発がんと突然変異誘発のジャーナル

概要

Characterizing Pathogenic Enhancer Activity at Single-Cell Resolution to Study the Mechanism of Limb Malformation Disorders through a Novel Deep Learning Approach

Raghav Sriram

The Zone of Polarizing Activity (ZPA) Regulatory Sequence (ZRS) enhancer, a regulatory sequence found in numerous organisms, plays an important role in early embryonic limb development. ZRS controls the expression of the Sonic Hedgehog gene (Shh), and therefore early limb development in an organism as Shh has been shown to control the width of the limb bud by stimulating mesenchyme cell proliferation due to its ability to regulate the anterior- posterior length of the apical ectodermal ridge. Several transcription factors, acting as repressors or activators of the Shh gene, coordinate this limb development process in tandem with the ZRS enhancer. While the significance of normal ZRS activity is evident, this study looks deeper into the effects of pathogenic changes to the ZRS enhancer and the development of associated limb disorders such as Preaxial Polydactyly (PPD) by focusing on several aspects of ZRS regulation and its relation to Shh expression. This was accomplished by characterizing the expression of Shh and mCherry, an introduced luminescence gene regulated by ZRS, through single-cell RNA sequenced cells from a developing limb bud of a mouse embryo. Additionally, this study characterized specific transcription factors as potential repressors or activators of ZRS by determining TF enrichment or depletion in highly expressive Shh and mCherry cells. Classifying such TFs is vital in identifying the regulatory elements that control the formation of limb malformation disorders such as preaxial polydactyly and aid in the development of future therapeutic interventions.