Figure 9

3' UTRs harbor cis -acting elements that dictate specific mRNA fates. (a) Experimental design. Plasmids encoding firefly-luciferase (F-luc) or Renilla-luciferase (R-luc) driven by early zygotic promoters were co-injected into embryos 0 to 1 h AEL (stages 1 to 2). Embryos were aged at 25°C for 4 h and homogenized in lysate buffer. Luciferase activities in lysates were quantified through luminometry. We analyzed 12 to 16 embryos for each reporter construct. (b) mRNA expression of endogenous genes: scute and sisA promoters support expression in early embryos [90] (see (c)); 3' UTRs of stable α-tubulin 84B (α-tub) and unstable cortex mRNAs were tested for their effect on luciferase expression (see (c,d)). Median microarray expression levels for each time point are shown (compare Figure 1). (c) Reporter gene construction. 3' UTRs of stable α-tub and unstable cortex mRNAs were coupled to coding sequences for F-luc; all DNA constructs share a SV40 terminator sequence (SV40 pA). (d) Reporter gene activity. Average median activities (ratio F-luc/R-luc) and standard error of the mean for three independent, biological replicates are shown (12 to 16 embryos analyzed for each replicate). A statistical test (two-tailed Mann-Whitney) for each replicate consistently showed a lack of significant changes in luciferase activity for the α-tub reporter and significantly lower levels for the cortex 3' UTR reporter. N.s., not significant.