Gene expression in the NMD- context. (a) Variations in the level of expression of YALI0C23496g splicing variants as a function of NMD context. RT-PCR products from spliced (S) and unspliced transcripts (intron 1 retained, R) from wild-type strains (WT) and NMD mutants (NMD-). Wild-type strains are E150 (lane 1) and PO1d (lane 2). NMD- strains are two independent knockouts of UPF1 (lane 3, upf1::LEU2 clone 7; lane 4, upf1::LEU2 clone C) and one UPF2 knockout (lane 5: upf2::LEU2 clone 7). The intensity of the unspliced transcripts is much stronger in the mutant strains. (b) Expression of the different transcripts of the Y. lipolytica YRA1 gene. Northern blot of total RNA of wild-type (WT) strain PO1d (lane 1) and NMD- mutant strains upf1::LEU2 clone 7 (lane 2), upf2::LEU2 clone 7 (lane 3), upf1::URA3 upf2::LEU2 (lane 4), xrn1::LEU2 (lane 5). The exon probe binding to exons 1 and 3 reveals the spliced transcript (S) in all strains and an additional splicing variant in NMD- mutants only. This variant corresponds to the retention of intron 1 (R). Hybridization with intron 1 confirmed that this intron is retained only in NMD- mutants, whereas it is efficiently spliced out in PO1d and xrn1- mutants.