The millimeter-wave spectrum of the SiP radical (X2Πi): Rotational perturbations and hyperfine structure.

The millimeter/submillimeter-wave spectrum of the SiP radical (X2Πi) has been recorded using direct absorption spectroscopy in the frequency range of 151-532 GHz. SiP was synthesized in an AC discharge from the reaction of SiH4 and gas-phase phosphorus, in argon carrier gas. Both spin-orbit ladders were observed. Fifteen rotational transitions were measured originating in the Ω = 3/2 ladder, and twelve in the Ω = 1/2 substate, each exhibiting lambda doubling and, at lower frequencies, hyperfine interactions from the phosphorus nuclear spin of I = 1/2. The lambda-doublets in the Ω = 1/2 levels appeared to be perturbed at higher J, with the f component deviating from the predicted pattern, likely due to interactions with the nearby excited A2Σ+ electronic state, where ΔEΠ-Σ âˆ¼ 430 cm-1. The data were analyzed using a Hund's case aß Hamiltonian and rotational, spin-orbit, lambda-doubling, and hyperfine parameters were determined. A 2Π/2Σ deperturbation analysis was also performed, considering spin-orbit, spin-electronic, and L-uncoupling interactions. Although SiP is clearly not a hydride, the deperturbed parameters derived suggest that the pure precession hypothesis may be useful in assessing the 2Π/2Σ interaction. Interpretation of the Fermi contact term, bF, the spin-dipolar constant, c, and the nuclear spin-orbital parameter, a, indicates that the orbital of the unpaired electron is chiefly pπ in character. The bond length in the v = 0 level was found to be r0 = 2.076 Å, suggestive of a double bond between the silicon and phosphorus atoms.

Outbreak of SARS-CoV-2 in a teenage discotheque in Northern Ireland-November 2021.

OBJECTIVES: To report key findings associated with an outbreak of SARS-CoV-2 following a teenage disco in Northern Ireland. STUDY DESIGN: Observational case series. METHODS: A case was defined as an individual who attended the event with a positive SARS-CoV-2 result between 6th and 20th November 2021. Demographic and clinical information, including symptom status, date of onset and school attended, were recorded during contact tracing. Vaccination status was derived from the COVID-19 Vaccine Management System. Forty-five samples associated with the outbreak were sequenced as part of the NI Whole Genome Sequencing (WGS) programme. RESULTS: Only 2.4% (5/205) of cases received a COVID-19 vaccine more than 14 days before the event. 84.9% (174/205) had received no vaccine at the time of the event and 12.7% (26/205) had been vaccinated within 14 days, offering only limited disease protection. The AY4.2.2 lineage of two cases who attended the event after symptom onset was found in 69% of sequenced outbreak cases. CONCLUSIONS: This study demonstrates extensive COVID-19 transmission in largely unvaccinated teenagers in an indoor venue with limited social distancing, close social contact and mixing, limited ventilation and singing and shouting. Public Health authorities developing COVID-19 entertainment regulations should consider congregations of teenagers in these settings, especially if vaccination rates are low in this group or they are not eligible for vaccination at that time. Public communications should be developed to ensure young people with COVID-19 symptoms follow public guidance regarding self-isolation and in particular avoid indoor events with larger numbers.

Salbutamol-induced electrophysiological changes show no correlation with electrophysiological changes during hyperinsulinaemic-hypoglycaemic clamp in young people with Type 1 diabetes.

