Tag: Charles Kimmel

Genetic Interaction in the Developing Danio Rerio Jaw

Presenter: Braden Larson, Biology, Russian and Eastern European Studies (REES)

Poster: B-7

Mentor: Charles Kimmel, Institute of Neuroscience

Genes act in concert during animal development to form complex anatomical structures. For example, the jaw skeleton requires precise expression and interaction of a multitude of genes to develop correctly. Our research focuses on the gene endothelin-1 (edn1), which encodes a signaling molecule required for ventral jaw development in the zebrafish, Danio rerio. Because elements of the jaw remain in the edn1 mutant, we hypothesized that other genes function within the edn1 genetic pathway. To test this, we generated double mutant zebrafish, pairing the edn1 mutant allele with mutant alleles of candidate genes based on previous literature. We then analyzed the double mutant skeletons for evidence of genetic interaction, and discovered an enhanced phenotype in one of the double mutants. Specifically, mutants for edn1 and fibroblast growth factor-8a (fgf8a), a gene that encodes another signaling molecule, are missing a portion of their upper jaw, a phenotype not present in either single mutant. To investigate the cell biology behind this phenotype we imaged the cells that constitute the jaw precursor tissue in live transgenic zebrafish. Strikingly, the cells that give rise to this structure appear missing in double mutant fish. Furthermore, we used Fluorescent In Situ Hybridization to observe gene expression of edn1, fgf8a, and potential shared target genes in wild type and mutant embryos. We conclude that cross talk between edn1 and fgf8a signaling is required for development of the jaw skeleton.

Interactions Between ihha and mef2ca in Bone Development

Presenters: Ashlin Larsen, Tyler R. Huycke and Charles B. Kimmel

Mentor: Charles Kimmel

PM Poster Presentation

Poster 24

Bones form through the differentiation of mesenchymal cells into bone-forming osteoblasts, and multiple genes regulate this process in order to ensure proper bone size and shape. Myocyte enhancer factor 2ca (mef2ca) encodes a transcription factor that negatively regulates development of the opercle (Op), a craniofacial bone of the zebrafish. Loss of mef2ca can lead to ectopic bone growth along the antero-ventral edge of the Op, yet the developmental mechanism underlying this process is unclear. We tested whether indian hedgehog a (ihha), a positive regulator of Op development acting in the same region of bone that mef2ca negatively regulates, is required for the complete ectopic bone growth seen in mef2ca mutants. With fluorescent in situ hybridization we show that expression of ihha and ptch1, a downstream target of active Hedgehog signaling, is present in ectopic bone growth of mef2ca mutants, suggesting that mef2ca may regulate Ihha signaling to pattern bone. Furthermore, analysis of mef2ca; ihha double mutant larvae reveals many ventrally reduced bones that resemble ihha single mutants; however, many also display the mef2ca mutant phenotype. Therefore, although Ihha is required for certain mef2ca mutant phenotypes, it is not imperative to induce expansion of bone in mef2ca mutants, and thus mef2ca must im- pose its negative regulation of bone development by acting through distinct gene networks in addition to the Hedgehog pathway.

Shaping the Face: Genetic Interactions in Zebrafish Jaw Development

Presenter: Braden Larson

Mentor: Charles Kimmel

AM Poster Presentation

Poster 25

Development of the jaw skeleton can be divided into three stages: (1) migration of neural crest cells, precursors of jaw skeletal cells, (2) aggregation of neural crest cells into pharyngeal arches, and (3) differentiation of pharyngeal arch cells into skeletal cells. Signaling between cells is instrumental in all three stages. One signal, Endothelin-1 (edn1), is known to be required for lower jaw development in zebrafish. Work in mice has shown that the signaling molecule Fibroblast growth factor-8 (fgf8) is required for proper edn1 expression, motivating the following hypothesis: edn1 and fgf8a genetically interact in zebrafish jaw development. / We tested this hypothesis by generating fish that carry mutations in both edn1 and fgf8a. These fish are missing cartilage cells from their upper jaw, a phenotype not present in either single mutant. To investigate the cause of this phenotype we examined cells of the pharyngeal arches. We found that the first arch, which is the precursor to the lower and upper jaw, to be dysmorphic. We then examined neural crest cell migration. While migration appeared normal, neural crest cells failed to aggregate into the intermediate region of the first arch in double mutant fish, placing the manifestation of the double mutant phenotype in stage two of jaw skeleton development. Our findings suggest a novel genetic interaction or synergy between edn1 and fgf8a in which they function to promote aggregation of neural crest cells into the intermediate region of the first pharyngeal arch.

