Differential developmental regulation and functional effects on pre-TCR surface expression of human pT alpha(a) and pT alpha(b) spliced isoforms

Functional rearrangement at the TCR beta locus leads to surface expression on developing pre-T cells of a pre-TCR complex composed of the TCR beta -ch ain paired with the invariant pre-TCR alpha (pT alpha) chain and associated with CD3 components. Pre-TCR signaling triggers the expansion and further differentiation of pre-T cells into TCR alpha beta mature T cells, a proces s known as beta selection. Besides the conventional pT alpha transcript (te rmed pT alpha (a)), a second, alternative spliced, isoform of the pT alpha gene (pT alpha (b)) has been described, whose developmental relevance remai ns unknown. In this study, phenotypic, biochemical, and functional evidence is provided that only pT alpha (a) is capable of inducing surface expressi on of a CD3-associated pre-TCR complex, which seems spontaneously recruited into lipid rafts, while pT alpha (b) pairs with and retains TCR beta intra cellularly. In addition, by using real-time quantitative RT-PCR approaches, we show that expression of pT alpha (a) and pT alpha (b) mRNA spliced prod ucts is differentially regulated along human intrathymic development, so th at pT alpha (b) transcriptional onset is developmentally delayed, but beta selection results in simultaneous shutdown of both isoforms, with a relativ e increase of pT alpha (b) transcripts in beta -selected vs nonselected pre -T cells in vivo. Relative increase of pT alpha (b) is also shown to occur upon pre-T cell activation in vitro. Taken together, our data illustrate th at transcriptional regulation of pT alpha limits developmental expression o f human pre-TCR to intrathymic stages surrounding beta selection, and are c ompatible with a role for pT alpha (b) in forming an intracellular TCR beta -pT alpha (b) complex- that may be responsible for limiting surface expres sion of a pT alpha (a)-containing pre-TCR and/or may be competent to signal from a subcellular compartment.