The ability of the pathogenic fungus Candida albicans to switch from a yeast to a hyphal morphology in response to external signals is implicated in its pathogenicity. We used glass DNA microarrays to investigate the transcription profiles of 6333 predicted ORFs in cells undergoing this transition and their responses to changes in temperature and culture medium. We have identified several genes whose transcriptional profiles are similar to those of known virulence factors that are modulated by the switch to hyphal growth caused by addition of serum and a 37 degrees C growth temperature. Time course analysis of this transition identified transcripts that are induced before germ tube initiation and shut off later in the developmental process. A strain deleted for the Efg1p and Cph1p transcription factors is defective in hyphae formation, and its response to serum and increased temperature is almost identical to the response of a wild-type strain grown at 37 degrees C in the absence of serum. Thus Efg1p and Cph1p are needed for the activation of the transcriptional program that is induced by the presence of serum.

Loss of CCM1/2 leads to destabilization of ICAP-1 and up-regulation of β1 integrin, resulting in the destabilization of intercellular junctions due to increased cell contractility and aberrant extracellular matrix remodeling.

The prion protein (PrP) propensity to adopt different structures is a clue to its biological role. PrP oligomers have been previously reported to bear prion infectivity or toxicity and were also found along the pathway of in vitro amyloid formation. In the present report, kinetic and structural analysis of ovine PrP (OvPrP) oligomerization showed that three distinct oligomeric species were formed in parallel, independent kinetic pathways. Only the largest oligomer gave rise to fibrillar structures at high concentration. The refolding of OvPrP into these different oligomers was investigated by analysis of hydrogen/deuterium exchange and introduction of disulfide bonds. These experiments revealed that, before oligomerization, separation of contacts in the globular part (residues 127-234) occurred between the S1-H1-S2 domain (residues 132-167) and the H2-H3 bundle (residues 174 -230), implying a conformational change of the S2-H2 loop (residues 168 -173). The type of oligomer to be formed depended on the site where the expansion of the OvPrP monomer was initiated. Our data bring a detailed insight into the earlier conformational changes during PrP oligomerization and account for the diversity of oligomeric entities. The kinetic and structural mechanisms proposed here might constitute a physicochemical basis of prion strain genesis.folding ͉ kinetics ͉ oligomers ͉ strain

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The leukocyte NADPH oxidase catalyzes the production of O(2)(-) from oxygen at the expense of NADPH. Activation of the enzyme requires interaction of the cytosolic factors p47(PHOX), p67(PHOX), and Rac2 with the membrane-associated cytochrome b(558). Activation of the oxidase in a semirecombinant cell-free system in the absence of an amphiphilic activator can be achieved by phosphorylation of the cytosolic factor p47(PHOX) by protein kinase C. Another cytosolic factor, p40(PHOX), was recently shown to be phosphorylated on serine and threonine residues upon activation of NADPH oxidase, but both stimulatory and inhibitory roles were reported. In the present study, we demonstrate that the addition of phosphorylated p40(PHOX) to the cell-free system inhibits NADPH oxidase activated by protein kinase C-phosphorylated p47(PHOX), an effect not observed with the unphosphorylated p40(PHOX). Moreover phosphorylated p40(PHOX) inhibits the oxidase if added before or after full activation of the enzyme. Direct mutagenesis of protein kinase C consensus sites enables us to conclude that phosphorylation of threonine 154 is required for the inhibitory effect of p40(PHOX) to occur. Although the phosphorylated mutants and nonphosphorylated mutants are still able to interact with both p47(PHOX) and p67(PHOX) in pull-down assays, their proteolysis pattern upon thrombin treatment suggests a difference in conformation between the phosphorylated and nonphosphorylated mutants. We postulate that phosphorylation of p40(PHOX) on threonine 154 leads to an inhibitory conformation that shifts the balance toward an inhibitory role and blocks oxidase activation.

A method to separate specific and nonspecific noncovalent interactions observed in ESI mass spectra between a protein and its ligands is presented. Assuming noncooperative binding, the specific ligand binding is modeled as a statistical distribution on identical binding sites. For the nonspecific fraction we assume a statistical distribution on a large number of "nonspecific" interacting sites. The model was successfully applied to the noncovalent interaction between the protein creatine kinase (CK) and its ligands adenosine diphosphate (ADP) and adenosine triphosphate (ATP) that both exhibit nonspecific binding in the mass spectrum. The two sequential dissociation constants obtained by applying our method are K 1,diss ϭ 11.8 Ϯ 1.5 M and K 2,diss ϭ 48 Ϯ 6 M for ADP. For ATP, the constants are K 1,diss ϭ 27 Ϯ 7 M and K 2,diss ϭ 114 Ϯ 27 M. All constants are in good correlation with reported literature values. The model should be valuable for systems with a large dissociation constant that require high ligand concentrations and thus have increased potential of forming nonspecific

The review will focus on how physicochemical properties of ECM might promote tumor growth or cell dissemination or on the contrary maintain quiescent state of cancer cells. It is crucial to clarify the molecular basis of mechanotransduction in the development and progression of tumors to identify new potential biomarkers and anticancer therapeutic targets.