Variation of Self-sorting Behavior and Jaw Bone Morphology in Early Development within Chinook Salmon (Oncorhynchus tshawytscha)

Presenter: Natasha Mckibben

Mentor: Charles Kimmel

Poster: 22

Major: Human Physiology

Research at Oregon State University has shown that two types of juvenile fish are present in a hatchery stock of spring run Chinook salmon derived from locations along the Upper Willamette River Basin in Oregon. The two groups differ in body morphology, agonistic behavior, growth, and whether they live at the surface or the bottom of the water column. However, they maintain the same diet and identical rearing conditions under captivity. The juveniles appear to resemble wild fish rearing in downstream and upstream sites in the Willamette River Basin respectively, and leave their home streams for the main stem of the river during the same time of year each wild group does. Based on understanding in another species, stickleback, I hypothesize that the fish growing at different depths also differ in their feeding strategies and, therefore, might exhibit changed jaw morphologies and mechanics, even though the genetic basis is unknown. To test this prediction, I quantitatively compared lower jaw bone morphologies in the two types of juveniles at the 50 mm stage. Supporting my hypothesis, I found significant differences between the groups in the shapes of both lower jaw bones, the angular-articular and the dentary. No difference was present in the bone sizes relative to body size. These results provide further evidence that early differences in behavior may be predictive of juvenile life history tactics.

Are Allometric Growth Patterns Consistent Throughout Development in Lower Jaw Bones of Chinook Salmon (Oncorhynchus tshawytscha)?

Presenter: Natasha Mckibben

Faculty Mentor: Charles Kimmel, Saywer Watson

Presentation Type: Poster 77

Primary Research Area: Science

Major: Human Physiology

In vertebrate species, bone morphology directly affects the function of the individual bone and the way it works within the skeletal system as a whole. Over the course of development, a variety of growth patterns are crucial in coordinating changes in bone morphology. Studying these changes in Chinook salmon, we hypothesized that the lower jaw bones, dentary and angular articular, grow allometrically, meaning that shape changes as a function of size. This study characterizes the growth pattern by quantitatively comparing bone shape of juvenile salmon using geometric morphometrics. We used two groups of juveniles at different ages, the first with an average length (fork length) of 48.5 mm and 85 mm for the second. We found that as the fish grows in length, the dentary becomes broader, while the posterior aspect of the angular articular rotates in a clockwise direction. These results show that growth between the two stages is allometric, supporting our hypothesis. We now can inquire whether the same allometric rules dictate the shape changes during other life history stages. To address this question we are examining both earlier and later time points of Chinook development. Studying such growth patterns across development can be used to evaluate how early growth patterns can impact the overall development of the individual and influence functionality, together resulting in constraints on evolution.

Severity of Cranial Phenotypes in Double Mutants Affected by the Interaction Between fras1 and ezh2 Genes

Presenter(s): Samuel Ahlquist – Human Physiology

Co presenter(s): Whitney Olivia

Faculty Mentor(s): Charles Kimmel

Poster 88

Research Area: Biology

This study assesses the interaction between two genes, fras1 and ezh2, in zebrafish craniofacial patterning. The Fras1 protein stabilizes facial development and is studied to understand Fraser syndrome in humans. Ezh2 is a protein subunit of a repressive complex that is important for early embryonic development and essential for tissue maintenance. Ezh2 is explored because of its role in controlling epigenetics, which is a background that can affect the expression of genes. Loss of function in these two genes individually results in mild reduction in the opercular bone (OP). We therefore hypothesized that the interaction between ezh2 and fras1 in double mutants will result in more severe reduction in OP bone size. Zebrafish embryos were stained to assess bone morphology and each set was phenotypically separated into WT, single, and double mutant groups using key phenotypes. The embryos will be genotypically verified through PCR in this ongoing study. As predicted, the double mutants displayed more severe reduction in the OP bone when compared to the single mutants. We concluded that the double mutants displayed a synergistic effect for the OP bone as the result appears more severe than both single mutants combined. This study suggests that changes in epigenetics, in this case loss of ezh2 function, influences the severity of the fras1 phenotype which could also influence the degree of severity of Fraser syndrome in humans.

Variation in Severity of Cartilaginous Fusions in fras1 Mutant Zebrafish Following Independent Transgene Insertions

Presenter(s): Alexander Wind

Co Presenter(s): Sam Ahlquist, Whitney Oliva

Faculty Mentor(s): Charles Kimmel

Poster 74

Session: Sciences

This study assesses the effect of the fli1a:gal4vp16 transgene insertion on fras1 mutant zebrafish. The Fras1 protein stabilizes craniofacial development and is studied to understand Fraser syndrome in humans. Fraser syndrome is a fatal disease that results in facial deformities and hearing loss. Zebrafish facial cartilages are homologous with human ear bones, making them good model organisms. The fras1 homozygous mutation causes fusions between the Meckel’s (m) and palatoquadrate (pq) cartilages in the zebrafish jaw. In previous experiments, we found that fras1 mutant embryos containing the transgene increased m-pq fusion severity. One hypothesis is that increased severity results from changes in host DNA sequences at the insertion site. An alternative hypothesis is that increased severity results from function of sequences within the transgene. In this study we hope to distinguish between these hypotheses by generating independent insertional lines with the expectation that the transgene has inserted in a unique location in each line. Mutant eggs were injected with the transgene and raised to score severity between transgene-containing and control groups. Polymerase Chain Reaction (PCR) can be used to verify the location of the independent insertions. As predicted by the second hypothesis, the majority of genetic lines with new transgene insertions exhibited increased m-pq severity, suggesting importance of the transgene sequences. This study gives us insight on the widespread variation of mutation in Fraser syndrome in humans to help us treat/prevent it, and helps us gain a better understanding of transgenic effects on mutants in general.