期刊
ACS SYNTHETIC BIOLOGY
卷 3, 期 5, 页码 264-272出版社
AMER CHEMICAL SOC
DOI: 10.1021/sb400053b
关键词
morphogen; synthetic patterning; MDCK; HGF; NK4
资金
- GABBA
- Portuguese Fundacao para a Ciencia e Tecnologia (FCT), Studentship [BD/15897/2005]
- La Caixa PhD Fellowships
- CONICET (Argentina)
- FP7 ERG [201249 ZINC-HUBS]
- Ministerio de Ciencia e Innovacion [BFU2010-17953]
- MEC-EMBL
Engineering spatial patterning in mammalian cells, employing entirely genetically encoded components, requires solving several problems. These include how to code secreted activator or inhibitor molecules and how to send concentration-dependent signals to neighboring cells, to control gene expression. The Madin-Darby Canine Kidney (MDCK) cell line is a potential engineering scaffold as it forms hollow spheres (cysts) in 3D culture and tubulates in response to extracellular hepatocyte growth factor (HGF). We first aimed to graft a synthetic patterning system onto single developing MDCK cysts. We therefore developed a new localized transfection method to engineer distinct sender and receiver regions. A stable reporter line enabled reversible EGFP activation by HGF and modulation by a secreted repressor (a truncated HGF variant, NK4). By expanding the scale to wide fields of cysts, we generated morphogen diffusion gradients, controlling reporter gene expression. Together, these components provide a toolkit for engineering cell-cell communication networks in 3D cell culture.
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