Biology subjects

Felker, A.

Publications and source records attributed to Felker, A..

5 recordsLinked to original sources

bioRxiv2018

Cre/lox-controlled spatio-temporal perturbation of FGF signaling in zebrafish

Fibroblast Growth Factor (FGF) signaling guides multiple developmental processes including body axis formation and specific cell fate patterning. In zebrafish, genetic mutants and chemical perturbations affecting FGF signaling have uncovered key developmental processes; however, these approaches cause embryo-wide FGF signaling perturbations, rendering assessment of cell-autonomous versus non-autonomous requirements for FGF signaling in individual processes difficult. Here, we created the novel transgenic line fgfr1-dn-cargo, encoding dominant-negative Fgfr1 with fluorescent tag under combined Cre/lox and heatshock control to provide spatio-temporal perturbation of FGF signaling. Validating efficient perturbation of FGF signaling by fgfr1-dn-cargo primed with ubiquitous CreERT2, we established that primed, heatshock-induced fgfr1-dn-cargo behaves akin to pulsed treatment with the FGFR inhibitor SU5402. Priming fgfr1-dn-cargo with CreERT2 in the lateral plate mesoderm, we observed selective cardiac and pectoral fin phenotypes without drastic impact on overall embryo patterning. Harnessing lateral plate mesoderm-specific FGF inhibition, we recapitulated the cell-autonomous and temporal requirement for FGF signaling in pectoral fin outgrow, as previously hypothesized from pan-embryonic FGF inhibition. Altogether, our results establish fgfr1-dn-cargo as a genetic tool to define the spatio-temporal requirements for FGF signaling in zebrafish.

developmental biology↗

bioRxiv2018

A conserved regulatory program drives emergence of the lateral plate mesoderm

Cardiovascular lineages develop together with kidney, smooth muscle, and limb connective tissue progenitors from the lateral plate mesoderm (LPM). How the LPM initially emerges and how its downstream fates are molecularly interconnected remain unknown. Here, we isolated a pan-LPM enhancer in the zebrafish draculin (drl) gene that provides specific LPM reporter activity from early gastrulation. In toto live imaging and lineage tracing of drl-based reporters captured the dynamic LPM emergence as lineage-restricted mesendoderm field. The drl pan-LPM enhancer responds to the transcription factors EomesoderminA, FoxH1, and MixL1 that combined with Smad activity drive LPM emergence. We uncovered specific drl reporter activity in LPM-corresponding territories of several chordates including chicken, axolotl, lamprey, Ciona, and amphioxus, revealing a universal upstream LPM program. Altogether, our work provides a mechanistic framework for LPM emergence as defined progenitor field, possibly representing an ancient mesodermal cell state that predates the primordial vertebrate embryo.

developmental biology↗

bioRxiv2018

Mutations in Bcl9 and Pygo genes cause congenital heart defects by tissue-specific perturbation of Wnt/β-catenin signaling

Genetic alterations in human BCL9 genes have repeatedly been found in congenital heart disease (CHD) with as-of-yet unclear causality. BCL9 proteins and their Pygopus (Pygo) co-factors can participate in canonical Wnt signaling via binding to {beta}-catenin. Nonetheless, their contributions to vertebrate heart development remain uncharted. Here, combining zebrafish and mouse genetics, we document tissue-specific functions in canonical Wnt signaling for BCL9 and Pygo proteins during heart development. In a CRISPR-Cas9-based genotype-phenotype association screen, we uncovered that zebrafish mutants for bcl9 and pygo genes largely retain {beta}-catenin activity, yet develop cardiac malformations. In mouse, both systemic and lineage-specific loss of the Pygo-BCL9-{beta}-catenin complex caused heart defects with outflow tract malformations, aberrant cardiac septation and valve formation, and compact myocardium hypoplasia. Mechanistically, these phenotypes coincide with transcriptional deregulation during heart patterning, and Pygo2 associates with {beta}-catenin at cis-regulatory regions of cardiac genes. Taken together, our results establish BCL9 and Pygo as tissue-specific {beta}-catenin co-factors during vertebrate heart development. Our results further implicate alterations in BCL9 and BCL9L in human CHDs as possibly causative.

genetics↗

bioRxiv2017

Continuous addition of progenitors forms the cardiac ventricle in zebrafish

The vertebrate heart develops from several progenitor lineages. After early-differentiating first heart field (FHF) progenitors form the linear heart tube, late-differentiating second heart field (SHF) progenitors extend atrium, ventricle, and form the inflow and outflow tracts (IFT/OFT). However, the position and migration of late-differentiating progenitors during heart formation remains unclear. Here, we tracked zebrafish heart development using transgenics based on the cardiopharyngeal transcription factor gene tbx1. Live-imaging uncovered a tbx1 reporter-expressing cell sheath that from anterior lateral plate mesoderm continuously disseminates towards the forming heart tube. High-speed imaging and optogenetic lineage tracing corroborated that the zebrafish ventricle forms through continuous addition from the undifferentiated progenitor sheath followed by late-phase accrual of the bulbus arteriosus (BA). FGF inhibition during sheath migration reduced ventricle size and abolished BA formation, refining the window of FGF action during OFT formation. Our findings consolidate previous end-point analyses and establish zebrafish ventricle formation as a continuous process.

developmental biology↗

bioRxiv2017

Early frameshift alleles of zebrafish tbx5a that fail to develop the heartstrings phenotype

Tbx5 is a key transcription factor for vertebrate heart and forelimb development that causes Holt-Oram syndrome when mutated in humans. The classic zebrafish mutant for tbx5a named heartstrings (hst) features recessive absence of pectoral fins and a spectrum of heart defects, most-prominently featuring the name-giving stretched heart tube. The mutation of the hst allele is a stop codon that is predicted to result in a truncated Tbx5a protein that might feature residual activity. Here, using CRISPR-Cas9 mutagenesis, we generated zebrafish strains for two new tbx5a frameshift alleles in the first coding exon: tbx5a c.21_25del and tbx5a c.22_31del, abbreviated as tbx5a{Delta}5 and tbx5a{Delta}10. Homozygous and trans-heterozygous combinations of these new tbx5a alleles cause fully penetrant loss of pectoral fins and heart defects including changes in cardiac marker expression akin to hst mutants. Nonetheless, distinct from hst mutants, homozygous and trans-heterozygous combinations of these tbx5a frameshift mutants do not readily manifest the stretched hst heart phenotype. Our observation points out the importance and value of comparing phenotypes from different classes of mutant alleles per gene.

developmental biology↗