Molecular basis of urostyle development in frogs: Genes and gene regulation underlying an evolutionary novelty

Evolutionary novelties entail the origin of morphologies that enable new functions. These features can arise through changes to gene function and regulation. One key novelty is the fused rod at the end of the vertebral column in anurans, the urostyle. This feature is composed of a coccyx and a hypochord, both of which ossify during metamorphosis. To elucidate the genetic basis of these features, we used laser capture microdissection of these tissues and did RNA-seq and ATAC-seq at three developmental stages in tadpoles of Xenopus tropicalis. RNA-seq reveals that the coccyx and hypochord have two different molecular signatures. Neuronal (TUBB3) and muscle markers (MYH3) are upregulated in coccygeal tissues, whereas T-box genes (TBXT, TBXT.2), corticosteroid stress hormones (CRCH.1) and matrix metallopeptidases (MMP1, MMP8 and MMP13) are upregulated in the hypochord. ATAC-seq reveals potential regulatory regions that are observed in proximity to candidate genes that regulate ossification identified from RNA-seq. Even though an ossifying hypochord is only present in anurans, this ossification between the vertebral column and the notochord resembles a congenital vertebral anomaly seen prenatally in humans caused by an ectopic expression of the TBXT/TBXT.2 gene. This work opens the way to functional studies that can elucidate anuran bauplan evolution

Diversification of shrub frogs (Rhacophoridae, Pseudophilautus) in Sri Lanka - Timing and geographic context

US National Science Foundation (DEB 0345885) to CJS and JH; National Geographic Society (7612-04) to CJS; and Society of Systematic Biologists Graduate Student Award for Research to MM. MM was also supported by a Harvard University Center for the Environment (HUCE) Ziff Environmental Postdoctoral Fellowship. (DEB 0345885 - US National Science Foundation; 7612-04 - National Geographic Society; Society of Systematic Biologists Graduate Student Award for Research; Harvard University Center for the Environment (HUCE) Ziff Environmental Postdoctoral Fellowship)Accepted manuscrip

Unearthing the Fossorial Tadpoles of the Indian Dancing Frog Family Micrixalidae

Tadpoles of the monotypic Indian dancing frog family Micrixalidae have remained obscure for over 125 years. Here we report the discovery of the elusive tadpoles of Micrixalus herrei from the sand beds of a forested stream in southern Western Ghats, and confirm their identity through DNA barcoding. These actively burrowing tadpoles lead an entirely fossorial life from eggs to late metamorphic stages. We describe their internal and external morphological characters while highlighting the following features: eel-like appearance, extensively muscularized body and tail, reduced tail fins, skin-covered eyes, delayed development of eye pigmentation in early pre-metamorphic stages (Gosner stages 25–29), prominent tubular sinistral spiracle, large transverse processes on vertebrae II and III, ankylosed ribs on transverse processes of vertebra II, notochord terminating before the atlantal cotyle-occipital condyle junction, absence of keratodonts, serrated well-formed jaw sheaths, and extensive calcified endolymphatic sacs reaching sacrum posteriorly. The tadpole gut contains mostly fine sediments and sand. We discuss the eel-like morphology and feeding habits of M. herrei in the context of convergence with other well-known fossorial tadpoles. This discovery builds the knowledge base for further comparative analyses and conservation of Micrixalus, an ancient and endemic lineage of Indian frogs

Osteology of <i>Polypedates cruciger</i>, adult male.

<p>(A) Cranium, dorsal view. (B) Cranium, ventral view. (C) Lower jaw. (D) Hyoid skeleton. (E) Pectoral girdle. (F) Left hind limb. (G) Left forelimb. (H) Axial skeleton. Abbreviations: AG, angulosplenial; AT, atlas; ALP, anterolateral process; CN, centrum; COP, coronoid process; CP, carpals; CR, coracoid; CT, cleithrum; CV, clavicle; DP, distal phalange digit; DT, dentary; EX, exoccipital; FP, frontoparietal; HU, humerus; HP, hyoid plate; HY, hyale; IL, illium; MC, metacarpal; MNT, mentomeckelian; MT, metatarsal; MX, maxilla; NS, nasal; OC, oblique cartilage; OS, omosternum; PA, parasphenoid; PAR, pars articularis; PH, prehallux; PL, palatine; PLP, posterolateral process; PM, premaxilla; PMP, posteromedial process; PR, prootic; PT, pterygoid; QJ, quadratojugal; RL, radiale; RU, radioulna; SC, scapula; SD, sacral diapophysis; SP, sphenethmoid; SQ, squamosal; SS, suprascapula; ST, sternum; TA, tympanum annulus; TP, transverse process; TR, tarsal; UL, ulnare; US, urostyle; VM, vomer. Scale bar: 5 mm.</p

Comparative Postembryonic Skeletal Ontogeny in Two Sister Lineages of Old World Tree Frogs (Rhacophoridae: <i>Taruga</i>, <i>Polypedates</i>) - Fig 4

<p>Modification and repatterning of the chondrocrania at stages 31, 38 and 43 of <i>Taruga longinasus</i> (A), <i>T</i>. <i>eques</i> (B), <i>Polypedates maculatus</i> (C), and <i>P</i>. <i>cruciger</i> (D).</p

Comparison of the osteology of <i>Polypedates</i> and <i>Taruga</i>, adult males.

<p>(A) Cranium, dorsal view. (B) Cranium, ventral view. (C) Lower jaw. (D) Hyoid skeleton. (E) Pectoral girdle. (F) Axial skeleton of <i>Polypedates cruciger</i>, <i>P</i>. <i>maculatus</i>, <i>Taruga eques</i> and <i>T</i>. <i>longinasus</i>.</p

Chondrocrania of the foam-nesting genera, <i>Polypedates</i> and <i>Taruga</i>.

<p>(A) A labeled chondrocranium of <i>Polypedates maculatus</i>. Comparative illustrations of the chondrocrania of the four species: <i>Taruga eques</i> (B), <i>Taruga longinasus</i> (C), <i>Polypedates cruciger</i> (D) and <i>Polypedates maculatus</i> (E). Abbreviations: AP, ascending process; ARP, articular process; ASO, arcus subocularis; IC, infrarostral cartilage; LON, lamina orbitonasalis; MC, Meckel’s cartilage; MP, muscular process; OC, otic capsule; OP, otic process; QC, anterior quadratocranial commissure; QEP, quadratoethmoidal process; SA, suprarostral ala; SC, suprarostral cartilage; SMC, suprarostral medial corpus; TH, trabecular horns; TTD, taenia tecti medialis; TTM, taenia tecti marginalis; TS, tectum synoticum. Scale bar: 5 mm.</p

Comparison of the distal phalanges of the forelimb and hind limb.

<p>(A) <i>Taruga eques</i>. (B) <i>T</i>. <i>longinasus</i>. (C) <i>Polypedates cruciger</i>. (D) <i>P</i>. <i>maculatus</i>.</p

Comparison of ossification sequences and ossification indices (cranial bones) of <i>Taruga</i> and <i>Polypedates</i>.

<p>(A) Initial appearance of cranial bones (<i>N</i> = 19) is plotted against the Gosner stage for the four species. (B) Ossification indices of the four species is calculated to each individual (circles) by dividing the number of present ossified skull bones by the total number of scored elements in the cranium. The initial ossification is relatively slow in the two species of <i>Polypedates</i> (green and yellow open dots), fast in the two species of <i>Taruga</i> (blue and red open dots). But as development progresses, ossification rate of <i>Polypedates</i> species increases.</p