Inducible and reversible in vivo protein degradation reveals T cell-intrinsic regulatory function of SHP-1
Year: 2026
Type: article
Abstract: Scientific problem Studying the acute, cell-intrinsic functions of signaling proteins in T cells has long been challenging. Existing constitutive or inducible gene deletion cannot overcome developmental adaptation that obscures true phenotypes. Similarly, RNA interference suffers from incomplete knockdown and off-target effects, and pharmacological inhibitors often lack selectivity. Thus, approaches allowing rapid, specific, and reversible protein depletion in defined cell populations in vivo are needed. Methods Here, we describe an Inducible and Reversible in vivo Protein Degradation (IRIP) system in T cells by combining auxin-induced degron technology with Cre-driver-mediated, lineage-specific expression of the plant E3 ligase adaptor OsTIR1. We first validated a minimal auxin-inducible degron tag (mAID-tag) inserted into the endogenous Ptpn6 locus encoding the tyrosine phosphatase SHP-1 in murine and human T cell lines. We then generated SHP-1 mAID knock-in mice and crossed them with mice carrying a floxed OsTIR1 construct in the Rosa26 locus, enabling Cre-mediated, T cell-specific, auxin-dependent SHP-1 degradation. We assessed SHP - 1 mAID protein degradation kinetics, specificity, reversibility, tissue restriction, and functional consequences. Results The mAID-tagged SHP-1 protein was expressed at wild-type levels and retained full signaling function. Auxin treatment induced complete degradation of SHP-1mAID protein restricted to cells co-expressing OsTIR1, with no loss of untagged SHP-1. Degradation was fully reversible, with SHP-1 protein restored to baseline within 2–3 hours of auxin withdrawal in vitro and 60 hours in vivo . Degradation was confined to the T cell lineage by restricting OsTIR1 expression through a T cell-specific Cre driver. Such targeting could be extended to additional cell populations using a ubiquitous Cre driver. Acute in vivo depletion of SHP-1 caused T cell hyperproliferation upon restimulation, which was reversed upon SHP-1 re-expression. Conclusion These findings demonstrate that SHP-1 restrains T cell proliferation in a manner that is direct, cell-intrinsic, acute, and independent of prior developmental conditioning. Our SHP-1 protein targeting approach also resolves a long-standing uncertainty in using the motheaten mouse line or the floxed Ptpn6 locus. Our IRIP system represents a versatile and broadly adaptable platform for dissecting acute physiological roles of proteins whose constitutive loss produces complex and difficult-to-interpret phenotypes.
Source: Frontiers in Immunology
Language: English
Subfield: Molecular Biology
Domain: Life Sciences
Open Access status: gold
Estimated APC paid: $3,801
OpenAlex ID: W7223121747