Phosphorylation by Akt within the ST loop of AMPK-α1 down-regulates its activation in tumour cells

The insulin/IGF-1 (insulin-like growth factor 1)-activated protein kinase Akt (also known as protein kinase B) phosphorylates Ser(487) in the ‘ST loop’ (serine/threonine-rich loop) within the C-terminal domain of AMPK-α1 (AMP-activated protein kinase-α1), leading to inhibition of phosphorylation by upstream kinases at the activating site, Thr(172). Surprisingly, the equivalent site on AMPK-α2, Ser(491), is not an Akt target and is modified instead by autophosphorylation. Stimulation of HEK (human embryonic kidney)-293 cells with IGF-1 caused reduced subsequent Thr(172) phosphorylation and activation of AMPK-α1 in response to the activator A769662 and the Ca(2+) ionophore A23187, effects we show to be dependent on Akt activation and Ser(487) phosphorylation. Consistent with this, in three PTEN (phosphatase and tensin homologue deleted on chromosome 10)-null tumour cell lines (in which the lipid phosphatase PTEN that normally restrains the Akt pathway is absent and Akt is thus hyperactivated), AMPK was resistant to activation by A769662. However, full AMPK activation could be restored by pharmacological inhibition of Akt, or by re-expression of active PTEN. We also show that inhibition of Thr(172) phosphorylation is due to interaction of the phosphorylated ST loop with basic side chains within the αC-helix of the kinase domain. Our findings reveal that a previously unrecognized effect of hyperactivation of Akt in tumour cells is to restrain activation of the LKB1 (liver kinase B1)–AMPK pathway, which would otherwise inhibit cell growth and proliferation

Androgen receptor phosphorylation at serine 515 by Cdk1 predicts biochemical relapse in prostate cancer patients

<br>Background:Prostate cancer cell growth is dependent upon androgen receptor (AR) activation, which is regulated by specific kinases. The aim of the current study is to establish if AR phosphorylation by Cdk1 or ERK1/2 is of prognostic significance.</br> <br>Methods: Scansite 2.0 was utilised to predict which AR sites are phosphorylated by Cdk1 and ERK1/2. Immunohistochemistry for these sites was then performed on 90 hormone-naive prostate cancer specimens. The interaction between Cdk1/ERK1/2 and AR phosphorylation was investigated in vitro using LNCaP cells.</br><br>Results:Phosphorylation of AR at serine 515 (pAR(S515)) and PSA at diagnosis were independently associated with decreased time to biochemical relapse. Cdk1 and pCdk1(161), but not ERK1/2, correlated with pAR(S515). High expression of pAR(S515) in patients with a PSA at diagnosis of ≤20 ng ml(-1) was associated with shorter time to biochemical relapse (P=0.019). This translated into a reduction in disease-specific survival (10-year survival, 38.1% vs 100%, P<0.001). In vitro studies demonstrated that treatment with Roscovitine (a Cdk inhibitor) caused a reduction in pCdk1(161) expression, pAR(S515)expression and cellular proliferation.</br> <br>Conclusion: In prostate cancer patients with PSA at diagnosis of ≤20 ng ml(-1), phosphorylation of AR at serine 515 by Cdk1 may be an independent prognostic marker.</br&gt

The biological mechanisms of resistance in selected lines of Nicotiana tabacum L. to Myzus nicotianae Blackman (Homoptera : Aphididae)

Since 1993, several aphid-resistant breeding lines of tobacco, Nicotiana tabacum L. have been developed at the Tobacco Experiment Station, Greeneville, TN. Crosses were developed between TI 1068, a resistant to the tobacco aphid, Myzus nicotianae Blackman, and ‘TN 86’ or ‘KY 17,’ two tobacco cultivars susceptible to the tobacco aphid. A research project was conducted to: 1) reevaluate levels of resistance of selected tobacco lines to the tobacco aphid, 2) determine the population growth parameters of the tobacco aphis that are affected by resistant tobacco, and 3) simulate aphid population growth on resistant and non-resistant tobacco using a computer-generated model. In 1996 & 1997, several entries were evaluated for aphid population growth in open field plots, greenhouse, and growth room. Field plots were allowed to become infested naturally by aphids. In greenhouse and growth experiments, adult aphids were placed individually onto each plant and allowed to colonize. Results indicated high resistance to the tobacco aphid on TI 1068 for a variety of experiments, compared to other resistant tobacco lines. Initially 3 aphid-resistant tobacco entries (TI 1068, 301, 3001) were evaluated to determine their mechanisms of resistance. Four aphid growth parameters (life cycle, fecundity, reproductive longevity, survival) were investigated for their effects on aphid populations. Aphid development was significantly slower on leaf discs of TI 1068, 301, and 3001 than on TN leaf discs. Aphid development, fecundity, reproductive longevity, and survival did not differ significantly among all tobacco entries tested when reared on excised leaves from greenhouse-grown plants. Aphids reared on leaves excised from field-grown TI 1068 plants had a longer life cycle, lower reproductive rate, shorter reproductive longevity, and shorter survival time than aphids reared on the standard cultivar, TN 86. A computer simulation, based on data from laboratory experiments,was developed to predict seasonal population development of tobacco aphids on TN 86 and TI 1068 aphid populations under field conditions, but in the absence of limiting factors such as weather, predators, disease, and parasitoids. It was estimated using this model that progeny from one aphid on TN 86 could exceed to 10,000,000 individuals within 40 days; whereas, less than 700 aphids would develop on TI 1068. Future work on the effects of weather, predators, diseases, and parasitoids on an aphid populations could prove useful in more accurately predicting aphid population growth in a typical tobacco field

