Gel-forming mucins appeared early in metazoan evolution

Abstract

Mucins are proteins that cover and protect epithelial cells and are characterized by domains rich in proline, threonine, and serine that are heavily glycosylated (PTS or mucin domains). Because of their sequence polymorphism, these domains cannot be used for evolutionary analysis. Instead, we have made use of the von Willebrand D (VWD) and SEA domains, typical for mucins. A number of animal genomes were examined for these domains to identify mucin homologues, and domains of the resulting proteins were used in phylogenetic studies. The frog Xenopus tropicalis stands out because the number of gel-forming mucins has markedly increased to at least 25 as compared with 5 for higher animals. Furthermore, the frog Muc2 homologues contain unique PTS domains where cysteines are abundant. This animal also has a unique family of secreted mucin-like proteins with alternating PTS and SEA domains, a type of protein also identified in the fishes. The evolution of the Muc4 mucin seems to have occurred by recruitment of a PTS domain to AMOP, NIDO, and VWD domains from a sushi domain-containing family of proteins present in lower animals, and Xenopus is the most deeply branching animal where a protein similar to the mammalian Muc4 was identified. All transmembrane mucins seem to have appeared in the vertebrate lineage, and the MUC1 mucin is restricted to mammals. In contrast, proteins with properties of the gel-forming mucins were identified also in the starlet sea anemone Nematostella vectensis, demonstrating an early origin of this group of mucins.

Fig. 1.

Proteins with VWD domains. (A) Domain structures of selected proteins containing the VWD domain, including proteins showed as colored branches in C. Organisms represented are human (h), X. tropicalis (Xt), C. intestinalis (Ci), S. purpuratus (Sp), N. vectensis (Nv), D. melanogaster (Dm), and B. floridae (Bf). (B) Phylogenetic tree of VWD domains. Four hundred thirty-one VWD domains from 147 proteins were aligned by using ClustalW, and shown in the figure is the neighbor-joining tree derived by the same program. Branches with the D1, D2, D3, and D4 domains (A) corresponding to the gel-forming mucins otogelin and VWF are indicated as well as Muc4 and a branch containing the group of proteins (IgG-binding, tectorin, zonadhesin, and SCO-spondin) that do not group with the D1–D4 domains. The D1–D4 domains are shown with different colors. A rectangular version of this tree with more details on the individual branches as well as bootstrapping data is shown at www.medkem.gu.se/mucinbiology/databases. (C) Tree as in B but with VWDs of proteins of N. vectensis (red), S. purpuratus (green), B. floridae (blue), and C. intestinalis (yellow). All of the VWD domains of these proteins clearly map to the D1–D4 clusters shown in B.

Fig. 2.

Distribution of mucins. Shaded boxes indicate where a protein was found. Organisms are Homo sapiens (Hs), Mus musculus (Mm), G. gallus (Gg), X. tropicalis (Xt), T. rubripes (Tr), D. rerio (Dr), C. intestinalis (Ci), B. floridae (Bf), D. melanogaster (Dm), C. elegans (Ce), and N. vectensis (Nv). In the case where MV proteins have been indicated, a classification into Muc2, Muc5, etc. is not possible. A more detailed version of this table is found in SI Table 1.

Fig. 3.

Phylogenetic distribution of gel-forming mucins (A) and Muc4 (B). The current phylogenetic distribution is shown in the context of metazoan taxonomy. Schematic domain structures are shown, and domains are not drawn to scale.

Fig. 4.

SEA domain proteins. (A) Domain structures of SEA domain-containing protein, including mucins and mucin-like proteins in Xenopus and fishes with alternating SEA and PTS domains. Proteins shown include previously known human mucins (hMUC) and human IMPG2, as well a previously uncharacterized type of mucins from T. rubripes (TrMuc), X. tropicalis (XtMuc), and D. rerio (DrMuc). (B) Phylogenetic tree of SEA domains. One hundred forty-nine VWD domains from 83 proteins were aligned by using ClustalW, and shown is the neighbor-joining tree derived by the same program. The mucin groups shown are Muc16 (green), Muc13 (blue), Muc1 (red), Muc3 (yellow), and the previously uncharacterized type of mucins with alternating PTS and SEA domains (pink). (C) Phylogenetic distribution of SEA domain mucins. The current phylogenetic distribution is shown in the context of metazoan taxonomy. Schematic domain structures are shown where domains are not drawn to scale. In the case of Monodelphis, only C-terminal parts of Muc1 and Muc3 have been identified, and the full domain structure is not known. Multiple SEA domains in the Homo and Gallus Muc16 have been left out.