AIMS: Hypoglycaemia causes QT-interval prolongation and appears pro-arrhythmogenic. Salbutamol, a ß2 -adrenoreceptor agonist also causes QT-interval prolongation. We hypothesized that the magnitude of electrophysiological changes induced by salbutamol and hypoglycaemia might relate to each other and that salbutamol could be used as a non-invasive screening tool for predicting an individual's electrophysiological response to hypoglycaemia. METHODS: Eighteen individuals with Type 1 diabetes were administered 2.5 mg of nebulized salbutamol. Participants then underwent a hyperinsulinaemic-hypoglycaemic clamp (2.5 mmol/l for 1 h). During both experiments, heart rate and serum potassium (and catecholamines during the clamp) were measured and a high-resolution electrocardiogram (ECG) was recorded at pre-set time points. Cardiac repolarization was measured by QT-interval duration adjusted for heart rate (QTc ), T-wave amplitude (Tamp ), T-peak to T-end interval duration (Tp Tend ) and T-wave area symmetry (Tsym ). The maximum changes vs. baseline in both experiments were assessed for their linear dependence. RESULTS: Salbutamol administration caused QTc and Tp Tend prolongation and a decrease in Tamp and Tsym . Hypoglycaemia caused increased plasma catecholamines, hypokalaemia, QTc and Tp Tend prolongation, and a decrease in Tamp and Tsym . No significant correlations were found between maximum changes in QTc [r = 0.15, 95% confidence interval (95% CI) -0.341 to 0.576; P = 0.553), Tp Tend (r = 0.075, 95% CI -0.406 to 0.524; P = 0.767), Tsym (r = 0.355, 95% CI -0.132 to 0.706; P = 0.149) or Tamp (r = 0.148, 95% CI -0.347 to 0.572; P = 0.558) in either experiment. CONCLUSIONS: Both hypoglycaemia and salbutamol caused pro-arrhythmogenic electrophysiological changes in people with Type 1 diabetes but were not related in any given individual. Salbutamol does not appear useful in assessing an individual's electrophysiological response to hypoglycaemia.

The AURKA/TPX2 axis drives colon tumorigenesis cooperatively with MYC.

BACKGROUND: The MYC oncogene has long been established as a central driver in many types of human cancers including colorectal cancer. However, the realization of MYC-targeting therapies remains elusive; as a result, synthetic lethal therapeutic approaches are alternatively being explored. A synthetic lethal therapeutic approach aims to kill MYC-driven tumors by targeting a certain co-regulator on the MYC pathway. PATIENTS AND METHODS: We analyzed copy number and expression profiles from 130 colorectal cancer tumors together with publicly available datasets to identify co-regulators on the MYC pathway. Candidates were functionally tested by in vitro assays using colorectal cancer and normal fibroblast cell lines. Additionally, survival analyses were carried out on another 159 colorectal cancer patients and public datasets. RESULTS: Our in silico screening identified two MYC co-regulator candidates, AURKA and TPX2, which are interacting mitotic regulators located on chromosome 20q. We found the two candidates showed frequent co-amplification with the MYC locus while expression levels of MYC and the two genes were positively correlated with those of MYC downstream target genes across multiple cancer types. In vitro, the aberrant expression of MYC, AURKA and TPX2 resulted in more aggressive anchorage-independent growth in normal fibroblast cells. Furthermore, knockdown of AURKA or TPX2, or treatment with an AURKA-specific inhibitor effectively suppressed the proliferation of MYC-expressing colorectal cancer cells. Additionally, combined high expression of MYC, AURKA and TPX2 proved to be a poor prognostic indicator of colorectal cancer patient survival. CONCLUSIONS: Through bioinformatic analyses and experiments, we proposed TPX2 and AURKA as novel co-regulators on the MYC pathway. Inhibiting the AURKA/TPX2 axis would be a novel synthetic lethal therapeutic approach for MYC-driven cancers.

Campylobacteriosis outbreak associated with consumption of undercooked chicken liver pâté in the East of England, September 2011: identification of a dose-response risk.

A foodborne outbreak with 49 cases (22 culture positive for Campylobacter sp.) following a wedding party in the East of England was investigated. A retrospective cohort study identified an association between consumption of chicken liver pâté and infection with Campylobacter jejuni/coli. There was a statistically significant association between dose (amount of chicken liver pâté eaten) and the risk of disease ['tasted': odds ratio (OR) 1·5, 95% confidence interval (CI) 0·04-∞; 'partly eaten': OR 8·4, 95% CI 1·4-87·5; 'most or all eaten': OR 36·1, 95% CI 3·3-2119). The local authority found evidence that the preparation of chicken livers breached Food Standards Agency's guidelines. This epidemiological investigation established a clear dose-response relationship between consumption of chicken liver pâté and the risk of infection with Campylobacter. The continuing need to raise public awareness of the risk to human health posed by undercooked chicken liver is evident.