ANIA:ANnotation and Integrated Analysis of the 14-3-3 interactome

The dimeric 14-3-3 proteins dock onto pairs of phosphorylated Ser and Thr residues on hundreds of proteins, and thereby regulate many events in mammalian cells. To facilitate global analyses of these interactions, we developed a web resource named ANIA: ANnotation and Integrated Analysis of the 14-3-3 interactome, which integrates multiple data sets on 14-3-3-binding phosphoproteins. ANIA also pinpoints candidate 14-3-3-binding phosphosites using predictor algorithms, assisted by our recent discovery that the human 14-3-3-interactome is highly enriched in 2R-ohnologues. 2R-ohnologues are proteins in families of two to four, generated by two rounds of whole genome duplication at the origin of the vertebrate animals. ANIA identifies candidate ‘lynchpins’, which are 14-3-3-binding phosphosites that are conserved across members of a given 2R-ohnologue protein family. Other features of ANIA include a link to the catalogue of somatic mutations in cancer database to find cancer polymorphisms that map to 14-3-3-binding phosphosites, which would be expected to interfere with 14-3-3 interactions. We used ANIA to map known and candidate 14-3-3-binding enzymes within the 2R-ohnologue complement of the human kinome. Our projections indicate that 14-3-3s dock onto many more human kinases than has been realized. Guided by ANIA, PAK4, 6 and 7 (p21-activated kinases 4, 6 and 7) were experimentally validated as a 2R-ohnologue family of 14-3-3-binding phosphoproteins. PAK4 binding to 14-3-3 is stimulated by phorbol ester, and involves the ‘lynchpin’ site phosphoSer99 and a major contribution from Ser181. In contrast, PAK6 and PAK7 display strong phorbol ester-independent binding to 14-3-3, with Ser113 critical for the interaction with PAK6. These data point to differential 14-3-3 regulation of PAKs in control of cell morphology. Database URL: https://ania-1433.lifesci.dundee.ac.uk/prediction/webserver/index.p

NetworKIN: a resource for exploring cellular phosphorylation networks

Protein kinases control cellular responses by phosphorylating specific substrates. Recent proteome-wide mapping of protein phosphorylation sites by mass spectrometry has discovered thousands of in vivo sites. Systematically assigning all 518 human kinases to all these sites is a challenging problem. The NetworKIN database (http://networkin.info) integrates consensus substrate motifs with context modelling for improved prediction of cellular kinase–substrate relations. Based on the latest human phosphoproteome from the Phospho.ELM and PhosphoSite databases, the resource offers insight into phosphorylation-modulated interaction networks. Here, we describe how NetworKIN can be used for both global and targeted molecular studies. Via the web interface users can query the database of precomputed kinase–substrate relations or obtain predictions on novel phosphoproteins. The database currently contains a predicted phosphorylation network with 20 224 site-specific interactions involving 3978 phosphoproteins and 73 human kinases from 20 families.Genome Canada (through Ontario Genomics Institute)National Institutes of Health (U.S.) (U54-CA112967)National Institutes of Health (U.S.) (GM60594)European Community’s Human Potential Programme (BioSapiens Network of Excellence (contract number LSHG-CT-2003-503265))European Community’s Human Potential Programme (ADIT Integrated Project (contract number LSHB-CT-2005511065)

Is Honesty the Best Policy? Examining the Effect of Product Safety Communication on Blame Attributions in Causal Chains

This paper extends research on attribution theory through three studies examining how the accuracy and explicitness of product safety information communicated to various entities within a causal chain influences blame attributions after an accident. Unlike prior research, we find consistent evidence that entities in the causal chain were able to limit blame attributions by communicating safety information that’s quality met or exceeded the quality of information available to that entity. Entities did not, however, benefit from providing more accurate information than what had been communicated to them by previous members of the causal chain. This insight suggests that the controllability of information communicated played an important role in the relationship between accurate communication and blame attributions. Our findings provide meaningful insight into steps that organizations can take to limit their potential for receiving blame following an accident, helping to bridge the gap between basic and applied research