Postveraison Deficit Irrigation Effects on Fruit Quality and Yield of "Flame Seedless" Table Grape Cultivated under Greenhouse and Net.

Lack of color in the skin of red table grape varieties is a serious problem in areas of warm climate. This problem is often addressed by the application of ethylene release products such as ethephon. Strict regulation in the use of this product in EU forces European grape producers to look for suitable alternatives. With the aim to increase red skin color, we applied regulated deficit irrigation (RDI) strategies from veraison until harvest on "Flame Seedless" table grape vines cultivated under nets and under a plastic greenhouse in South East Spain, and compared yield and fruit quality with vines fully irrigated under the same net and plastic greenhouses. Our results show a modest improvement in the percentage of commercial clusters with better skin color, probably because the short duration of the deficit irrigation period only caused a slight decrease in soil water content and a mild water stress in RDI vines. Larger differences were observed under the more limiting conditions of the plastic greenhouse for light environment, especially when berry skin color was measured by CIRG (color index of red grape). More noticeable effect of RDI was noted on fruit earliness. Water savings were also remarkable. Negative effects of RDI on berry size or total soluble solid content were not perceived. Our results suggest that RDI is a suitable strategy to save irrigation water without substantial negative effects on yield and berry size. However, the effects on skin color were insufficient in the trial conditions.

Expression, purification and crystallization of human CD5 domain III, a nano-scale crystallization example.

The human lymphocyte receptor CD5, a key regulator of immune responses, is involved in the modulation of antigen specific receptor-mediated T cell activation and differentiation signals. CD5 is a membrane glycoprotein which belongs to the group B scavenger receptor cysteine-rich (SRCR) superfamily for which no structural information is available. The most conserved membrane-proximal SRCR domain of CD5 (domain III) has been expressed in HEK-EBNA-293 cells. Although the yield of the purified protein was at the level of micrograms, well diffracting crystals have been obtained. The crystals belong to a tetragonal space group P4(1)22 or P4(3)22. They contain two molecules per asymmetric unit and diffracted to 2.5A resolution using synchrotron radiation. The strategy shown here to produce, isolate and crystallize CD5 domain III can be used for other mammalian proteins difficult to produce for structural or other biophysical studies.

Molecular and structural basis of polo-like kinase 1 substrate recognition: Implications in centrosomal localization.

Polo-like kinase (Plk1) is crucial for cell cycle progression through mitosis. Here we present the molecular and structural mechanisms that regulate the substrate recognition of Plk1 and influence its centrosomal localization and activity. Our work shows that Plk1 localization is controlled not only by the polo box domain (PBD); remarkably, the kinase domain is also involved in Plk1 targeting mechanism to the centrosome. The crystal structures of the PBD in complex with Cdc25C and Cdc25C-P target peptides reveal that Trp-414 is fundamental in their recognition regardless of its phosphorylation status. Binding measurements demonstrate that W414F mutation abolishes molecular recognition and diminishes centrosomal localization. Therefore, Plk1 centrosomal localization is not controlled by His-538 and Lys-540, the residues involved in phosphorylated target binding. The different conformations of the loop, which connects the polo boxes in the apo and the PBD-Cdc25C and PBD-Cdc25C-P complex structures, together with changes in the proline adjacent to the phosphothreonine in the target peptide, suggest a regulatory mechanism to detect binding of unphosphorylated or phosphorylated target substrates. Altogether, these data propose a model for the interaction between Plk1 and Cdc25C.

Crystallization and preliminary X-ray diffraction studies on the human Plk1 Polo-box domain in complex with an unphosphorylated and a phosphorylated target peptide from Cdc25C.

Polo-like kinase (Plk1) is crucial for cell-cycle progression via mitosis. Members of the Polo-like kinase family are characterized by the presence of a C-terminal domain termed the Polo-box domain (PBD) in addition to the N-terminal kinase domain. The PBD of Plk1 was cloned and overexpressed in Escherichia coli. Crystallization experiments of the protein in complex with an unphosphorylated and a phosphorylated target peptide from Cdc25C yield crystals suitable for X-ray diffraction analysis. Crystals of the PBD in complex with the phosphorylated peptide belong to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 38.23, b = 67.35, c = 88.25 angstroms, alpha = gamma = beta = 90 degrees, and contain one molecule per asymmetric unit. Crystals of the PBD in complex with the unphosphorylated peptide belong to the monoclinic space group P2(1), with unit-cell parameters a = 40.18, b = 49.17, c = 56.23 angstroms, alpha = gamma = 90, beta = 109.48 degrees, and contain one molecule per asymmetric unit. The crystals diffracted to resolution limits of 2.1 and 2.85 angstroms using synchrotron radiation at the European Synchrotron Radiation Facility (ESRF) and the Swiss Light Source (SLS), respectively.