Trends in alkali metal hydrosulfides: a combined Fourier transform microwave/millimeter-wave spectroscopic study of KSH (X1A').

The pure rotational spectrum of KSH (X(1)A') has been measured using millimeter-wave direct absorption and Fourier transform microwave (FTMW) techniques. This work is the first gas-phase experimental study of this molecule and includes spectroscopy of KSD as well. In the millimeter-wave system, KSH was synthesized in a DC discharge from a mixture of potassium vapor, H2S, and argon; a discharge-assisted laser ablation source, coupled with a supersonic jet expansion, was used to create the species in the FTMW instrument. Five and three rotational transitions in the range 3-57 GHz were recorded with the FTMW experiment for KSH and KSD, respectively, in the K(a) = 0 component; in these data, potassium quadrupole hyperfine structure was observed. Five to six transitions with K(a) = 0-5 were measured in the mm-wave region (260-300 GHz) for the two species. The presence of multiple asymmetry components in the mm-wave spectra indicates that KSH has a bent geometry, in analogy to other alkali hydrosulfides. The data were analyzed with an S-reduced asymmetric top Hamiltonian, and rotational, centrifugal distortion, and potassium electric quadrupole coupling constants were determined for both isotopolgues. The r0 geometry for KSH was calculated to be r(S-H) = 1.357(1) Å, r(K-S) = 2.806(1) Å, and θ(M-S-H) (°) = 95.0 (1). FTMW measurements were also carried out on LiSH and NaSH; metal electric quadrupole coupling constants were determined for comparison with KSH. In addition, ab initio computations of the structures and vibrational frequencies at the CCSD(T)/6-311++G(3df,2pd) and CCSD(T)/aug-cc-pVTZ levels of theory were performed for LiSH, NaSH, and KSH. Overall, experimental and computational data suggest that the metal-ligand bonding in KSH is a combination of electrostatic and covalent forces.

Obesity is associated with impaired immune response to influenza vaccination in humans.

BACKGROUND: Obesity is an independent risk factor for morbidity and mortality from pandemic influenza H1N1. Influenza is a significant public health threat, killing an estimated 250,000-500,000 people worldwide each year. More than one in ten of the world's adult population is obese and more than two-thirds of the US adult population is overweight or obese. No studies have compared humoral or cellular immune responses to influenza vaccination in healthy weight, overweight and obese populations despite clear public health importance. OBJECTIVE: The study employed a convenience sample to determine the antibody response to the 2009-2010 inactivated trivalent influenza vaccine (TIV) in healthy weight, overweight and obese participants at 1 and 12 months post vaccination. In addition, activation of CD8⁺ T cells and expression of interferon-γ and granzyme B were measured in influenza-stimulated peripheral blood mononuclear cell (PBMC) cultures. RESULTS: Body mass index (BMI) correlated positively with higher initial fold increase in IgG antibodies detected by enzyme-linked immunosorbent assay to TIV, confirmed by HAI antibody in a subset study. However, 12 months post vaccination, higher BMI was associated with a greater decline in influenza antibody titers. PBMCs challenged ex vivo with vaccine strain virus, demonstrated that obese individuals had decreased CD8⁺ T-cell activation and decreased expression of functional proteins compared with healthy weight individuals. CONCLUSION: These results suggest obesity may impair the ability to mount a protective immune response to influenza virus.

Oral DNA vaccination of rainbow trout, Oncorhynchus mykiss (Walbaum), against infectious haematopoietic necrosis virus using PLGA [Poly(D,L-Lactic-Co-Glycolic Acid)] nanoparticles.

