Advances in molecular techniques have allowed for genome-wide studies to examine which genes are associated with many different human diseases, including those of the skeletal system. In one such study, mutations in the human SOST gene were linked to the skeletal disease Sclerosteosis 1, which presents as hyperostosis in the skull and long bones of the axial skeleton. Functional SOST genes influence the production of sclerostin, a protein which inhibits osteoblastic bone formation. The homologous gene in zebrafish is hypothesized to have a similar function based on ongoing work examining the axial skeleton in mutant zebrafish. Our goal for the current study is to test how the zebrafish cranial skeleton is affected by the SOST gene. I used the open-source 3DSlicer software to place landmarks on micro-CT scans of 27 zebrafish (9 SOST mutants, 9 SOST heterozygotes, and 9 wildtype fish) from the same clutch. I also used 3DSlicer to generate digital models of the cranial skeleton as well as to place 308 pseudolandmark points on the models. From here, I used geometric morphometric methods implemented in R to analyze the complex shape differences between the three groups. Preliminary results suggest that SOST mutants have narrower posterior cranial skeletons than heterozygous or wildtype fish, and that groups may vary in their degree of cranial asymmetry. In addition to quantifying the effect of SOST on zebrafish cranial morphology, this study is part of a larger project to establish a baseline craniofacial analysis method, and create a screening tool for examining genotype-phenotype relationships in genes associated with human skeletal diseases within zebrafish models.

Genome Wide Association Studies enable researchers to identify which genes are associated with human diseases. Once genes are identified, gene editing of zebrafish models allows researchers to further examine the link between genotype and phenotype, with the long-term goal of treatment development. One such gene is MEOX1, which encodes for transcription factor Mesenchyme Homeobox 1. MEOX1 mutations in humans have been associated with Klippel-Feil syndrome, a congenital disorder with the most frequent symptoms being a shortened neck caused by the fusion of cervical vertebrae, leading to a decreased range of motion. Previous studies have established that zebrafish with loss-of-function meox1 mutations present with similar phenotypes in the axial skeleton. However, fusion of the vertebral column could also impact other aspects of fish morphology. The goal of this study is to test if a semi-automated screening tool can be used to quantify craniofacial variations in meox1 crispant zebrafish. Here, I utilized micro-CT scans of CRISPR-modified zebrafish with meox1 mutations (n=12) and compared them to wild type zebrafish (N=12) from the same clutch. I used 3D Slicer to manually landmark 23 major anatomical points on each individual wildtype and crispant zebrafish. I then used a semi-automated process to distribute pseudolandmarks on the surface of each zebrafish. Anatomical differences between the groups were quantified using a geometric morphometrics approach. Preliminary results show that meox1 mutations are associated with a wider posterior section of the skull and a shorter skull length. There were also differences found in the degree of asymmetry between groups. This last result in particular aligns with previous human studies of Klippel-Feil syndrome. The findings from this investigation are important for the understanding of how diseases from meox1 mutations present clinically as well as the testing of a semiautomated pipeline that will be used as a screening tool for crispant zebrafish.