Hoxd13 contribution to the evolution of vertebrate appendages.

Fossil data suggest that limbs evolved from fish fins by sequential elaboration of their distal endoskeleton, giving rise to the autopod close to the tetrapod origin. This elaboration may have occurred by a simultaneous reduction of the distal ectodermal fold of fish fins. Modulation of 5'Hoxd gene transcription, through tetrapod-specific digit enhancers, has been suggested as a possible evolutionary mechanism involved in these morphological transformations. Here, we overexpress hoxd13a in zebrafish to investigate the impact of increasing 5'Hoxd expression during fin development. This overexpression causes increased proliferation, distal expansion of chondrogenic tissue and finfold reduction. In addition, we also show that the tetrapod-specific 5'Hoxd enhancer CsC promotes similar expression in zebrafish fins and mouse limbs. Our results support the idea that modulation of 5'Hoxd gene expression, by acquisition of novel enhancer elements, offered the substrate for the evolution of fins and the origin of tetrapod limbs.

The evolution of pepsinogen C genes in vertebrates: duplication, loss and functional diversification.

BACKGROUND: Aspartic proteases comprise a large group of enzymes involved in peptide proteolysis. This collection includes prominent enzymes globally categorized as pepsins, which are derived from pepsinogen precursors. Pepsins are involved in gastric digestion, a hallmark of vertebrate physiology. An important member among the pepsinogens is pepsinogen C (Pgc). A particular aspect of Pgc is its apparent single copy status, which contrasts with the numerous gene copies found for example in pepsinogen A (Pga). Although gene sequences with similarity to Pgc have been described in some vertebrate groups, no exhaustive evolutionary framework has been considered so far. METHODOLOGY/PRINCIPAL FINDINGS: By combining phylogenetics and genomic analysis, we find an unexpected Pgc diversity in the vertebrate sub-phylum. We were able to reconstruct gene duplication timings relative to the divergence of major vertebrate clades. Before tetrapod divergence, a single Pgc gene tandemly expanded to produce two gene lineages (Pgbc and Pgc2). These have been differentially retained in various classes. Accordingly, we find Pgc2 in sauropsids, amphibians and marsupials, but not in eutherian mammals. Pgbc was retained in amphibians, but duplicated in the ancestor of amniotes giving rise to Pgb and Pgc1. The latter was retained in mammals and probably in reptiles and marsupials but not in birds. Pgb was kept in all of the amniote clade with independent episodes of loss in some mammalian species. Lineage specific expansions of Pgc2 and Pgbc have also occurred in marsupials and amphibians respectively. We find that teleost and tetrapod Pgc genes reside in distinct genomic regions hinting at a possible translocation. CONCLUSIONS: We conclude that the repertoire of Pgc genes is larger than previously reported, and that tandem duplications have modelled the history of Pgc genes. We hypothesize that gene expansion lead to functional divergence in tetrapods, coincident with the invasion of terrestrial habitats.

Is there a compromise between nutrient uptake and gas exchange in the gut of Misgurnus anguillicaudatus, an intestinal air-breathing fish?

The Asian weatherloach, Misgurnus anguillicaudatus (Cobitidae), is a facultative air-breathing teleost fish that makes use of its hindgut or intestine as an accessory air-breathing organ (ABO). The hindgut is highly modified, being well vascularized with intraepithelial capillaries, which makes it well suited for gas exchange. However, the consequences for nutrient uptake, the traditional function of the intestine are unknown. The alimentary canal was examined histologically to assess differences between the fore-, mid- and hindgut regions that have been considered as the digestive, spiral and respiratory zones, respectively. In order to characterise the potential digestive (absorptive) function of the respiratory zone we used semi-quantitative polymerase chain reaction (PCR) to detect the presence of the intestinal Na(+):glucose cotransporter (SGLT1; SLC5A1) and H(+):peptide cotransporter (PEPT1a; SLC15A1) and partially sequenced the SGLT1 and PEPT1a cDNAs. These two transporters play important roles in the absorption of carbohydrate and di-/tripeptides, respectively, in the gut of fishes and other vertebrates and were therefore used as markers for potential nutrient uptake function. We also determined their tissue distributions through semi-quantitative RT-PCR. The effects of diet composition (high protein or high carbohydrate) or fasting on gene expression were also examined. SGLT1 expression was found in kidney, liver, heart, as well as in the three zones of the gut except the most distal part of the hindgut. PEPT1a mRNA was found in heart, brain, liver, and fore- and midgut, but absent in the hindgut. Our results clearly show high expression of SGLT1 (both mRNA and protein by immunolocalization) and PEPT1a (mRNA) in the foregut and midgut correlated with the digestive region of the gut. Modulatory effects of diet on the gene expression for both SGLT1 and PEPT1a were not observed. The presence of SGLT1 transcripts in the respiratory zone of the intestine suggests an overlap in function. However, in the case of PEPT1a, the distal limit was the midgut. Thus, despite its highly modified structure, the hindgut of the loach retains some potential nutrient uptake function.

Cadmium tolerance in the Nile tilapia (Oreochromis niloticus) following acute exposure: assessment of some ionoregulatory parameters.

The Nile tilapia (Oreochromis niloticus) can tolerate very high levels of waterborne cadmium. It has one of the highest 96 h LC50 recorded for a freshwater teleost fish (14.8 mg/L Cd; hardness 50 mg/L CaCO(3)). Cadmium is known to perturb ion balance in teleost fishes. However, in an acute time course experiment, plasma Na(+) concentrations were unaffected, and plasma Ca(2+) values only decreased after 96 h exposure in a dose-independent manner. Branchial Na(+)/K(+)-ATPase activity and alpha-subunit protein level expression in crude gill homogenates were not affected by Cd exposure during this period. Branchial chloride cell numbers, identified as Na(+)/K(+)-ATPase immunoreactive cells using immunohistochemistry, decreased 24 h after exposure but recovered thereafter. Histopathological changes did not follow a consistent pattern of variation with exposure time, and the alterations noted in gill epithelium were basically nonspecific to cadmium. Because of its tolerance, it can be concluded that the tilapia O. niloticus would not be a suitable test organism to evaluate sublethal toxicity of cadmium and the realistic impact of this pollutant in the environment. However, it certainly could contribute significantly to our understanding of the toxic mechanism of cadmium exposure in aquatic organisms. This is the first work to investigate the effect of waterborne pollutants on Na(+)/K(+)-ATPase alpha-subunit protein expression in fish gills.