show an appropriate expression pattern in zebrafish. The zebrafish genomic sequence is not available yet but the genome of Fugu rubripes was recently published. To allow comparative analysis, the currently known Her proteins from zebrafish were used to screen the genomic sequence database of Fugu rubripes. RESULTS: 20 h/E(spl)/hey-related genes were identified in Fugu, which is twice the number of corresponding zebrafish genes known so far. A novel class of c-Hairy proteins was identified in the genomes of Fugu and Tetraodon. A screen of the human genome database with the Fugu proteins yielded 10 h/E(spl)/hey-related genes. By analysing the upstream sequences of the c-hairy class genes in zebrafish, Fugu and Tetraodon highly similar sequence stretches were identified that harbour Suppressor of hairless paired binding sites (SPS). This motif was also discovered in the upstream sequences of the her1 gene in the examined fish species. Here, the Su(h) sites are separated by longer intervening sequences. CONCLUSIONS: Our study indicates that not all her homologues in zebrafish have been isolated. Comparison to the human genome suggests a selective duplication of h/E(spl) genes in pufferfish or loss of members of these genes during evolution to the human lineage

Formin homology 2 domains occur in multiple contexts in angiosperms

BACKGROUND: Involvement of conservative molecular modules and cellular mechanisms in the widely diversified processes of eukaryotic cell morphogenesis leads to the intriguing question: how do similar proteins contribute to dissimilar morphogenetic outputs. Formins (FH2 proteins) play a central part in the control of actin organization and dynamics, providing a good example of evolutionarily versatile use of a conserved protein domain in the context of a variety of lineage-specific structural and signalling interactions. RESULTS: In order to identify possible plant-specific sequence features within the FH2 protein family, we performed a detailed analysis of angiosperm formin-related sequences available in public databases, with particular focus on the complete Arabidopsis genome and the nearly finished rice genome sequence. This has led to revision of the current annotation of half of the 22 Arabidopsis formin-related genes. Comparative analysis of the two plant genomes revealed a good conservation of the previously described two subfamilies of plant formins (Class I and Class II), as well as several subfamilies within them that appear to predate the separation of monocot and dicot plants. Moreover, a number of plant Class II formins share an additional conserved domain, related to the protein phosphatase/tensin/auxilin fold. However, considerable inter-species variability sets limits to generalization of any functional conclusions reached on a single species such as Arabidopsis. CONCLUSIONS: The plant-specific domain context of the conserved FH2 domain, as well as plant-specific features of the domain itself, may reflect distinct functional requirements in plant cells. The variability of formin structures found in plants far exceeds that known from both fungi and metazoans, suggesting a possible contribution of FH2 proteins in the evolution of the plant type of multicellularity

Investigations into a putative role for the novel BRASSIKIN pseudokinases in compatible pollen-stigma interactions in Arabidopsis thaliana.

BACKGROUND: In the Brassicaceae, the early stages of compatible pollen-stigma interactions are tightly controlled with early checkpoints regulating pollen adhesion, hydration and germination, and pollen tube entry into the stigmatic surface. However, the early signalling events in the stigma which trigger these compatible interactions remain unknown. RESULTS: A set of stigma-expressed pseudokinase genes, termed BRASSIKINs (BKNs), were identified and found to be present in only core Brassicaceae genomes. In Arabidopsis thaliana Col-0, BKN1 displayed stigma-specific expression while the BKN2 gene was expressed in other tissues as well. CRISPR deletion mutations were generated for the two tandemly linked BKNs, and very mild hydration defects were observed for wild-type Col-0 pollen when placed on the bkn1/2 mutant stigmas. In further analyses, the predominant transcript for the stigma-specific BKN1 was found to have a premature stop codon in the Col-0 ecotype, but a survey of the 1001 Arabidopsis genomes uncovered three ecotypes that encoded a full-length BKN1 protein. Furthermore, phylogenetic analyses identified intact BKN1 orthologues in the closely related outcrossing Arabidopsis species, A. lyrata and A. halleri. Finally, the BKN pseudokinases were found to be plasma-membrane localized through the dual lipid modification of myristoylation and palmitoylation, and this localization would be consistent with a role in signaling complexes. CONCLUSION: In this study, we have characterized the novel Brassicaceae-specific family of BKN pseudokinase genes, and examined the function of BKN1 and BKN2 in the context of pollen-stigma interactions in A. thaliana Col-0. Additionally, premature stop codons were identified in the predicted stigma specific BKN1 gene in a number of the 1001 A. thaliana ecotype genomes, and this was in contrast to the out-crossing Arabidopsis species which carried intact copies of BKN1. Thus, understanding the function of BKN1 in other Brassicaceae species will be a key direction for future studies