Divergent cis-regulatory evolution underlies the convergent loss of sodium channel expression in electric fish.

South American and African weakly electric fish independently evolved electric organs from muscle. In both groups, a voltage-gated sodium channel gene independently lost expression from muscle and gained it in the electric organ, allowing the channel to become specialized for generating electric signals. It is unknown how this voltage-gated sodium channel gene is targeted to muscle in any vertebrate. We describe an enhancer that selectively targets sodium channel expression to muscle. Next, we demonstrate how the loss of this enhancer, but not trans-activating factors, drove the loss of sodium channel gene expression from muscle in South American electric fish. While this enhancer is also altered in African electric fish, key transcription factor binding sites and enhancer activity are retained, suggesting that the convergent loss of sodium channel expression from muscle in these two electric fish lineages occurred via different processes.

Elongation during segmentation shows axial variability, low mitotic rates, and synchronized cell cycle domains in the crustacean, Thamnocephalus platyurus.

BACKGROUND: Segmentation in arthropods typically occurs by sequential addition of segments from a posterior growth zone. However, the amount of tissue required for growth and the cell behaviors producing posterior elongation are sparsely documented. RESULTS: Using precisely staged larvae of the crustacean, Thamnocephalus platyurus, we systematically examine cell division patterns and morphometric changes associated with posterior elongation during segmentation. We show that cell division occurs during normal elongation but that cells in the growth zone need only divide ~ 1.5 times to meet growth estimates; correspondingly, direct measures of cell division in the growth zone are low. Morphometric measurements of the growth zone and of newly formed segments suggest tagma-specific features of segment generation. Using methods for detecting two different phases in the cell cycle, we show distinct domains of synchronized cells in the posterior trunk. Borders of cell cycle domains correlate with domains of segmental gene expression, suggesting an intimate link between segment generation and cell cycle regulation. CONCLUSIONS: Emerging measures of cellular dynamics underlying posterior elongation already show a number of intriguing characteristics that may be widespread among sequentially segmenting arthropods and are likely a source of evolutionary variability. These characteristics include: the low rates of posterior mitosis, the apparently tight regulation of cell cycle at the growth zone/new segment border, and a correlation between changes in elongation and tagma boundaries.

Silencing the Spark: CRISPR/Cas9 Genome Editing in Weakly Electric Fish.

Electroreception and electrogenesis have changed in the evolutionary history of vertebrates. There is a striking degree of convergence in these independently derived phenotypes, which share a common genetic architecture. This is perhaps best exemplified by the numerous convergent features of gymnotiforms and mormyrids, two species-rich teleost clades that produce and detect weak electric fields and are called weakly electric fish. In the 50 years since the discovery that weakly electric fish use electricity to sense their surroundings and communicate, a growing community of scientists has gained tremendous insights into evolution of development, systems and circuits neuroscience, cellular physiology, ecology, evolutionary biology, and behavior. More recently, there has been a proliferation of genomic resources for electric fish. Use of these resources has already facilitated important insights with regards to the connection between genotype and phenotype in these species. A major obstacle to integrating genomics data with phenotypic data of weakly electric fish is a present lack of functional genomics tools. We report here a full protocol for performing CRISPR/Cas9 mutagenesis that utilizes endogenous DNA repair mechanisms in weakly electric fish. We demonstrate that this protocol is equally effective in both the mormyrid species Brienomyrus brachyistius and the gymnotiform Brachyhypopomus gauderio by using CRISPR/Cas9 to target indels and point mutations in the first exon of the sodium channel gene scn4aa. Using this protocol, embryos from both species were obtained and genotyped to confirm that the predicted mutations in the first exon of the sodium channel scn4aa were present. The knock-out success phenotype was confirmed with recordings showing reduced electric organ discharge amplitudes when compared to uninjected size-matched controls.

Electric fish genomics: Progress, prospects, and new tools for neuroethology.

Electric fish have served as a model system in biology since the 18th century, providing deep insight into the nature of bioelectrogenesis, the molecular structure of the synapse, and brain circuitry underlying complex behavior. Neuroethologists have collected extensive phenotypic data that span biological levels of analysis from molecules to ecosystems. This phenotypic data, together with genomic resources obtained over the past decades, have motivated new and exciting hypotheses that position the weakly electric fish model to address fundamental 21st century biological questions. This review article considers the molecular data collected for weakly electric fish over the past three decades, and the insights that data of this nature has motivated. For readers relatively new to molecular genetics techniques, we also provide a table of terminology aimed at clarifying the numerous acronyms and techniques that accompany this field. Next, we pose a research agenda for expanding genomic resources for electric fish research over the next 10years. We conclude by considering some of the exciting research prospects for neuroethology that electric fish genomics may offer over the coming decades, if the electric fish community is successful in these endeavors.

Wnt repertoire and developmental expression patterns in the crustacean Thamnocephalus platyurus.

Wnt genes are a family of conserved glycoprotein ligands that play a role in a wide variety of cell and developmental processes, from cell proliferation to axis elongation. There are 13 Wnt subfamilies found among metazoans. Eleven of these appear conserved in arthropods with a pattern of loss during evolution of as many as six subfamilies among hexapods. Here we report on Wnt genes in the branchiopod crustacean, Thamnocephalus platyurus, including the first documentation of the expression of the complete Wnt gene family in a crustacean. Our results suggest fewer Wnt genes were retained in Thamnocephalus than in the related crustacean, Daphnia, although the Thamnocephalus Wnt repertoire is larger than that found in insects. We also find an intriguing pattern of staggered expression of Wnts-an anterior-posterior stagger within the posterior growth zone and a dorsal-ventral stagger in the developing segments-suggesting a potential for subfunctionalization of Wnts in these regions.