More Than One-to-Four via 2R: Evidence of an Independent Amphioxus Expansion and Two-Gene Ancestral Vertebrate State for MyoD-Related Myogenic Regulatory Factors (MRFs)

Abstract

The evolutionary transition from invertebrates to vertebrates involved extensive gene duplication, but understanding precisely how such duplications contributed to this transition requires more detailed knowledge of specific cases of genes and gene families. Myogenic differentiation (MyoD) has long been recognized as a master developmental control gene and member of the MyoD family of bHLH transcription factors (myogenic regulatory factors [MRFs]) that drive myogenesis across the bilaterians. Phylogenetic reconstructions within this gene family are complicated by multiple instances of gene duplication and loss in several lineages. Following two rounds of whole-genome duplication (2R WGD) at the origin of the vertebrates, the ancestral function of MRFs is thought to have become partitioned among the daughter genes, so that MyoD and Myf5 act early in myogenic determination, whereas Myog and Myf6 are expressed later, in differentiating myoblasts. Comparing chordate MRFs, we find an independent expansion of MRFs in the invertebrate chordate amphioxus, with evidence for a parallel instance of subfunctionalization relative to that of vertebrates. Conserved synteny between chordate MRF loci supports the 2R WGD events as a major force in shaping the evolution of vertebrate MRFs. We also resolve vertebrate MRF complements and organization, finding a new type of vertebrate MRF gene in the process, which allowed us to infer an ancestral two-gene state in the vertebrates corresponding to the early- and late-acting types of MRFs. This necessitates a revision of previous conclusions about the simple one-to-four origin of vertebrate MRFs.

Somite Compartments in Amphioxus and Its Implications on the Evolution of the Vertebrate Skeletal Tissues

“…Later during development, MRF2 is also found in the dorsolateral somite, adjacent to dermomyotome markers like Pax3/7 ( Figures 4F,G ). This pattern corresponds to the expression of the newly identified amphioxus MRF4 , which is also found in the dorsolateral portion of the lateral somite ( Aase-Remedios et al, 2020 ). While these expression data suggest a possible DML-like location, further lineage tracing experiments will be needed to determine whether this region behaves like the DML and contributes to myotome precursors.…”

An Updated Staging System for Cephalochordate Development: One Table Suits Them All

“…Developing B. lanceolatum embryos were reared at three different temperatures: 16 • C, 19 • C, and 22 • C. At regular intervals, animals were collected and fixed for subsequent in situ hybridization analyses. A 874-bp fragment containing the complete coding sequence of the B. lanceolatum mrf1 (myogenic regulatory factor 1) gene, a member of the myoD gene family (Schubert et al, 2003;Aase-Remedios et al, 2020), was amplified by PCR from cDNA and cloned into the pGEM-T Easy Vector (GenBank accession number of B. lanceolatum mrf1: MT452570). In situ hybridization experiments were carried out with a mrf1-specific antisense riboprobe as previously described (Yu and Holland, 2009;Carvalho et al, 2017c).…”

Improved Understanding of the Role of Gene and Genome Duplications in Chordate Evolution With New Genome and Transcriptome Sequences

“…Mirroring this cluster, a cryptic fifth MRF ohnologue is located adjacent to MyoD in the genomes of the coelacanth, sterlet, and spotted gar (Aase-Remedios et al, 2020). Only with the recent availability of the genomes of these "non-model" organisms could this fifth ohnologue, Myf7, be found, since it has been lost from most other vertebrate lineages to which common model organisms belong, including tetrapods, cartilaginous fish, and teleosts.…”

“…The finding of this fifth vertebrate MRF upended previous interpretations of the evolution of the four MRFs shared amongst all vertebrates. Instead of a single ancestral gene, duplicated in 2R into four ohnologues, and a highly unusual translocation event, it is much more likely that there was an ancestral two-gene state resulting from a tandem duplication that generated the two types of MRF, early and late (Aase-Remedios et al, 2020). This cluster nevertheless shares synteny with the MyoD locus across the vertebrates, potentially as a result of constraint on regulatory elements that overlap with the MRFs and their gene neighbours.…”

“…In many important developmental gene families, amphioxus has undergone its own expansions of gene clusters, formed by tandem duplications (Minguillón et al, 2002). This ranges from expansions within existing clusters, possibly like the Posterior Hox gene AmphiHox15 within the Hox cluster (Holland et al, 2008), to the advent of new clusters, like the expansion of the amphioxus MRFs (Schubert et al, 2003;Urano et al, 2003;Yuan et al, 2003;Somorjai et al, 2008;Tan et al, 2014;Aase-Remedios et al, 2020). The five genes in this cluster are expressed in different temporal and spatial patterns during amphioxus muscle development, indicating these genes have subfunctionalised as well, providing an independent case of MRF subfunctionalisation from that seen with the vertebrate MRFs (Aase-Remedios et al, 2020).…”