Extremely slow rate of evolution in the HOX cluster revealed by comparison between Tanzanian and Indonesian coelacanths
Highlights
► We sequenced the HOX cluster of the Tanzanian coelacanth. ► The genetic divergence between two coelacanth species is very small. ► The slow substitution rate may account for the slow rate of coelacanth evolution.
Introduction
Coelacanths were initially recognized as a distinct taxonomic group by fossil records, which range in age from the Early Devonian to the Late Cretaceous periods (Maisey, 1996). Because no fossil records of coelacanths have been found since 65 million years ago (MYA), coelacanths were believed to have been extinct (Maisey, 1996). Therefore, the discovery in 1938 of the first living coelacanth, Latimeria chalumnae, off the coast of South Africa created a sensation in the field of evolutionary biology (Smith, 1939). One of the most interesting observations is that coelacanth morphology has changed very little (Holder et al., 1999, Smith, 1939); most of the characteristics unique to coelacanths (fleshy-lobed fins, hollow nerve cord, poor ossification of the skeleton, lack of defined ribs, and bilobed caudal region) have been maintained from the Devonian era (Carroll, 1988). Accordingly, coelacanths are called “evolutionary relics” or “living fossils”. The elucidation of the molecular mechanism of such slow morphological change in coelacanths is of primary importance to understand the morphological evolution of animals from genotype to phenotype.
After the discovery of a second living coelacanth in the Comoros archipelagos (Smith, 1953), the existence of a viable coelacanth population in this region was confirmed. In addition to the Comoros archipelagos, several coelacanths have been captured off the coasts of Mozambique (Schliewen et al., 1993), Madagascar (Heemstra et al., 1996), and Kenya (De Vos and Oyugi, 2002). Nikaido et al. (2011) recently found a genetically distinct coelacanth population off the northern coastal region of Tanzania, indicating that coelacanths are widely distributed throughout the western Indian Ocean. Apart from the western Indian Ocean, two coelacanth individuals were captured off the coast of Manado, Sulawesi, Indonesia (Erdman et al., 1998). These coelacanths are the first individuals recorded from a location outside the western Indian Ocean and were described as a new species, Latimeria menadoensis. The divergence time between the two coelacanth species was estimated in three independent studies. Holder et al. (1999) used partial mtDNA sequences and estimated the divergence time at 6 MYA. On the other hand, using the entire mtDNA sequences (except for the d-loop) and Bayesian methods, Inoue et al. (2005) estimated the divergence time at about 30 MYA. Sudarto et al. (2010) also used Bayesian analysis and proposed the divergence time to be 28 MYA.
HOX genes encode a highly conserved family of transcription factors possessing a 60‐amino acid residue motif called a homeodomain, and they are involved in morphogenesis during embryonic development (Krumlauf, 1994). Most of the jawed vertebrates have four separate HOX clusters–HOXA, HOXB, HOXC and HOXD–in which about 40 HOX genes are arranged. However, the number and composition of HOX clusters of teleost fish are distinct from those of the other vertebrates owing to a whole-genome duplication event specific to teleost fish (Meyer and Málaga-Trillo, 1999). In particular, duplication of the HOX clusters led to eight HOX clusters in an ancestor of teleost fish. Subsequently, some HOX genes were lost independently in the lineage of each teleost fish species during evolution. As a result, euteleosts have seven HOX clusters, in which 46 (medaka) to 49 (zebrafish) HOX genes were identified (Kurosawa et al., 2006). Amemiya et al. (2010) characterized the complete HOX clusters in the coelacanth genome. Although the HOX clusters of coelacanth were not remarkable relative to those from other species with four clusters, characterization of the complete HOX genes of coelacanth allowed us to reconstruct the evolutionary history of HOX clusters among vertebrates.
Consistent with the slow rate of phenotypic changes in coelacanth, several genetic studies showed a slow rate of evolution at the molecular level. Noonan et al. (2004) indicated that the content and organization of the procadherin gene cluster were more conserved in coelacanths than in teleost fish. Furthermore, they indicated fewer amino acid substitutions in coelacanths than in zebrafish and humans. Amemiya et al. (2010) also showed a significantly slower rate of amino acid substitution in the coelacanth HOX cluster than in teleost fish and tetrapods. These observations suggested that the coelacanth genes have evolved under strong purifying selection or that mutation rate has been slowed down in the coelacanth genome. To examine these possibilities, we directly estimated the absolute value of synonymous divergence of the two coelacanths. Namely, we determined the entire HOX cluster sequence for Tanzanian coelacanth L. chalumnae and compared it to that of L. menadoensis, which was already available in the literature (Amemiya et al., 2010).
Section snippets
Genomic DNA of Tanzanian coelacanth
Frozen or ethanol-preserved coelacanth materials were transferred from the Tanzania Fisheries Research Institute to the Tokyo Institute of Technology in accordance with international regulations under the Convention on International Trade in Endangered Species of Wild Fauna and Flora (CITES). In the present study, we used a juvenile coelacanth individual TCC041-1 to determine the HOX cluster sequence. The juvenile was found in the body of female coelacanth individual (TCC041) captured off the
Whole-genome sequencing and mapping
We performed two sequencing runs on the Illumina Genome Analyzer II platform and produced 85.9 Gbp of sequence data (Table 1). The short reads were aligned with the Burrows–Wheeler Alignment tool (Li and Durbin, 2009); we mapped 2.8% of the total reads to the L. menadoensis HOX cluster sequences, and 1.6% of the total reads were uniquely mapped. Insert size distribution showed not only major broad peaks around 150 bp but also a minor peak around 320 bp. This minor peak was created by the reads
Acknowledgments
This work was supported by the JSPS AA Science Platform Program, a Grant-in-Aid for Scientific Research (S) from the Ministry of Education, Culture, Sports, Science and Technology of Japan (to N.O.), and the Global COE Program “Deciphering Biosphere from Genome Big Bang” (to S.M.).
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These authors equally contributed to this work.