Role of extracellular charged amino acids in the yeast alpha-factor receptor

The yeast pheromone receptor, Ste2p, is a G protein coupled receptor that initiates cellular responses to alpha-mating pheromone, a 13 residue peptide that carries a net positive charge at physiological pH. We have examined the role of extracellular charged groups on the receptor in response to the pheromone. Substitutions of Asn or Ala for one extracellular residue, Asp275, affected both pheromone binding and signaling, suggesting that this position interacts directly with ligand. The other seven extracellular acidic residues could be individually replaced by polar residues with no detectable effects on receptor function. However, substitution of Ala for each of these seven residues resulted in impairment of signaling without affecting pheromone binding, implying that the polar nature of these residues promotes receptor activation. In contrast, substitution of Ala for each of the six positively charged residues at the extracellular surface of Ste2p did not affect signaling.     

Arabidopsis At5g39790 encodes a chloroplast-localized,carbohydrate-binding,coiled-coil domain-containing putative scaffold protein

Background  

Starch accumulation and degradation in chloroplasts is accomplished by a suite of over 30 enzymes. Recent work has emphasized the importance of multi-protein complexes amongst the metabolic enzymes, and the action of associated non-enzymatic regulatory proteins. Arabidopsis At5g39790 encodes a protein of unknown function whose sequence was previously demonstrated to contain a putative carbohydrate-binding domain.     

Clinical Utility of Waist Circumference in Predicting All‐cause Mortality in a Preventive Cardiology Clinic Population: A PreCIS Database Study

Although obesity is a risk factor for mortality, it is unclear whether waist circumference (WC) is a better predictor of mortality than BMI in a clinical setting of patients at high risk for coronary artery disease (CAD). Thus, we compared the association between WC and BMI with all‐cause mortality in relation to traditional CAD risk factors in a high‐risk cohort. Study population included 5,453 consecutive new patients seen between 1996 and 2005 for management of CAD risk factors in a preventive cardiology clinic. Mortality was determined from the Social Security Death Index. There were 359 deaths over a median follow‐up of 5.2 years. Mortality was greater in high (>102 cm in men and >88 cm in women) vs. normal WC in both genders (P < 0.01). The unadjusted Cox proportional hazard ratio (HR) for continuous WC (per cm) was 1.02 (P < 0.001) in both genders and remained significant after adjustment for CAD risk factors (HR = 1.01 in men, HR = 1.03 in women, both P < 0.05). BMI did not associate statistically with mortality. WC associated with diabetes mellitus (DM) and CAD prevalence (P < 0.001). BMI associated only with DM (P < 0.001) and this association disappeared when WC was added to the model. We conclude that WC is an independent predictor of all‐cause mortality in a preventive cardiology population. These data affirm the clinical importance of WC measurements for mortality, DM, and CAD risk prediction and suggest that obesity‐specific interventions targeting WC in addition to traditional risk factor management may favorably impact these outcomes.     

Differential interactions of fluorescent agonists and antagonists with the yeast G protein coupled receptor Ste2p

We describe a rapid method to probe for mutations in cell surface ligand-binding proteins that affect the environment of bound ligand. The method uses fluorescence-activated cell sorting to screen randomly mutated receptors for substitutions that alter the fluorescence emission spectrum of environmentally sensitive fluorescent ligands. When applied to the yeast α-factor receptor Ste2p, a G protein-coupled receptor, the procedure identified 22 substitutions that red shift the emission of a fluorescent agonist, including substitutions at residues previously implicated in ligand binding and at additional sites. A separate set of substitutions, identified in a screen for mutations that alter the emission of a fluorescent α-factor antagonist, occurs at sites that are unlikely to contact the ligand directly. Instead, these mutations alter receptor conformation to increase ligand-binding affinity and provide signaling in response to antagonists of normal receptors. These results suggest that receptor-agonist interactions involve at least two sites, of which only one is specific for the activated conformation of the receptor.     

A mathematical model for insulin kinetics and its application to protein-deficient (malnutrition-related) diabetes mellitus (PDDM)

A nonlinear mathematical model which incorporates both beta-cell kinetics and a glucose-insulin feedback system is proposed for describing the time variations of plasma glucose and insulin levels. Numerical simulations show that this model is consistent with experimental observations on normal groups. An analysis of the changes in the solutions with variations in the parameters showed that a decrease in a single parameter gave results consistent with experimental findings in protein-deficient (malnutrition-related) diabetes mellitus (PDDM). The model predicts that it is the function and not the number of beta cells which is reduced in PDDM.     

Lysine acetylome profiling uncovers novel histone deacetylase substrate proteins in Arabidopsis

Histone deacetylases have central functions in regulating stress defenses and development in plants. However, the knowledge about the deacetylase functions is largely limited to histones, although these enzymes were found in diverse subcellular compartments. In this study, we determined the proteome‐wide signatures of the RPD3/HDA1 class of histone deacetylases in Arabidopsis. Relative quantification of the changes in the lysine acetylation levels was determined on a proteome‐wide scale after treatment of Arabidopsis leaves with deacetylase inhibitors apicidin and trichostatin A. We identified 91 new acetylated candidate proteins other than histones, which are potential substrates of the RPD3/HDA1‐like histone deacetylases in Arabidopsis, of which at least 30 of these proteins function in nucleic acid binding. Furthermore, our analysis revealed that histone deacetylase 14 (HDA14) is the first organellar‐localized RPD3/HDA1 class protein found to reside in the chloroplasts and that the majority of its protein targets have functions in photosynthesis. Finally, the analysis of HDA14 loss‐of‐function mutants revealed that the activation state of RuBisCO is controlled by lysine acetylation of RuBisCO activase under low‐light conditions.