A DNA vaccine against infectious haematopoietic necrosis virus (IHNV) is effective at protecting rainbow trout, Oncorhynchus mykiss, against disease, but intramuscular injection is required and makes the vaccine impractical for use in the freshwater rainbow trout farming industry. Poly (D,L-lactic-co-glycolic acid) (PLGA) is a U.S. Food and Drug Administration (FDA) approved polymer that can be used to deliver DNA vaccines. We evaluated the in vivo absorption of PLGA nanoparticles containing coumarin-6 when added to a fish food pellet. We demonstrated that rainbow trout will eat PLGA nanoparticle coated feed and that these nanoparticles can be detected in the epithelial cells of the lower intestine within 96 h after feeding. We also detected low levels of gene expression and anti-IHNV neutralizing antibodies when fish were fed or intubated with PLGA nanoparticles containing IHNV G gene plasmid. A virus challenge evaluation suggested a slight increase in survival at 6 weeks post-vaccination in fish that received a high dose of the oral vaccine, but there was no difference when additional fish were challenged at 10 weeks post-vaccination. The results of this study suggest that it is possible to induce an immune response using an orally delivered DNA vaccine, but the current system needs improvement.

Cardiac arrhythmias and electrophysiologic responses during spontaneous hyperglycaemia in adults with type 1 diabetes mellitus.

AIM: We examined the effect of spontaneous hyperglycaemia in adults with type 1 diabetes mellitus (T1DM) and without history of cardiovascular disease on heart rate variability (HRV), cardiac repolarisation and incidence of cardiac arrhythmias. METHODS: Thirty-seven individuals with T1DM (age 17-50 years, 19 males, mean duration of diabetes 19.3 SD(9.6) years) underwent 96 h of simultaneous ambulatory 12-lead Holter ECG and blinded continuous interstitial glucose (IG) monitoring (CGM). HRV, QT interval and cardiac repolarisation were assessed during hyperglycaemia (IG ≥ 15 mmol/l) and compared with matched euglycaemia (IG 5-10 mmol/l) on a different day, separately during the day and night. Rates of arrhythmias were assessed by calculating incidence rate differences. RESULTS: Simultaneous ECG and CGM data were recorded for 2395 hours. During daytime hyperglycaemia vs euglycaemia the mean QTc interval duration was 404 SD(21)ms vs 407 SD(20)ms, P = 0.263. T-peak to T-end interval duration corrected for heart rate (TpTendc) shortened: 74.8 SD(16.1)ms vs 79.0 SD(14.8)ms, P = 0.033 and T-wave symmetry increased: 1.62 SD(0.33) vs 1.50 SD(0.39), P = 0.02. During night-time hyperglycaemia vs euglycaemia, the mean QTc interval duration was 401 SD(26)ms vs 404 SD(27)ms, P = 0.13 and TpTend shortened: 62.4 SD(12.0)ms vs 67.1 SD(11.8)ms, P = 0.003. The number of cardiac arrhythmias was low and confined to bradycardia and isolated ectopic beats. A considerable inter-subject and diurnal variability was observed. CONCLUSIONS: Hyperglycaemia in individuals with T1DM without known cardiovascular disease was not associated with clinically important cardiac arrhythmias.

Effects of ethnicity on diabetes incidence and prevention: results of the Diabetes REduction Assessment with ramipril and rosiglitazone Medication (DREAM) trial.

AIMS: Risk of Type 2 diabetes varies by ethnicity, but whether ethnicity remains important among those who have impaired glucose tolerance or impaired fasting glucose is uncertain. Whether the effect of thiazolidinedione treatment on diabetes prevention in persons with non-diabetic dysglycaemia varies by ethnicity is also not known. We addressed these questions using data collected in the DREAM trial. METHODS: A 2-by-2 factorial double-blind randomized controlled trial to compare the effects of rosiglitazone and ramipril on the primary outcome of diabetes or death in persons meeting criteria for impaired glucose tolerance or impaired fasting glucose. The effect of these interventions by ethnicity was estimated using Cox regression analysis. RESULTS: Of 5269 adults, 2365 were randomly assigned to rosiglitzone and 2634 to placebo. South Asians showed a higher hazard for the primary outcome compared with Europeans (hazard ratio, 95% confidence interval 2.21, 1.41-3.47) adjusted for age, gender, BMI, waist-hip ratio and geographic region. A lesser increase in risk was seen in Black people (1.37, 1.04-1.81). A significant reduction in risk of the primary outcome with rosiglitazone treatment assignment was seen in all ethnic groups, but the treatment effect significantly differed by ethnicity (P=0.0242), with South Asians experiencing a smaller, and Latinos a larger preventive effect. CONCLUSIONS: Ethnicity is an important risk factor for Type 2 diabetes in dysglycaemic persons. All ethnic groups experienced a large significant reduction in diabetes risk because of rosiglitazone. The magnitude of this reduction differed by ethnicity. Given the post hoc nature of this analysis, further confirmation of these findings is needed.

