Phocid Seal Leptin: Tertiary Structure and Hydrophobic Receptor Binding Site Preservation during Distinct Leptin Gene Evolution

The cytokine hormone leptin is a key signalling molecule in many pathways that control physiological functions. Although leptin demonstrates structural conservation in mammals, there is evidence of positive selection in primates, lagomorphs and chiropterans. We previously reported that the leptin genes of the grey and harbour seals (phocids) have significantly diverged from other mammals. Therefore we further investigated the diversification of leptin in phocids, other marine mammals and terrestrial taxa by sequencing the leptin genes of representative species. Phylogenetic reconstruction revealed that leptin diversification was pronounced within the phocid seals with a high dN/dS ratio of 2.8, indicating positive selection. We found significant evidence of positive selection along the branch leading to the phocids, within the phocid clade, but not over the dataset as a whole. Structural predictions indicate that the individual residues under selection are away from the leptin receptor (LEPR) binding site. Predictions of the surface electrostatic potential indicate that phocid seal leptin is notably different to other mammalian leptins, including the otariids. Cloning the grey seal leptin binding domain of LEPR confirmed that this was structurally conserved. These data, viewed in toto, support a hypothesis that phocid leptin divergence is unlikely to have arisen by random mutation. Based upon these phylogenetic and structural assessments, and considering the comparative physiology and varying life histories among species, we postulate that the unique phocid diving behaviour has produced this selection pressure. The Phocidae includes some of the deepest diving species, yet have the least modified lung structure to cope with pressure and volume changes experienced at depth. Therefore, greater surfactant production is required to facilitate rapid lung re-inflation upon surfacing, while maintaining patent airways. We suggest that this additional surfactant requirement is met by the leptin pulmonary surfactant production pathway which normally appears only to function in the mammalian foetus

Combined analysis of transcriptome and metabolite data reveals extensive differences between black and brown nearly-isogenic soybean (Glycine max) seed coats enabling the identification of pigment isogenes

<p>Abstract</p> <p>Background</p> <p>The <it>R </it>locus controls the color of pigmented soybean (<it>Glycine max</it>) seeds. However information about its control over seed coat biochemistry and gene expressions remains limited. The seed coats of nearly-isogenic black (<it>iRT</it>) and brown (<it>irT</it>) soybean (<it>Glycine max</it>) were known to differ by the presence or absence of anthocyanins, respectively, with genes for only a single enzyme (anthocyanidin synthase) found to be differentially expressed between isolines. We recently identified and characterized a UDP-glycose:flavonoid-3-<it>O</it>-glycosyltransferase (<it>UGT78K1</it>) from the seed coat of black (<it>iRT</it>) soybean with the aim to engineer seed coat color by suppression of an anthocyanin-specific gene. However, it remained to be investigated whether <it>UGT78K1 </it>was overexpressed with anthocyanin biosynthesis in the black (<it>iRT</it>) seed coat compared to the nearly-isogenic brown (<it>irT</it>) tissue.</p> <p>In this study, we performed a combined analysis of transcriptome and metabolite data to elucidate the control of the R locus over seed coat biochemistry and to identify pigment biosynthesis genes. Two differentially expressed late-stage anthocyanin biosynthesis isogenes were further characterized, as they may serve as useful targets for the manipulation of soybean grain color while minimizing the potential for unintended effects on the plant system.</p> <p>Results</p> <p>Metabolite composition differences were found to not be limited to anthocyanins, with specific proanthocyanidins, isoflavones, and phenylpropanoids present exclusively in the black (<it>iRT</it>) or the brown (<it>irT</it>) seed coat. A global analysis of gene expressions identified <it>UGT78K1 </it>and 19 other anthocyanin, (iso)flavonoid, and phenylpropanoid isogenes to be differentially expressed between isolines. A combined analysis of metabolite and gene expression data enabled the assignment of putative functions to biosynthesis and transport isogenes. The recombinant enzymes of two genes were validated to catalyze late-stage steps in anthocyanin biosynthesis <it>in vitro </it>and expression profiles of the corresponding genes were shown to parallel anthocyanin biosynthesis during black (<it>iRT</it>) seed coat development.</p> <p>Conclusion</p> <p>Metabolite composition and gene expression differences between black (<it>iRT</it>) and brown (<it>irT</it>) seed coats are far more extensive than previously thought. Putative anthocyanin, proanthocyanidin, (iso)flavonoid, and phenylpropanoid isogenes were differentially-expressed between black (<it>iRT</it>) and brown (<it>irT</it>) seed coats, and <it>UGT78K2 </it>and <it>OMT5 </it>were validated to code UDP-glycose:flavonoid-3-<it>O</it>-glycosyltransferase and anthocyanin 3'-<it>O</it>-methyltransferase proteins <it>in vitro</it>, respectively. Duplicate gene copies for several enzymes were overexpressed in the black (<it>iRT</it>) seed coat suggesting more than one isogene may have to be silenced to engineer seed coat color using RNA interference.</p

A supermatrix analysis of genomic, morphological, and paleontological data from crown Cetacea

<p>Abstract</p> <p>Background</p> <p>Cetacea (dolphins, porpoises, and whales) is a clade of aquatic species that includes the most massive, deepest diving, and largest brained mammals. Understanding the temporal pattern of diversification in the group as well as the evolution of cetacean anatomy and behavior requires a robust and well-resolved phylogenetic hypothesis. Although a large body of molecular data has accumulated over the past 20 years, DNA sequences of cetaceans have not been directly integrated with the rich, cetacean fossil record to reconcile discrepancies among molecular and morphological characters.</p> <p>Results</p> <p>We combined new nuclear DNA sequences, including segments of six genes (~2800 basepairs) from the functionally extinct Yangtze River dolphin, with an expanded morphological matrix and published genomic data. Diverse analyses of these data resolved the relationships of 74 taxa that represent all extant families and 11 extinct families of Cetacea. The resulting supermatrix (61,155 characters) and its sub-partitions were analyzed using parsimony methods. Bayesian and maximum likelihood (ML) searches were conducted on the molecular partition, and a molecular scaffold obtained from these searches was used to constrain a parsimony search of the morphological partition. Based on analysis of the supermatrix and model-based analyses of the molecular partition, we found overwhelming support for 15 extant clades. When extinct taxa are included, we recovered trees that are significantly correlated with the fossil record. These trees were used to reconstruct the timing of cetacean diversification and the evolution of characters shared by "river dolphins," a non-monophyletic set of species according to all of our phylogenetic analyses.</p> <p>Conclusions</p> <p>The parsimony analysis of the supermatrix and the analysis of morphology constrained to fit the ML/Bayesian molecular tree yielded broadly congruent phylogenetic hypotheses. In trees from both analyses, all Oligocene taxa included in our study fell outside crown Mysticeti and crown Odontoceti, suggesting that these two clades radiated in the late Oligocene or later, contra some recent molecular clock studies. Our trees also imply that many character states shared by river dolphins evolved in their oceanic ancestors, contradicting the hypothesis that these characters are convergent adaptations to fluvial habitats.</p