Engineering modular intracellular protein sensor-actuator devices

Velia Siciliano,Blandine Monel,Kiera L Clayton,Jin Huh,Bruce D Walker,Jacob Beal,Annemarie Mckeon,Liliana Wroblewska,Ron Weiss,Breanna Diandreth

doi:10.1038/s41467-018-03984-5

Abstract

Understanding and reshaping cellular behaviors with synthetic gene networks requires the ability to sense and respond to changes in the intracellular environment. Intracellular proteins are involved in almost all cellular processes, and thus can provide important information about changes in cellular conditions such as infections, mutations, or disease states. Here we report the design of a modular platform for intrabody-based protein sensing-actuation devices with transcriptional output triggered by detection of intracellular proteins in mammalian cells. We demonstrate reporter activation response (fluorescence, apoptotic gene) to proteins involved in hepatitis C virus (HCV) infection, human immunodeficiency virus (HIV) infection, and Huntington’s disease, and show sensor-based interference with HIV-1 downregulation of HLA-I in infected T cells. Our method provides a means to link varying cellular conditions with robust control of cellular behavior for scientific and therapeutic applications.

Highlights

Understanding and reshaping cellular behaviors with synthetic gene networks requires the ability to sense and respond to changes in the intracellular environment
We report on our development of modular sensing-actuation devices that initiate programmed transcriptional response when detecting target intracellular proteins in mammalian cells
One intrabody is fused at the N-terminus to a membrane-tethered fluorescent tag and at the Cterminus to a Tobacco Etch Virus (TEV) cleavage site (TCS) and to a GAL4-VP16 transcriptional activator, forming a chimeric protein sequestered in the cytosol

Summary

Introduction

Understanding and reshaping cellular behaviors with synthetic gene networks requires the ability to sense and respond to changes in the intracellular environment. While circuits that tune output gene expression in response to specific miRNA signatures have been demonstrated[6], a customizable framework to link intracellular protein sensing to programmed cellular responses still lags behind To this end, we report on our development of modular sensing-actuation devices that initiate programmed transcriptional response when detecting target intracellular proteins in mammalian cells. We demonstrate the modularity of this platform by creating devices that sense four different proteins associated with diseases and respond with either fluorescent reporter activation or biological activity where applicable (cell death or receptor downregulation) We envision these devices will find use in enabling protein-responsive therapeutic gene circuits as well as in basic research

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