The heart is a target organ for androgen.

Autoradiographic and biochemical analyses of the hearts of female rhesus monkeys and baboons indicate that atrial and ventricular myocardial cells contain androgen receptors. Although the specific effects of nuclear uptake and retention of androgen on the function of heart muscle cells are not known, the presence of this receptor suggests that sex steroid hormones may affect myocardial function directly and may explain some of the peculiar differences in heart disease between men and women.

Reduced leucine-enkephalin--like immunoreactive substance in hamster basal ganglia after long-term ethanol exposure.

Golden Syrian hamsters were placed individually in cages with three drinking bottles--one empty, one containing water, and the third containing water and ethanol. Control hamsters received water only. After 1 year the experimental hamsters showed a significantly lower concentration of leucine-enkephalin-like immunoreactive substance in the basal ganglia than the control hamsters. This finding indicates that the action of ethanol involves endogenous peptidyl opiates.

Androgen receptors in the brain: what are we measuring?

Androgen has long been known to act on the brain to modify behavior and other brain functions. In the past, two methods have been used to characterize the putative receptors which mediate these actions. Autoradiography has been used to map and identify androgen binding neurons. Binding studies have been conducted to quantify and characterize the system(s). The resultant data are discordant and a new model is proposed to resolve the apparent differences. It is proposed that there are three categories of receptors for androgen in the brain. One receptor preferentially binds testosterone and a second one preferentially binds DHT. Both of these receptors are in equilibrium between nucleus and cytoplasm according to the free water content of the compartments. Both of these receptors can be activated and transformed by steroid and thus concentrate in the nucleus. It is proposed that a third receptor binds both steroids with the same relative affinity. However, this third receptor can only be activated but not transformed (i.e. it does not concentrate in the nucleus). The proposed system implies that testosterone acts on a few discrete populations of neurons in the brain while DHT has a very diffuse action on the central nervous system.

Effect of rosiglitazone on the frequency of diabetes in patients with impaired glucose tolerance or impaired fasting glucose: a randomised controlled trial.

BACKGROUND: Rosiglitazone is a thiazolidinedione that reduces insulin resistance and might preserve insulin secretion. The aim of this study was to assess prospectively the drug's ability to prevent type 2 diabetes in individuals at high risk of developing the condition. METHODS: 5269 adults aged 30 years or more with impaired fasting glucose or impaired glucose tolerance, or both, and no previous cardiovascular disease were recruited from 191 sites in 21 countries and randomly assigned to receive rosiglitazone (8 mg daily; n=2365) or placebo (2634) and followed for a median of 3 years. The primary outcome was a composite of incident diabetes or death. Analyses were done by intention to treat. This trial is registered at ClinicalTrials.gov, number NCT00095654. FINDINGS: At the end of study, 59 individuals had dropped out from the rosiglitazone group and 46 from the placebo group. 306 (11.6%) individuals given rosiglitazone and 686 (26.0%) given placebo developed the composite primary outcome (hazard ratio 0.40, 95% CI 0.35-0.46; p<0.0001); 1330 (50.5%) individuals in the rosiglitazone group and 798 (30.3%) in the placebo group became normoglycaemic (1.71, 1.57-1.87; p<0.0001). Cardiovascular event rates were much the same in both groups, although 14 (0.5%) participants in the rosiglitazone group and two (0.1%) in the placebo group developed heart failure (p=0.01). INTERPRETATION: Rosiglitazone at 8 mg daily for 3 years substantially reduces incident type 2 diabetes and increases the likelihood of regression to normoglycaemia in adults with impaired fasting glucose or impaired glucose tolerance, or both.

