Coupled motions in the SH2 and kinase domains of Csk control Src phosphorylation

The C-terminal Src kinase (Csk) phosphorylates and down-regulates Src family tyrosine kinases. The Csk-binding protein (Cbp) localizes Csk close to its substrates at the plasma membrane, and increases the specific activity of the kinase. To investigate this long-range catalytic effect, the phosphorylation of Src and the conformation of Csk were investigated in the presence of a high-affinity phosphopeptide derived from Cbp. This peptide binds tightly to the SH2 domain and enhances Src recognition (lowers K(m)) by increasing the apparent phosphoryl transfer rate in the Csk active site, a phenomenon detected in rapid quench flow experiments. Previous studies demonstrated that the regulation of Csk activity is linked to conformational changes in the enzyme that can be probed with hydrogen-deuterium exchange methods. We show that the Cbp peptide impacts deuterium incorporation into its binding partner (the SH2 domain), and into the SH2-kinase linker and several sequences in the kinase domain, including the glycine-rich loop in the active site. These findings, along with computational data from normal mode analyses, suggest that the SH2 domain moves in a cantilever fashion with respect to the small lobe of the kinase domain, ordering the active site for catalysis. The binding of a small Cbp-derived peptide to the SH2 domain of Csk modifies these motions, enhancing Src recognition.     

Arylphthalazines as potent,and orally bioavailable inhibitors of VEGFR-2

A series of arylphthalazine derivatives were synthesized and evaluated as antagonists of VEGF receptor II (VEGFR-2). IM-094482 57, which was prepared in two steps from commercially available starting materials, was found to be a potent inhibitor of VEGFR-2 in enzymatic, cellular and mitogenic assays (comparable activity to ZD-6474). Additionally, 57 inhibited the related receptor, VEGF receptor I (VEGFR-1), and showed excellent exposure when dosed orally to female CD-1 mice.     

The Effects of the Organic Flame‐Retardant 1,2‐Dibromo‐4‐(1,2‐dibromoethyl) Cyclohexane (TBECH) on Androgen Signaling in Human Prostate Cancer Cell Lines

The effects of the organic flame retardant 1,2‐Dibromo‐4‐(1,2‐dibromoethyl) cyclohexane (TBECH) on androgen receptor target gene expression were examined in the human LNCaP prostate cancer cell line. While γ‐/δ‐TBECH alone led to a significant increase in prostate‐specific antigen (PSA) mRNA accumulation, both the α‐/‐TBECH and γ‐/δ‐TBECH mixtures repressed androgen‐inducible PSA mRNA and protein accumulation in human LNCaP cells. Thus, we hypothesize that isomeric mixtures of TBECH may act as partial agonists of the androgen receptor.     

Spatial arrangement of kin affects recruitment success in young male red grouse

Models have shown that population cycles might be driven by time lags resulting from positive feedback between kin structure and population change, coupled with negative feedback between density and population change. One such model operates through kin favouritism facilitating the recruitment of young cock red grouse. We investigated whether recruitment by young cocks depended on the presence and spatial arrangement of elder relatives in the territorial population. We used molecular genetic estimates of relatedness, and checked for effects of covariates including natal territory size, hatching date, body size, parasite burdens and local density. Philopatric recruitment by cock red grouse led to the formation of clusters of contiguous territories owned by kin. The probability that an individual young cock would establish a territory increased with the number of kin in his father's cluster. This pattern might have been due to genetic quality determining both recruitment success and the size of the paternal cluster. If so, there should have been a positive correlation between a young cock's probability of recruitment and the number of his relatives in the population, irrespective of their spatial distribution. This did not occur and so the effect of cluster size is unlikely to have been confounded by genetic quality. The only morphological measure correlated with recruitment success was supraorbital comb size. The results are consistent with the prediction that kin tolerance affects recruitment but were at the level of the individual within years, rather than the population among years. Hence an experimental test of the kin favouritism hypothesis for population cycles, by manipulation of relatedness in populations among years, is now required.     

Plasminogen activator inhibitor-1 regulates tumor growth and angiogenesis.

Elevated expression of plasminogen activator inhibitor-1 (PAI-1) in tumors is associated with a poor prognosis in many cancers. Reduced tumor growth and angiogenesis have also been reported in mice deficient in PAI-1. These results suggest that PAI-1 may be required for efficient angiogenesis and tumor growth. In the present study, we demonstrate that PAI-1 can both enhance and inhibit the growth of M21 human melanoma tumors in nude mice and that this appears to be due to PAI-1 regulation of angiogenesis. Quantitative analysis of angiogenesis in a Matrigel implant assay indicated that in PAI-1 null mice angiogenesis was reduced approximately 60% compared with wild-type mice, while in mice overexpressing PAI-1, angiogenesis was increased nearly 3-fold. Furthermore, addition of PAI-1 to implants in wild-type mice enhanced angiogenesis up to 3-fold at low concentrations but inhibited angiogenesis nearly completely at high concentrations. Together, these data demonstrate that PAI-1 is a potent regulator of angiogenesis and hence of tumor growth and suggest that understanding the mechanism of this activity may lead to the development of important new therapeutic agents for controlling pathologic angiogenesis.     

Flushuffle and Fluresort: new algorithms to identify reassorted strains of the influenza virus by mass spectrometry

ABSTRACT: BACKGROUND: Influenza is one of the oldest and deadliest infectious diseases known to man. Reassorted strains of the virus pose the greatest risk to both human and animal health and have been associated with all pandemics of the past century, with the possible exception of the 1918 pandemic, resulting in tens of millions of deaths. We have developed and tested new computer algorithms, FluShuffle and FluResort, which enable reassorted viruses to be identified by the most rapid and direct means possible. These algorithms enable reassorted influenza, and other, viruses to be rapidly identified to allow prevention strategies and treatments to be more efficiently implemented. RESULTS: The FluShuffle and FluResort algorithms were tested with both experimental and simulated mass spectra of whole virus digests. Flu Shuffle considers different combinations of viral protein identities that match the mass spectral data using a Gibbs sampling algorithm employing a mixed protein Markov chain Monte Carlo (MCMC) method. Flu Resort utilizes those identities to calculate the weighted distance of each across two or more different phylogenetic trees constructed through viral protein sequence alignments. Each weighted mean distance value is normalized by conversion to a Z-score to establish a reassorted strain. CONCLUSIONS: The new Flu Shuffle and Flu Resort algorithms can correctly identify the origins of influenza viral proteins and the number of reassortment events required to produce the strains from the high resolution mass spectral data of whole virus proteolytic digestions. This has been demonstrated in the case of constructed vaccine strains as well as common human seasonal strains of the virus. The algorithms significantly improve the capability of the proteotyping approach to identify reassorted viruses that pose the greatest pandemic risk.