Human TRPS1 Gene: Endogenous Expression and Cis-acting Regulatory Control

Abstract

Background: Early embryological events and patterning are tightly controlled and orchestrated by the precise expression of developmental genes both in space and time. Disruptions in gene regulation and irregular expression results in developmental disorders. One such gene TRPS1, serves as the causative gene for tricho-rhino phalangeal syndrome, known for its craniofacial and skeletal abnormalities. The Trps1 gene encodes a protein that represses Wnt signaling through strong interactions with Wnt signaling inhibitors. The identification of genomic cisacting regulatory sequences governing Trps1 expression is crucial for understanding its role in embryogenesis. Nevertheless, to date, no investigations have been conducted concerning these aspects of Trps1. Results: To identify deeply conserved noncoding elements (CNEs) within the Trps1 locus, we employed a comparative genomics approach, utilizing slowly evolving fish such as coelacanth and spotted gar. These analysis resulted in the identification of eight CNEs in the intronic region of the Trps1 gene. Comparative synteny along with Hi-C interaction analysis confirmed strong association of these CNEs with TRPS1 gene. Functional characterization of these CNEs in zebrafish revealed their regulatory potential in various tissues, including pectoral fins, heart, and pharyngeal arches. The redundant expression of multiple CNEs in each of these tissues represent the precise expression and regulation of trps1 in the development of respective tissues. RNA in-situ hybridization experiments revealed concordance between the reporter expression pattern induced by the identified set of CNEs and the spatial expression pattern of the trps1 gene in zebrafish. Comparative in vivo data from zebrafish and mice for CNE7/hs919 revealed conserved functions of these enhancers. Each of these eight CNEs was further investigated in cell line-based reporter assays, revealing their repressive potential predominantly. The repressive potential of these CNEs was different in different type of cell lines. This differential activity appears to be corelated with wnt signaling. As each cell type responds to wnt signaling depending on the type of ligands or receptors it possesses. Taken together, in vivo and in vitro assays suggest a context-dependent dual functionality for the identified set of Trps1-associated CNE enhancers. Conclusion: This functionally characterized set of CNE-enhancers will contribute to a more comprehensive understanding of the developmental roles of Trps1 and can aid in the identification of non-coding DNA variants associated with human diseases.