Strigolactones are a recently discovered class of plant hormone involved in branching, leaf senescence, root development, and plant-microbe interactions. They are carotenoid-derived lactones, synthesized in the roots and transported acropetally to modulate axillary bud outgrowth (i.e., branching). However, a receptor for strigolactones has not been identified. We have identified the DAD2 gene from petunia, an ortholog of the rice and Arabidopsis D14 genes, and present evidence for its roles in strigolactone perception and signaling. DAD2 acts in the strigolactone pathway, and the dad2 mutant is insensitive to the strigolactone analog GR24. The crystal structure of DAD2 reveals an α/β hydrolase fold containing a canonical catalytic triad with a large internal cavity capable of accommodating strigolactones. In the presence of GR24 DAD2 interacts with PhMAX2A, a central component of strigolactone signaling, in a GR24 concentration-dependent manner. DAD2 can hydrolyze GR24, with mutants of the catalytic triad abolishing both this activity and the ability of DAD2 to interact with PhMAX2A. The hydrolysis products can neither stimulate the protein-protein interaction nor modulate branching. These observations suggest that DAD2 acts to bind the mobile strigolactone signal and then interacts with PhMAX2A during catalysis to initiate an SCF-mediated signal transduction pathway.

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Understanding direct salt effects on protein crystal polymorphism is addressed by comparing different crystal forms (triclinic, monoclinic, tetragonal and orthorhombic) for hen, turkey, bob white quail and human lysozymes. Four new structures of hen egg‐white lysozyme are reported: crystals grown in the presence of NapTS diffracted to 1.85 Å, of NaI to 1.6 Å, of NaNO3 to 1.45 Å and of KSCN to 1.63 Å. These new structures are compared with previously published structures in order to draw a mapping of the surface of different lysozymes interacting with monovalent anions, such as nitrate, chloride, iodide, bromide and thiocyanate. An analysis of the structural sites of these anions in the various lysozyme structures is presented. This study shows common anion sites whatever the crystal form and the chemical nature of anions, while others seem specific to a given geometry and a particular charge environment induced by the crystal packing.

Cold storage of tubers of potato (Solanum tuberosum L.) compromises tuber quality in many cultivars by the accumulation of hexose sugars in a process called cold-induced sweetening. This is caused by the breakdown of starch to sucrose, which is cleaved to glucose and fructose by vacuolar acid invertase. During processing of affected tubers, the high temperatures involved in baking and frying cause the Maillard reaction between reducing sugars and free amino acids, resulting in the accumulation of acrylamide. cDNA clones with deduced proteins homologous to known invertase inhibitors were isolated and the two most abundant forms, termed INH1 and INH2, were shown to possess apoplastic and vacuolar localization, respectively. The INH2 gene showed developmentally regulated alternative splicing, so, in addition to the INH2α transcript encoding the full-length protein, two hybrid mRNAs (INH2β*A and INH2β*B) that encoded deduced vacuolar invertase inhibitors with divergent C-termini were detected, the result of mRNA splicing of an upstream region of INH2 to a downstream region of INH1. Hybrid RNAs are common in animals, where they may add to the diversity of the proteome, but are rarely described in plants. During cold storage, INH2α and the hybrid INH2β mRNAs accumulated to higher abundance in cultivars resistant to cold-induced sweetening than in susceptible cultivars. Increased amounts of invertase inhibitor may contribute to the suppression of acid invertase activity and prevent cleavage of sucrose. Evidence for increased RNA splicing activity was detected in several resistant lines, a mechanism that in some circumstances may generate a range of proteins with additional functional capacity to aid adaptability.

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SUMMARYThe 'fruity' attributes of ripe apples (Malus 3 domestica) arise from our perception of a combination of volatile ester compounds. Phenotypic variability in ester production was investigated using a segregating population from a 'Royal Gala' (RG; high ester production) 3 'Granny Smith' (GS; low ester production) cross, as well as in transgenic RG plants in which expression of the alcohol acyl transferase 1 (AAT1) gene was reduced. In the RG 3 GS population, 46 quantitative trait loci (QTLs) for the production of esters and alcohols were identified on 15 linkage groups (LGs). The major QTL for 35 individual compounds was positioned on LG2 and co-located with AAT1. Multiple AAT1 gene variants were identified in RG and GS, but only two (AAT1-RGa and AAT1-GSa) were functional. AAT1-RGa and AAT1-GSa were both highly expressed in the cortex and skin of ripe fruit, but AAT1 protein was observed mainly in the skin. Transgenic RG specifically reduced in AAT1 expression showed reduced levels of most key esters in ripe fruit. Differences in the ripe fruit aroma could be perceived by sensory analysis. The transgenic lines also showed altered ratios of biosynthetic precursor alcohols and aldehydes, and expression of a number of ester biosynthetic genes increased, presumably in response to the increased substrate pool. These results indicate that the AAT1 locus is critical for the biosynthesis of esters contributing to a 'ripe apple' flavour.

Type III secretion (TTS) systems are used by many Gram-negative pathogens to inject virulence proteins into the cells of their hosts. Several of these virulence effectors require TTS chaperones that maintain them in a secretion-competent state. Whereas most chaperones bind only one effector, Spa15 from the human pathogen Shigella flexneri and homologous chaperones bind several seemingly unrelated effectors, and were proposed to form a special subgroup. Its 1.8 Å crystal structure confirms this specific classification, showing that Spa15 has the same fold as other TTS effector chaperones, but forms a different dimer. The presence of hydrophobic sites on the Spa15 surface suggests that the different Spa15 effectors all possess similar structural elements that can bind these sites. Furthermore, the Spa15 structure reveals larger structural differences between class I chaperones than previously anticipated, which does not support the hypothesis that chaperone-effector complexes are structurally conserved and function as three-dimensional secretion signals.

The structure of a monoclinic form of bovine pancreatic trypsin inhibitor (BPTI) crystallized from a thiocyanate solution has been determined and re®ned at 2.7 A Ê resolution. The space group is P2 1 with a = 71.56, b = 73.83, c = 64.47 A Ê , = 93.9and Z = 20. The ten independent molecules were located by a multi-body molecular-replacement search as developed in the AMoRe program, starting from a single monomer model (PDB code: 6PTI). The molecular arrangement of the subunits is a decamer resulting from the combination of two orthogonal ®vefold and twofold noncrystallographic axes. This builds a globular micelle-like particle which minimizes hydrophobic interactions with the solvent. The re®nement was conducted with non-crystallographic symmetry constraints up to a ®nal residual of R = 0.20 (R free = 0.26). The root-mean-square deviations from ideal geometry were 0.015 A Ê and 1.6 on bond distances and bond angles, respectively. Several sites for thiocyanate ions were analyzed.