Venous thromboembolism in association with features of the metabolic syndrome.

BACKGROUND: Central obesity, diabetes mellitus, dyslipidaemia and chronic hypertension--features of the metabolic syndrome--have been individually associated with venous thromboembolism (VTE). However, whether each of these factors additively increases the risk of VTE is uncertain. AIM: To determine whether features of the metabolic syndrome independently increase the risk of VTE. DESIGN: Prospective cohort study derived from the Heart Outcomes Prevention Evaluation 2 (HOPE-2) randomized clinical trial. SETTING: One hundred and forty-five clinical centres in 13 countries. METHODS: We studied 5522 adults aged > or =55 years with cardiovascular disease or diabetes mellitus. At enrollment, 35% had 0-1 features of the metabolic syndrome, 30% had two, 24% had three and 11% had four. We defined symptomatic VTE as an objectively confirmed new episode of deep-vein thrombosis or pulmonary embolism. RESULTS: VTE occurred in 88 individuals during a median 5.0 years of follow-up. The incidence rate of VTE (per 100 person-years) was 0.30 with 0-1 features, 0.36 with two features, 0.38 with three features and 0.40 with four features of the metabolic syndrome (trend p = 0.43). Relative to the presence of 0-1 features of the metabolic syndrome, the adjusted hazard ratio (95%CI) for VTE was 1.22 (0.71-2.08) with two features, 1.25 (0.70-2.24) with three features, and 1.26 (0.59-2.69) with four features. DISCUSSION: The number of features of the metabolic syndrome present was not a clinically important risk factor for VTE in older adults with vascular arterial disease.

Rationale, design and baseline characteristics of a large, simple, randomized trial of combined folic acid and vitamins B6 and B12 in high-risk patients: the Heart Outcomes Prevention Evaluation (HOPE)-2 trial.

BACKGROUND: Epidemiological studies suggest that mild to moderate elevation in plasma homocysteine concentration is associated with increased risk of atherothrombotic cardiovascular (CV) disease. Simple, inexpensive and nontoxic therapy with folic acid and vitamins B6 and B12 reduces plasma homocysteine levels by approximately 25% to 30% and may reduce CV events. Therefore, a large, randomized clinical trial--the Heart Outcomes Prevention Evaluation (HOPE)-2 study--is being conducted to evaluate this therapy in patients at high risk for CV events. OBJECTIVES: To evaluate whether long-term therapy with folic acid and vitamins B6 and B12 reduces the risk of major CV events in a high-risk population. The primary study outcome is the composite of death from CV causes, myocardial infarction and stroke. METHODS: A total of 5522 patients aged 55 years or older with pre-existing CV disease or with diabetes and additional risk factor(s) at 145 centres in 13 countries were randomly assigned to daily therapy with combined folic acid 2.5 mg, vitamin B6 50 mg and vitamin B12 1 mg, or to placebo. Follow-up will average five years, to be completed by the end of 2005. RESULTS: The patients' baseline characteristics confirmed their high-risk status. Baseline homocysteine levels varied between countries and regions. HOPE-2 is one of the largest trials of folate and vitamins B6 and B12 and is expected to significantly contribute to the evaluation of the role of homocysteine lowering in CV prevention.

Effect of zinc administration on the growth of L1210 and BW5147 tumors in mice.

Aqueous zinc acetate injected ip prevented tumor growth in 50-70% of BDF male mice previously inoculated ip with L1210 leukemia cells. However, aqueous zinc acetate injected sc did not prevent tumor growth in AKR/J mice inoculated im with BW5147 lymphatic leukemia cells. In the latter mice, only a small but statistically significant increase in mean survival was noted.

Approaches for deciphering the structural basis of low temperature enzyme activity.

