Warm nights drive Coffea arabica ripening in Tanzania.

Studies have demonstrated that plant phenophases (e.g. budburst, flowering, ripening) are occurring increasingly earlier in the season across diverse ecologies globally. Despite much interest that climate change impacts have on coffee (Coffea arabica), relatively little is known about the driving factors determining its phenophases. Using high-resolution microclimatic data, this study provides initial insights on how climate change is impacting C. arabica phenophases in Tanzania. In particular, we use generalized additive models to show how warming nocturnal temperatures (Tnight), as opposed to day-time or maximum temperatures, have a superseding effect on the ripening of coffee and subsequent timing of harvest. A warm night index (WNI), generated from mean nocturnal temperature, permits accurate prediction of the start of the harvest season, which is superior to existing methods using growing degree days (GDD). The non-linear function indicates that a WNI of 15 °C is associated with the latest ripening coffee cherries (adjusted R2 = 0.95). As the WNI increases past the inflection point of ~ 16 °C, ripening occurs earlier and progresses more or less linearly at a rate of ~ 17 ± 1.95 days for every 1 °C increase in WNI. Using the WNI will thus not only allow farmers to more accurately predict their harvest start date, but also assist with identifying the most suitable adaptation strategies which may reduce harvest-related costs and buffer potential losses in quality and production.

Diagnosis of dental and mandibular asymmetries in children according to Levandoski Panoramic Analysis.

AIM: The purpose of this study was to evaluate the use of the Levandoski Panoramic Analysis in the diagnosis of dental and mandibular asymmetries and its contribution to clinical patient's evaluation and treatment planning. MATERIALS AND METHODS: Thirty-one randomly selected panoramic radiographs of children from 7 to 14 year old were analysed using 10 linear measurements. Right and left values were compared with Student's paired T tests. For each value, mean and standard deviation were computed separately for each side. RESULTS: Statistics. A dominance for the left side over the right side was observed. The data obtained were not statistically significant with the exception of maxillary length: the right side length of the maxilla was shorter (p<0.05) compared to the left side. CONCLUSIONS: Levandosky Panoramic Analisys represents a useful screening method in the diagnosis of dental and mandibular asymmetries.

Characterization of an interaction between insulin receptor substrate 1 and the insulin receptor by using the two-hybrid system.

Insulin receptor substrate 1 (IRS-1) is a major substrate of the insulin receptor and has been implicated in insulin signaling. Although IRS-1 is thought to interact with the insulin receptor, the nature of the interaction has not been defined. In this study, we used the two-hybrid assay of protein-protein interaction in the yeast Saccharomyces cerevisiae to study the interaction between human IRS-1 and the insulin receptor. We demonstrate that IRS-1 forms a specific complex with the cytoplasmic domain of the insulin receptor when both are expressed as hybrid proteins in yeast cells. We show that the interaction is strictly dependent upon receptor tyrosine kinase activity, since IRS-1 shows no interaction with a kinase-inactive receptor hybrid containing a mutated ATP-binding site. Furthermore, mutation of receptor tyrosine 960 to phenylalanine eliminates IRS-1 interaction in the two-hybrid assay. These data suggest that the interaction between IRS-1 and the receptor is direct and provide evidence that the juxtamembrane domain of the receptor is involved. Furthermore, we show that a 356-amino-acid region encompassed by amino acids 160 through 516 of IRS-1 is sufficient for interaction with the receptor in the two-hybrid assay. Lastly, in agreement with our findings for yeast cells, we show that the insulin receptor is unable to phosphorylate an IRS-1 protein containing a deletion of amino acids 45 to 516 when expressed in COS cells. The two-hybrid assay should provide a facile means by which to pursue a detailed understanding of this interaction.

Phosphotyrosine-dependent interaction of SHC and insulin receptor substrate 1 with the NPEY motif of the insulin receptor via a novel non-SH2 domain.

The SHC proteins have been implicated in insulin receptor (IR) signaling. In this study, we used the sensitive two-hybrid assay of protein-protein interaction to demonstrate that SHC interacts directly with the IR. The interaction is mediated by SHC amino acids 1 to 238 and is therefore independent of the Src homology 2 domain. The interaction is dependent upon IR autophosphorylation, since the interaction is eliminated by mutation of the IR ATP-binding site. In addition, mutational analysis of the Asn-Pro-Glu-Tyr (NPEY) motif within the juxtamembrane domain of the IR showed the importance of the Asn, Pro, and Tyr residues to both SHC and IR substrate 1 (IRS-1) binding. We conclude that SHC interacts directly with the IR and that phosphorylation of Tyr-960 within the IR juxtamembrane domain is necessary for efficient interaction. This interaction is highly reminiscent of that of IRS-1 with the IR, and we show that the SHC IR-binding domain can substitute for that of IRS-1 in yeast and COS cells. We identify a homologous region within the IR-binding domains of SHC and IRS-1, which we term the SAIN (SHC and IRS-1 NPXY-binding) domain, which may explain the basis of these interactions. The SAIN domain appears to represent a novel motif which is able to interact with autophosphorylated receptors such as the IR.

Application of thermography for monitoring stomatal conductance of Coffea arabica under different shading systems.

