Stem Loop Mediated Transgene Modulation in Human T Cells

ACS Synth Biol. 2024 Dec 20;13(12):3897-3907. doi: 10.1021/acssynbio.4c00152. Epub 2024 Dec 6.

Abstract

Controlling gene expression is useful for many applications, but current methods often require external user inputs, such as the addition of a drug. We present an alternative approach using cell-autonomous triggers based on RNA stem loop structures in the 3' untranslated regions (UTRs) of mRNA. These stem loops are targeted by the RNA binding proteins Regnase-1 and Roquin-1, allowing us to program stimulation-induced transgene regulation in primary human T cells. By incorporating engineered stem loops into the 3' UTRs of transgenes, we achieved transgene repression through Regnase-1 and Roquin-1 activity, dynamic upregulation upon stimulation, and orthogonal tunability. To demonstrate the utility of this system, we employed it to modulate payloads in CAR-T cells. Our findings highlight the potential of leveraging endogenous regulatory machinery in T cells for transgene regulation and suggest RNA structure as a valuable layer for regulatory modulation.

MeSH terms

  • 3' Untranslated Regions* / genetics
  • Gene Expression Regulation
  • Humans
  • RNA, Messenger / genetics
  • RNA, Messenger / metabolism
  • RNA-Binding Proteins / genetics
  • RNA-Binding Proteins / metabolism
  • T-Lymphocytes* / metabolism
  • Transgenes*

Substances

  • 3' Untranslated Regions
  • RNA-Binding Proteins
  • RNA, Messenger