The genome of the choanoflagellate Monosiga brevicollis and the origin of metazoans

Abstract

Choanoflagellates are the closest known relatives of metazoans. To discover potential molecular mechanisms underlying the evolution of metazoan multicellularity, we sequenced and analysed the genome of the unicellular choanoflagellate Monosiga brevicollis. The genome contains approximately 9,200 intron-rich genes, including a number that encode cell adhesion and signalling protein domains that are otherwise restricted to metazoans. Here we show that the physical linkages among protein domains often differ between M. brevicollis and metazoans, suggesting that abundant domain shuffling followed the separation of the choanoflagellate and metazoan lineages. The completion of the M. brevicollis genome allows us to reconstruct with increasing resolution the genomic changes that accompanied the origin of metazoans.

Figure 1. Introduction to the choanoflagellate Monosiga brevicollis

a, The close phylogenetic affinity between choanoflagellates and metazoans highlights the value of the M. brevicollis genome for investigations into metazoan origins, the biology of the last common ancestor of metazoans (filled circle) and the biology of the last common ancestor of choanoflagellates and metazoans (open circle). Genomes from species shown with their abbreviation were used for protein domain comparisons in this study: human (Homo sapiens, Hsap), ascidian (Ciona intestinalis, Cint), Drosophila (Drosophila melanogaster,Dmel), cnidarian (N. vectensis, Nvec), M. brevicollis (Mbre), zygomycete (Rhizopus oryzae, Rory), basidiomycete (Coprinus cinereus, Ccin), ascomycete (Neurospora crassa, Ncra), hemiascomycete (Saccharomyces cerevisiae, Scer), slime mould (Dictyostelium discoideum, Ddis) and Arabidopsis (Arabidopsis thaliana, Atha). b–d, Choanoflagellate cells bear a single apical flagellum(arrow, b) and an apical collar of actin-filled microvilli (bracket, c). d, An overlay of β-tubulin (green), polymerized actin (red) and DNA localization (blue) reveals the position of the flagellum within the collar of microvilli. Scale bar, 2 µm.

Figure 2. Intron gain preceded the origin and diversification of metazoans

Ancestral intron content, intron gains and intron losses were inferred by the Csuros maximum likelihood method from a sample of 1,054 intron positions in 473 highly conserved genes in representative metazoans (humans, D. melanogaster and N. vectensis), M. brevicollis, intron-rich fungi (Cryptococcus neoformans A and Phanerochaete chrysosporium), plants and green algae (A. thaliana and Chlamydomonas reinhardtii), and a ciliate (Tetrahymena thermophila). Branches with more gain than loss are blue, those with more loss than gain are red, and those with comparable amounts of each are black. The inferred or observed number of introns present in ancestral and extant species is indicated by proportionally sized circles. As in Fig. 1, the last common ancestor of metazoans and the last common ancestor of choanoflagellates and metazoans are represented by a filled circle and an open circle, respectively. Other ancestral nodes are indicated by grey circles.

Figure 3. Phylogenetic distribution of metazoan-type cell adhesion domains and sequence-specific transcription factor families

M. brevicollis possesses diverse adhesion and ECM domains previously thought to be unique to metazoans (magenta). In contrast, many metazoan sequence-specific transcription factors are absent from the M. brevicollis gene catalogue. For adhesion and ECM domains, a filled box indicates a domain identified by both SMART and Pfam,, a half-filled box indicates a domain identified by either SMART or Pfam, and an open box indicates a domain that is not encoded by the current set of gene models. The presence (filled box) or absence (empty box) of transcription factor families was determined by reciprocal BLAST and SMART/Pfam domain annotations (Supplementary Note 3.5). Species names follow the convention from Fig. 1. EC, extracellular domain; cyto, cytoplasmic domain; asterisk, collagen triple-helix-domains occur in the extended tandem arrays diagnostic of collagen proteins found only in metazoans and choanoflagellates.

Figure 4. Domain shuffling and the evolution of Notch and hedgehog

Analysis of the draft gene set reveals that M. brevicollis possesses proteins containing domains characteristic of metazoan Notch (a, N1–N3) and hedgehog (b, H1 and H2). Some of these protein domains were previously thought to be unique to metazoans. The presence of these domains in separate M. brevicollis proteins implicates domain shuffling in the evolution of Notch and Hedgehog. Hh, hedgehog; N-hh, hedgehog N-terminal signalling domain; Hint, hedgehog/intein domain; TM, transmembrane domain; VWA, von Willebrand A domain. See Supplementary Note 3.6 for protein accession numbers and Supplementary Fig. 6 for identification of all displayed protein domains. Species names follow the convention from Fig. 1.

Figure 5. Divergent usage of protein domains involved in pTyr-based signalling between M. brevicollis and metazoans

A metric for functional usage of a domain within a genome is the number of other domains with which it co-occurs in a single protein. Numbers of pairwise domain combinations are indicated for classes of signalling domains involved in Ras, Rho, pSer/Thr and pTyr signalling. In cases in which a domain combination occurs multiple times within an individual protein or genome, it is only counted once. All combinations observed in M. brevicollis are indicated either as those that are only observed in the M. brevicollis genome (magenta) or as those that are observed both in M. brevicollis and metazoan genomes(grey). pTyr signalling domains in M. brevicollis are unique in that most of their observed pairwise domain combinations are distinct from those observed in metazoans. GEF, guanine-nucleotide exchange factor; GAP, GTPase-activating protein.