Stomatal regulation is a key process in the physiology of Coffea arabica (C. arabica). Intrinsically linked to photosynthesis and water relations, it provides insights into the plant's adaptive capacity, survival and growth. The ability to rapidly quantify this parameter for C. arabica under different agroecological systems would be an indispensable tool. Using a Flir E6 MIR Camera, an index that is equivalent to stomatal conductance (Ig) was compared with stomatal conductance measurements (gs) in a mature coffee plantation. In order to account for varying meteorological conditions between days, the methods were also compared under stable meteorological conditions in a laboratory and Ig was also converted to absolute stomatal conductance values (g1). In contrast to typical plant-thermography methods which measure indices once per day over an extended time period, we used high resolution hourly measurements over daily time series with 9 sun and 9 shade replicates. Eight daily time series showed a strong correlation between methods, while the remaining 10 were not significant. Including several other meteorological parameters in the calculation of g1 did not contribute to any stronger correlation between methods. Total pooled data (combined daily series) resulted in a correlation of ρ=0.66 (P≤2.2e-16), indicating that our approach is particularly useful for situations where absolute values of stomatal conductance are not required, such as for comparative purposes, screening or trend analysis. We use the findings to advance the protocol for a more accurate methodology which may assist in quantifying advantageous microenvironment designs for coffee, considering the current and future climates of coffee growing regions.

14-3-3 (epsilon) interacts with the insulin-like growth factor I receptor and insulin receptor substrate I in a phosphoserine-dependent manner.

The 14-3-3 proteins have been implicated as potential regulators of diverse signaling pathways. Here, using two-hybrid assays and in vitro assays of protein interaction, we show that the epsilon isoform of 14-3-3 interacts with the insulin-like growth factor I receptor (IGFIR) and with insulin receptor substrate I (IRS-1), but not with the insulin receptor (IR). Coprecipitation studies demonstrated an IGFI-dependent in vitro interaction between 14-3-3-glutathione S-transferase proteins and the IGFIR. In similar studies no interaction of 14-3-3 with the IR was observed. We present evidence to suggest that 14-3-3 interacts with phosphoserine residues within the COOH terminus of the IGFIR. Specifically, peptide competition studies combined with mutational analysis suggested that the 14-3-3 interaction was dependent upon phosphorylation of IGFIR serine residues 1272 and/or 1283, a region which has been implicated in IGFIR-dependent transformation. Phosphorylation of these serines appears to be dependent upon prior IGFIR activation since no interaction of 14-3-3 was observed with a kinase-inactive IGFIR in the two-hybrid assay nor was any in vitro interaction with unstimulated IGFIR derived from mammalian cells. We show that the interaction of 14-3-3 with IRS-1 also appears to be phosphoserine-dependent. Interestingly, 14-3-3 appears to interact with IRS-1 before and after hormonal stimulation. In summary, our data suggest that 14-3-3 interacts with phosphoserine residues within the COOH terminus of the IGFIR and within the central domain of IRS-1. The potential functional roles which 14-3-3 may play in IGFIR and IRS-1-mediated signaling remain to be elucidated.

Non-SH2 domains within insulin receptor substrate-1 and SHC mediate their phosphotyrosine-dependent interaction with the NPEY motif of the insulin-like growth factor I receptor.

Insulin receptor substrate-1 (IRS-1) and SHC become rapidly phosphorylated upon tyrosines after insulin-like growth factor I receptor (IGFIR) activation. In this study we demonstrate that IRS-1, SHC, and the p85 subunit of phosphatidylinositol 3-kinase interact directly and specifically with the IGFIR. The interaction of all three proteins is dependent upon IGFIR kinase activity and, furthermore, substitution of Tyr-950 with Phe within the NPEY motif of the IGFIR eliminated interaction with both SHC and IRS-1 but had no effect upon p85 interaction. We show that residues 160-516 of IRS-1 and 1-238 of SHC are sufficient and necessary for receptor interaction in the yeast two-hybrid assay. We also demonstrate a direct in vitro interaction between the IGFIR and a fusion protein containing SHC amino acids 1-238. No interaction was observed with a SHC protein containing only the SH2 domain. We conclude that SHC and IRS-1 interact with the tyrosine-phosphorylated NPEY motif of the IGFIR, and that both proteins interact via related motifs located in their amino termini. We conclude that the interactions of SHC and IRS-1 with the IGFIR are similar to those which we have previously defined with the insulin receptor.

Interaction of insulin receptor substrate-2 (IRS-2) with the insulin and insulin-like growth factor I receptors. Evidence for two distinct phosphotyrosine-dependent interaction domains within IRS-2.

Insulin receptor substrate 2 (IRS-2) has recently been shown to be a substrate of the insulin receptor (IR). In this study we utilize the yeast two-hybrid system and assays of in vitro interaction to demonstrate that IRS-2 interacts directly with the IR and the insulin-like growth factor I receptor. We show that, like IRS-1, the region of IRS-2 that contains the putative phosphotyrosine binding and SAIN elements (188-591) is sufficient for receptor interaction and that this interaction is dependent upon the NPX(p)Y (where (p)Y is phosphotyrosine) motifs within the juxtamembrane domains of the receptors. In addition to this amino-terminal NPX(p)Y-binding domain, an additional domain of strong interaction was identified in the central region of IRS-2 and was localized between amino acids 591 and 733. This interaction was found to be dependent upon receptor phosphorylation but was NPX(p)Y-independent. This region does not appear to have either an SH2 or a phosphotyrosine binding domain. Both of the interactions could also be demonstrated in vitro using IRS-2 glutathione S-transferase fusion proteins. We conclude that IRS-2, unlike IRS-1, can interact with tyrosine-phosphorylated receptors such as the IR and insulin-like growth factor I receptor via multiple independent binding motifs. Our findings suggest the existence of a previously unidentified phosphotyrosine-dependent binding domain within the central region of IRS-2.