A pre-metazoan origin of the CRK gene family and co-opted signaling network.

CRK and CRKL adapter proteins play essential roles in development and cancer through their SRC homology 2 and 3 (SH2 and SH3) domains. To gain insight into the origin of their shared functions, we have investigated their evolutionary history. We propose a term, crk/crkl ancestral (crka), for orthologs in invertebrates before the divergence of CRK and CRKL in the vertebrate ancestor. We have isolated two orthologs expressed in the choanoflagellate Monosiga brevicollis, a unicellular relative to the metazoans. Consistent with its highly-conserved three-dimensional structure, the SH2 domain of M. brevicollis crka1 can bind to the mammalian CRK/CRKL SH2 binding consensus phospho-YxxP, and to the SRC substrate/focal adhesion protein BCAR1 (p130(CAS)) in the presence of activated SRC. These results demonstrate an ancient origin of the CRK/CRKL SH2-target recognition specificity. Although BCAR1 orthologs exist only in metazoans as identified by an N-terminal SH3 domain, YxxP motifs, and a C-terminal FAT-like domain, some pre-metazoan transmembrane proteins include several YxxP repeats in their cytosolic region, suggesting that they are remotely related to the BCAR1 substrate domain. Since the tyrosine kinase SRC also has a pre-metazoan origin, co-option of BCAR1-related sequences may have rewired the crka-dependent network to mediate adhesion signals in the metazoan ancestor

Appendix B. Detailed methods for all studies.

Detailed methods for all studies

Appendix C. The multidimensional scaling analysis of all measured floral traits for individual Silene plants.

The multidimensional scaling analysis of all measured floral traits for individual Silene plants

Premetazoan genome evolution and the regulation of cell differentiation in the choanoflagellate Salpingoeca rosetta

Abstract Background Metazoan multicellularity is rooted in mechanisms of cell adhesion, signaling, and differentiation that first evolved in the progenitors of metazoans. To reconstruct the genome composition of metazoan ancestors, we sequenced the genome and transcriptome of the choanoflagellate Salpingoeca rosetta, a close relative of metazoans that forms rosette-shaped colonies of cells. Results A comparison of the 55 Mb S. rosetta genome with genomes from diverse opisthokonts suggests that the origin of metazoans was preceded by a period of dynamic gene gain and loss. The S. rosetta genome encodes homologs of cell adhesion, neuropeptide, and glycosphingolipid metabolism genes previously found only in metazoans and expands the repertoire of genes inferred to have been present in the progenitors of metazoans and choanoflagellates. Transcriptome analysis revealed that all four S. rosetta septins are upregulated in colonies relative to single cells, suggesting that these conserved cytokinesis proteins may regulate incomplete cytokinesis during colony development. Furthermore, genes shared exclusively by metazoans and choanoflagellates were disproportionately upregulated in colonies and the single cells from which they develop. Conclusions The S. rosetta genome sequence refines the catalog of metazoan-specific genes while also extending the evolutionary history of certain gene families that are central to metazoan biology. Transcriptome data suggest that conserved cytokinesis genes, including septins, may contribute to S. rosetta colony formation and indicate that the initiation of colony development may preferentially draw upon genes shared with metazoans, while later stages of colony maturation are likely regulated by genes unique to S. rosetta

FoxJ1 from <i>T. adhaerens</i> and <i>S. purpuratus</i> are nuclear localized and can regulate the expression of ciliary genes.

<p>Anti-myc antibodies were used to detect Placozoa (A) and sea urchin (B) FoxJ1 (red, white arrow). Nuclei were stained with DAPI (blue). (C) Expression of <i>dynein intermediate chain</i> in the spinal cord (long arrow) and pronephric (kidney) duct (short arrow) of a wild-type zebrafish embryo. The <i>wdr78</i> and <i>efhc1</i> genes are expressed in a similar pattern in wild-type embryos (see <a href="http://www.plosgenetics.org/article/info:doi/10.1371/journal.pgen.1003019#pgen-1003019-g002" target="_blank">Figure 2A</a> and data not shown). Ectopic expression of <i>dynein intermediate chain</i> in embryos ectopically expressing placozoan (D) and sea urchin (E) FoxJ1, respectively. Ectopic expression of <i>wdr78</i> in embryos ectopically expressing placozoan (F) and sea urchin (G) FoxJ1, respectively. Ectopic expression of <i>efhc1</i> in embryos ectopically expressing placozoan (H) and sea urchin (I) FoxJ1, respectively. Mis-expression of the different ciliary genes in D–I is indicated by the arrows. Embryos depicted are at 20 hpf, oriented anterior to the left, dorsal to the top.</p

Zebrafish FoxJ2 and FoxJ3 are unable to induce the expression of ciliary genes.

<p>(A) Expression of <i>efhc1</i> in the spinal cord (long arrow) and pronephric (kidney) duct (short arrow) of a wild-type zebrafish embryo, and in embryos ectopically expressing zebrafish FoxJ2 (B) and FoxJ3 (C), respectively. (D) Expression of <i>spag6</i> in the spinal cord (long arrow) and pronephric (kidney) duct (short arrow) of a wild-type zebrafish embryo, and in embryos ectopically expressing zebrafish FoxJ2 (E) and FoxJ3 (F), respectively. Embryos depicted are at 20 hpf, oriented anterior to the left, dorsal to the top.</p