Biology subjects

Eggenschwiler, J.

Publications and source records attributed to Eggenschwiler, J..

3 recordsLinked to original sources

bioRxiv2023

A new cell culture resource for investigations of reptilian gene function

The recent establishment of CRISPR/Cas9 gene editing in A. sagrei lizards makes it a powerful model system for studies of reptilian gene function. To enhance the versatility of this model, we developed an immortalized lizard fibroblast cell line (ASEC-1) for the exploration of reptilian gene function in cellular processes. We demonstrate the use of this in vitro system by scrutinizing the role of primary cilia in lizard Hedgehog (Hh) signaling. Through CRISPR/Cas9 mutagenesis we disrupted the ift88 gene, which is required for ciliogenesis in diverse organisms. We find that the loss of itf88 from lizard cells results in an absence of primary cilia, a partial derepression of gli1 transcription, and an inability of the cells to respond to the Smoothened agonist, SAG. Through a cross-species analysis of SAG-induced transcriptional responses in cultured limb bud cells, we further determined that [~]46% of genes induced as a response to Hh pathway activation in A. sagrei, are also SAG-responsive in M. musculus limb bud cells. Our results highlight conserved and diverged aspects of Hh signaling in anoles and establish a new resource for investigations of reptilian gene function.

developmental biology↗

bioRxiv2021

Evolution of a chordate-specific mechanism for myoblast fusion

The size of an animal is determined by the size of its musculoskeletal system. Myoblast fusion is an innovative mechanism that allows for multinucleated muscle fibers to compound the size and strength of individual mononucleated cells. However, the evolutionary history of the control mechanism underlying this important process is currently unknown. The phylum Chordata hosts closely related groups that span distinct myoblast fusion states: no fusion in cephalochordates, restricted fusion and multinucleation in tunicates, and extensive, obligatory fusion in vertebrates. To elucidate how these differences may have evolved, we studied the evolutionary origins and function of membrane-coalescing agents Myomaker and Myomixer in various groups of chordates. Here we report that Myomaker likely arose through gene duplication in the last common ancestor of tunicates and vertebrates, while Myomixer appears to have evolved de novo in early vertebrates. Functional tests revealed an unexpectedly complex evolutionary history of myoblast fusion in chordates. A pre-vertebrate phase of muscle multinucleation driven by Myomaker was followed by the later emergence of Myomixer that enables the highly efficient fusion system of vertebrates. Thus, our findings reveal the evolutionary origins of chordate-specific fusogens and illustrate how new genes can shape the emergence of novel morphogenetic traits and mechanisms.

developmental biology↗

bioRxiv2020

Development of a method for revertable CRISPR/Cas9-based mutagenesis in cell culture

CRISPR/Cas9 mutagenesis is a revolutionary tool for genetics in organismal and cell culture systems. One notable caveat with this system is the potential for phenotype-inducing off-target/background mutations. There has been considerable success in modifying the methodology to minimize these potential confounds. Here we have developed a tool to functionally demonstrate that a targeted mutation of interest is responsible for the phenotype observed. This approach creates revertable mutations in cell culture systems using CRISPR/Cas9-induced homology-directed repair (HDR) to insert a LoxP-flanked transcriptional stop sequence into an early intron of a target gene. This method has the potential to be used in multiplexed and inducible scenarios to restore gene function within a given experiment.

cell biology↗