Tissue morphogenesis in plants requires communication between cells, a process involving the trafficking of molecules through plasmodesmata (PD). PD conductivity is regulated by endogenous and exogenous signals. However, the underlying signaling mechanisms remain enigmatic. In Arabidopsis, signal transduction mediated by the receptor-like kinase STRUBBELIG (SUB) contributes to inter-cell layer signaling during tissue morphogenesis. Previous analysis has revealed that SUB acts non-cell-autonomously suggesting that SUB controls tissue morphogenesis by participating in the formation or propagation of a downstream mobile signal. A genetic screen identified QUIRKY (QKY), encoding a predicted membrane-anchored C2-domain protein, as a component of SUB signaling. Here, we provide further insight into the role of QKY in this process. We show that like SUB, QKY exhibits non-cell-autonomy when expressed in a tissue-specific manner and that non-autonomy of QKY extends across several cells. In addition, we report on localization studies indicating that QKY and SUB localize to PD but independently of each other. FRET-FLIM analysis suggests that SUB and QKY are in close contact at PD in vivo. We propose a model where SUB and QKY interact at PD to promote tissue morphogenesis, thereby linking RLK-dependent signal transduction and intercellular communication mediated by PD.

The crystal structure of recombinant human GTP cyclohydrolase I was solved by Patterson search methods by using the coordinates of the Escherichia coli enzyme as a model. The human as well as bacterial enzyme were shown to contain an essential zinc ion coordinated to a His side chain and two thiol groups in each active site of the homodecameric enzymes that had escaped detection during earlier studies of the E. coli enzyme. The zinc ion is proposed to generate a hydroxyl nucleophile for attack of imidazole ring carbon atom eight of the substrate, GTP. It may also be involved in the hydrolytic release of formate from the intermediate, 2-amino-5-formylamino-6-ribosylamino-4(3H)-pyrimidinone 5-triphosphate, and in the consecutive Amadori rearrangement of the ribosyl moiety.

GTP cyclohydrolase I catalyzes a ring expansion affording dihydroneopterin triphosphate from GTP. [1,2,3,4,5-13 C 5 ,2-2 H 1 ]GTP was prepared enzymatically from [U-13 C 6 ]glucose for use as enzyme substrate. Multinuclear NMR experiments showed that the reaction catalyzed by GTP cyclohydrolase I involves the release of a proton from C-2 of GTP that is exchanged with the bulk solvent. Subsequently, a proton is reintroduced stereospecifically from the bulk solvent. This is in line with an Amadori rearrangement mechanism. The proton introduced from solvent occupies the pro-7R position in the enzyme product. The data also confirm that the reaction catalyzed by pyruvoyltetrahydropterin synthase results in the incorporation of solvent protons into positions C-6 and C-3 of the enzyme product. On the other hand, the reaction catalyzed by sepiapterin reductase does not involve any detectable incorporation of solvent protons into tetrahydrobiopterin.Pteridines serve as cofactors for a variety of enzyme-catalyzed reactions. Specifically, tetrahydrofolate (in bacteria and eukaryotic organisms) and tetrahydromethanopterin (in archaea) mediate the transfer of one-carbon fragments, tetrahydrobiopterin (BH 4 ) 1 is implicated in the hydroxylation of aromatic amino acids and the formation of nitric oxide in animals (1, 2), and molybdopterin is required as cofactor by a variety of redox enzymes, e.g. xanthine dehydrogenase. The metabolic roles of these cofactors have been reviewed repeatedly (3-6).The formation of pterins by ring expansion of guanosine, including an Amadori rearrangement of the ribose moiety, was first suggested by Weygand et al. (7) on basis of in vivo studies using 14 C-labeled precursors. Subsequent studies by Brown and Burg (8) and by Shiota et al. (9) showed that the first committed step in the biosynthesis of tetrahydrofolate and BH 4 is catalyzed by the enzyme GTP cyclohydrolase I. More specifically, C-8 of GTP (Fig. 1, compound 1) is released as formate, carbon atoms 1Ј and 2Ј of the ribose moiety are utilized for the formation of the dihydropyrazine ring, and carbon atoms 3Ј-5Ј of GTP afford the position 6 side chain of dihydroneopterin triphosphate (NH 2 TP) (Fig. 1, compound 2) (for a review, see Ref.3).The product of GTP cyclohydrolase I, NH 2 TP, is converted to BH 4 (compound 4) by the consecutive action of pyruvoyltetrahydropterin synthase (PPH 4 synthase) and sepiapterin reductase (10 -12). PPH 4 synthase catalyzes the elimination of triphosphate from NH 2 TP as well as a series of tautomerization reactions that are conducive to the formation of a tetrahydropterin from the dihydropterin substrate. Both carbonyl groups of the resulting pyruvoyltetrahydropterin (PPH 4 , compound 3) are subsequently reduced by the action of sepiapterin reductase.The three-dimensional structures of GTP cyclohydrolase I from Escherichia coli (13, 14), PPH 4 synthase from rat (15, 16), and sepiapterin reductase (17) from mouse have been determined by x-ray crystallography. The folding patterns of GTP cyclohydrolase I ...

Two types of genomic DNA hybridizing with a chalcone synthase cDNA were isolated from cell suspension cultures of parsley (Petroselinum crispum cv. Mooskrause) and cloned in lambda EMBL4. Their fragmentation patterns with several common restriction enzymes were identical, except for the occurrence of a 927 base pair insertion in one type relative to the other. This insertion is located 538 base pairs upstream of the first of two transcription start sites and has characteristic features of a transposable element. The two types of cloned DNA most likely represent two alleles of a chalcone synthase gene occurring in one copy per haploid parsley genome. The nucleotide sequence and exon-intron structure of the larger allele were determined. Analysis of plants either heterozygous or homozygous with respect to the chalcone synthase gene revealed that both allelic forms were expressed and activated by UV light.