T-cell receptor retrogenic mice: a rapid, flexible alternative to T-cell receptor transgenic mice

Abstract

The T-cell receptor (TCR) is unique in its complexity. It determines not only positive (life) and negative (death) selection in the thymus, but also mediates proliferation, anergy, differentiation, cytotoxicity and cytokine production in the periphery. Through its association with six CD3 signalling chains (εγ, δε and ζζ), the TCR is capable of recognizing an extensive variety of antigenic peptides, from both pathogens and self-antigens, and translating these interactions into multiple signalling pathways that mediate diverse T-cell developmental and functional responses. The analysis of TCR biology has been revolutionized by the development of TCR transgenic mice, which express a single clonotypic T-cell population, with diverse specificities and genetic backgrounds. However, they are time consuming to generate and characterize, limiting the analysis of large numbers of TCR over a short period of time in multiple genetic backgrounds. The recent development of TCR retrogenic technology resolves these limitations and could in time have a similarly important impact on our understanding of T-cell development and function. In this review, we will discuss the advantages and limitations of retrogenic technology compared with the generation and use of TCR transgenic mice for studying all aspects of T-cell biology.

Figure 1

Picornavirus-like ‘self-cleaving’ 2A peptide-linked, multi-cistronic retroviral vector. (a) Schematic of the retroviral vector and 2A-linked T-cell receptor (TCR) -α : TCR-β construct. The murine stem cell virus long terminal repeat (MSCV LTR) promoter is used to express the 2A-linked construct. The internal ribosomal entry site (IRES) is used to direct translation of the fluorescent protein. (b) Amino acid sequence of the 2A regions of foot-and-mouth disease virus (F2A), equine rhinitis A virus (E2A), Thosea asigna virus (T2A) and porcine teschovirus-1 (P2A). Conserved residues are boxed. The cleavage point between the 2A and 2B peptides, and hence the N- and C-terminal cistrons, is indicated by the arrow.

Figure 2

Flow chart of steps required to create T-cell receptor retrogenic mice. The standard time required for each step is indicated. 5FU, 5-fluorouracil; IL-3, interleukin-3; i.p., intraperitoneal; i.v., intravenous; SCF, stem cell factor.