Biology subjects

Gomez, A. E.

Publications and source records attributed to Gomez, A. E..

3 recordsLinked to original sources

bioRxiv2024

Loss of cped1 does not affect bone and lean mass in zebrafish

Human genetic studies have nominated Cadherin-like and PC-esterase Domain-containing 1 (CPED1) as a candidate target gene mediating bone mineral density (BMD) and fracture risk heritability. Recent efforts to define the role of CPED1 in bone in mouse and human models have revealed complex alternative splicing and inconsistent results arising from gene targeting, making its function in bone difficult to interpret. To better understand the role of CPED1 in adult bone mass and morphology, we conducted a comprehensive genetic and phenotypic analysis of cped1 in zebrafish, an emerging model for bone and mineral research. We analyzed two different cped1 mutant lines and performed deep phenotyping to characterize more than 200 measures of adult vertebral, craniofacial, and lean tissue morphology. We also examined alternative splicing of zebrafish cped1 and gene expression in various cell/tissue types. Our studies fail to support an essential role of cped1 in adult zebrafish bone. Specifically, homozygous mutants for both cped1 mutant alleles, which are expected to result in loss-of-function and impact all cped1 isoforms, exhibited no significant differences in the measures examined when compared to their respective wildtype controls, suggesting that cped1 does not significantly contribute to these traits. We identified sequence differences in critical residues of the catalytic triad between the zebrafish and mouse orthologs of CPED1, suggesting that differences in key residues, as well as distinct alternative splicing, could underlie different functions of CPED1 orthologs in the two species. Our studies fail to support a requirement of cped1 in zebrafish bone and lean tissue, adding to evidence that variants at 7q31.31 can act independently of CPED1 to influence BMD and fracture risk. Lay summaryBone mineral density (BMD) is a key indicator for predicting and diagnosing osteoporosis and fracture risk, and it has been estimated that up to 89% of variation in BMD is determined by genetics. Multiple human genetics studies have nominated CPED1 as a potential gene underlying BMD and fracture risk heritability, however the function of CPED1 remains poorly understood. In this study, we examined the role of cped1 in bone by quantifying over 200 morphological measures of vertebral and craniofacial bone size, shape, and density in two different mutant lines of zebrafish in which cped1 function was reduced or eliminated. We also examined lean tissue mass because co-heritability of this trait with BMD has also been hypothesized to involve CPED1. Surprisingly, despite the loss of cped1 function, there were no significant differences between the mutant zebrafish and their respective controls. Our study therefore fails to support a role for cped1 in bone and lean tissue, suggesting that hereditary influence on BMD and fracture risk can occur independently of CPED1.

genetics↗

bioRxiv2024

Dominant nonsense mutations in efemp1 alter vertebral and craniofacial characteristics in adult zebrafish

Heritable Disorders of Connective Tissues (HDCT) are a heterogenous, pleiotropic group of conditions that broadly affect connective tissues. EFEMP1 is a member of the fibulin family of extracellular matrix (ECM) glycoproteins which is expressed in various human tissues. Individuals with EFEMP1 variants have recently been identified and appear to have Marfan-like characteristics. Clinical phenotypes of these individuals include hernias, advanced bone age, tall stature, myopia, joint laxity, and thin skin. EFEMP1-associated HDCTs have been identified in individuals with biallelic and monoallelic variants. There is an urgent need to better understand the role of EFEMP1 in regulating connective tissues including bone, and the pathophysiological mechanisms underlying the spectrum of genotype-phenotype relationships seen in EFEMP1-associated HDCTs. To investigate the role of EFEMP1 in developing and adult bone, we used CRISPR-based editing to generate two efemp1 zebrafish alleles encoding for premature termination codons (PTCs) predicted to delete or severely alter the fibulin-type domain. Both alleles exhibited similar phenotypes in juvenile and adult fish. In juvenile fish, we did not identify changes in body size or vertebral development. In adults, we found significant changes in body length, bone microarchitecture, and craniofacial measurements in both heterozygous and homozygous mutant fish. These results expand our understanding of the role of efemp1 in the skeleton and highlight the potential for dominant nonsense variants to play a role in manifestation of clinical phenotypes in EFEMP1-associated HDCTs.

genetics↗

bioRxiv2022

Systematic analysis of cilia characteristics and Hedgehog signaling in five immortal cell lines

Dysfunction of the primary cilium, a microtubule-based signaling organelle, leads to genetic conditions called ciliopathies. Hedgehog (Hh) signaling is mediated by the primary cilium in vertebrates and is therefore implicated in ciliopathies; however, it is not clear which immortal cell lines are the most appropriate for modeling pathway response in human disease; therefore we systematically evaluated Hh in five commercially available, immortal mammalian cell lines: ARPE-19, HEK293T, hTERT RPE-1, NIH/3T3, and SH-SY5Y. All of the cell lines ciliated adequately for our subsequent experiments, except for SH-SY5Y which were excluded from further analysis. hTERT RPE-1 and NIH/3T3 cells relocalized Hh pathway components Smoothened (SMO) and GPR161 and upregulated Hh target genes in response to pathway stimulation. In contrast, pathway stimulation did not induce target gene expression in ARPE-19 and HEK293T cells, despite SMO and GPR161 relocalization. These data indicate that human hTERT RPE-1 cells and murine NIH/3T3 cells, but not ARPE-19 and HEK293T cells, are suitable for modeling the role of Hh signaling in ciliopathies.

molecular biology↗