An increasing number of enzymes active at low temperature are being studied to help determine the structural features important for cold-activity. This review examines the diversity of prokaryotic cold-active enzymes and the features proposed to account for low temperature activity. We then consider the difficulty of identifying the key structural features needed for cold-activity and the need to compare enzymes having different temperature optima from phylogenetically related organisms to determine features responsible for low temperature activity. In addition to studying naturally occurring enzymes, directed evolution experiments are discussed as methods for examining the proposed mechanisms influencing the thermal dependence of activity.

Allelic association of the D2 dopamine receptor gene with receptor-binding characteristics in alcoholism.

The allelic association of the human D2 dopamine receptor gene with the binding characteristics of the D2 dopamine receptor was determined in 66 brains of alcoholic and non-alcoholic subjects. In a blinded experiment, DNA from the cerebral cortex was treated with the restriction endonuclease Taql and probed with a 1.5-kilobase (kb) digest of a clone (lambda hD2G1) of the human D2 dopamine receptor gene. The binding characteristics (Kd [binding affinity] and Bmax [number of binding sites]) of the D2 dopamine receptor were determined in the caudate nuclei of these brains using tritiated spiperone as the ligand. The adjusted Kd was significantly lower in alcoholic than in nonalcoholic subjects. In subjects with the A1 allele, in whom a high association with alcoholism was found, the Bmax was significantly reduced compared with the Bmax of subjects with the A2 allele. Moreover, a progressively reduced Bmax was found in subjects with A2/A2, A1/A2, and A1/A1 alleles, with subjects with A2/A2 having the highest mean values, and subjects with A1/A1, the lowest. The polymorphic pattern of the D2 dopamine receptor gene and its differential expression of receptors suggests the involvement of the dopaminergic system in conferring susceptibility to at least one subtype of severe alcoholism.

Human progesterone receptor transformation and nuclear down-regulation are independent of phosphorylation.

We have studied the phosphorylation of progesterone receptors (PR) in T47Dco human breast cancer cells using a monoclonal antibody directed against human PR called AB-52. This antibody recognizes both the A- (Mr approximately 94,000) and B- (Mr approximately 120,000) hormone binding proteins of PR, and was used to immunoprecipitate phosphorylated receptors isolated from cells incubated in vivo with [32P]orthophosphate. The specific activity, or phosphorylation levels, relative to protein levels was quantified by combined immunoblotting and autoradiography followed by densitometry. We find that immunopurified untransformed hormone-free receptors, which have a characteristic triplet B, singlet A structure, are phosphoproteins with similar levels of phosphate incorporation in all protein bands. If PR are first transformed to the nuclear binding form by treatment of cells with progesterone, and then labeled with [32P]orthophosphate, the receptor proteins are additionally phosphorylated. These chromatin-bound hormone occupied receptors incorporate five to 10 times more labeled phosphate per total receptor protein than do PR from untreated cells during the same [32P]incubation time. The second round of phosphorylation may also account for mobility shifts of transformed A- and B-receptors observed in sodium dodecyl sulfate-polyacrylamide gels. Both untransformed and transformed species of A- and B-receptors are phosphorylated only on serine residues, and neither the extent of phosphorylation, nor the phosphoamino acids, are affected by treatment of the cells with epidermal growth factor or insulin. We previously reported that after hormone binding and transformation of receptors to the tight chromatin binding state, PR undergo processing, or nuclear down-regulation. AB-52 was used to compare PR protein and phosphorylation levels when cells were treated for 0.5-48 h with progesterone or the synthetic progestin R5020. Both agonists lead to hyperphosphorylation of nuclear PR before phosphorylation levels decrease, in parallel with the drop in protein levels as receptors down-regulate. Treatment of cells with RU 486, an antiprogestin, leads to PR transformation as determined by immunoblotting, but subsequent down-regulation does not occur. After transformation, chromatin-bound RU 486-occupied receptors become intensely phosphorylated however, with specific activities 15 times greater than those of untransformed PR. Since these receptors are phosphorylated but not processed, the hormone-induced nuclear phosphorylation of PR is unlikely to be a signal for receptor processing.(ABSTRACT TRUNCATED AT 400 WORDS)