Nonsense Codons Trigger an RNA Partitioning Shift

T-cell receptor-β (TCRβ) genes naturally acquire premature termination codons (PTCs) as a result of programmed gene rearrangements. PTC-bearing TCRβ transcripts are dramatically down-regulated to protect T-cells from the deleterious effects of the truncated proteins that would otherwise be produced. Here we provide evidence that two responses collaborate to elicit this dramatic down-regulation. One is rapid mRNA decay triggered by the nonsense-mediated decay (NMD) RNA surveillance pathway. We demonstrate that this occurs in highly purified nuclei lacking detectable levels of three different cytoplasmic markers, but containing an outer nuclear membrane marker, suggesting that decay occurs either in the nucleoplasm or at the outer nuclear membrane. The second response is a dramatic partitioning shift in the nuclear fraction-to-cytoplasmic fraction mRNA ratio that results in few TCRβ transcripts escaping to the cytoplasmic fraction of cells. Analysis of TCRβ mRNA kinetics after either transcriptional repression or induction suggested that this nonsense codon-induced partitioning shift (NIPS) response is not the result of cytoplasmic NMD but instead reflects retention of PTC+ TCRβ mRNA in the nuclear fraction of cells. We identified TCRβ sequences crucial for NIPS but found that NIPS is not exclusively a property of TCRβ transcripts, and we identified non-TCRβ sequences that elicit NIPS. RNA interference experiments indicated that NIPS depends on the NMD factors UPF1 and eIF4AIII but not the NMD factor UPF3B. We propose that NIPS collaborates with NMD to retain and degrade a subset of PTC+ transcripts at the outer nuclear membrane and/or within the nucleoplasm.