To evaluate the effect of increased expression of genes involved in flower pigmentation, additional dihydroflavonol-4-reductase (DFR) or chalcone synthase (CHS) genes were transferred to petunia. In most transformants, the increased expression had no measurable effect on floral pigmentation. Surprisingly, however, in up to 25% of the transformants, a reduced floral pigmentation, accompanied by a dramatic reduction of DFR or CHS gene expression, respectively, was observed. This phenomenon was obtained with both chimeric gene constructs and intact CHS genomic clones. The reduction in gene expression was independent of the promoter driving transcription of the transgene and involved both the endogenous gene and the homologous transgene. The gene-specific collapse in expression was obtained even after introduction of only a single gene copy. The similarity between the sense transformants and regulatory CHS mutants suggests that this mechanism of gene silencing may operate in naturally occurring regulatory circuits.

No abstract

. 11. Complementary DNAs encoding the short forms of G␣ s WT and G␣ s * (9) were subcloned into pUAST (7) and used to transform flies [A. Spradling, in Drosophila: A Practical Approach, D. Roberts, Ed. (IRL Press, Oxford, 1986), pp. 75 -197]. The UAS-G␣ s lines were verified by Msc I digestion of polymerase chain reaction-amplified G␣ s transgenes to detect the presence of a site found in G␣ s WT but lacking as a result of the Q215L mutation. 12. 238Y and 201Y are described in (17) and C232 in (18). C747 shows identical expression to that of C772 (17) (24). C309 is described by J. D. Armstrong [thesis, University of Glasgow, Scotland (1995)]. The O'Kane laboratory screened 500 P-GAL4 lines to identify OK66, OK86, OK62, OK107, OK348, and OK415. 13. T. Tully and W. Quinn, J. Comp. Physiol. 157, 263 (1985); T. Tully, T. Preat, S. Boynton, M. Del Vecchio, Cell 79, 35 (1994). 14. In total, we analyzed behaviorally 16 P-GAL4 insertions that expressed to some degree in MBs. Flies with four insertions (OK66, OK86, KL65, and KL107) showed reduced learning in the absence of expression of constitutively activated G␣ s *, most likely because of genetic background differences. These were not tested further. Flies with eight insertions showed defects in olfactory acuity (30Y, C35, OK62, OK107, C302, OK415, and C532) or shock reactivity (C772), as transheterozygotes with UAS-G␣ s *. These were eliminated from further behavioral testing. Flies with the remaining four P-GAL4 insertions (201Y, 238Y, C309, and C747) showed disrupted learning but normal olfactory acuity and shock reactivity, as transheterozygotes with at least one UAS-G␣ s * insertion. We characterized a total of two P-GAL4 insertions (C232 and OK348) that expressed to some degree in the CC. 15. We suggest that olfactory learning results from G sdependent convergence of the conditional stimulus (CS) and unconditional stimulus (US) in the MBs. This belief is reinforced by the observation that expression of G␣ s * in the CC does not affect learning.

In this paper we report the isolation of cDNA clones encoding the flavonoid-biosynthetic enzyme chalcone flavanone isomerase (CHI)

In this paper we describe the organization and expression of the genes encoding the flavonoid-biosynthetic enzyme dihydroflavonol-4-reductase (DFR) in Petunia hybrida. A nearly full-size DFR cDNA clone (1.5 kb), isolated from a corolla-specific cDNA library was compared at the nucleotide level with the pallida gene from Antirrhinum majus and at the amino acid level with enzymes encoded by the pallida gene and the A1 gene from Zea mays. The P. hybrida and A. majus DFR genes transcribed in flowers contain 5 introns, at identical positions; the three introns of the A1 gene from Z. mays coincide with the first three introns of the other two species. P. hybrida line V30 harbours three DFR genes (A, B, C) which were mapped by RFLP analysis on three different chromosomes (IV, II and VI respectively). Steady-state levels of DFR mRNA in the line V30 follow the same pattern during development as chalcone synthase (CHS) and chalcone flavanone isomerase (CHI) mRNA. Six mutants that accumulate dihydroflavonols in mature flowers were subjected to Northern blot analysis for the presence of DFR mRNA. Five of these mutants lack detectable levels of DFR mRNA. Four of these five also show drastically reduced levels of activity for the enzyme UDPG: flavonoid-3-O-glucosyltransferase (UFGT), which carries out the next step in flavonoid biosynthesis; these mutants might be considered as containing lesions in regulatory genes, controlling the expression of the structural genes in this part of the flavonoid biosynthetic pathway. Only the an6 mutant shows no detectable DFR mRNA but a wild-type level for UFGT activity. Since both an6 and DFR-A are located on chromosome IV and DFR-A is transcribed in floral tissues, it is postulated that the An6 locus contains the DFR structural gene. The an9 mutant shows a wild-type level of DFR mRNA and a wild-type UFGT activity.