Significance of MDM2 protein in the cell cycle

The basis of oncogenesis underlies the modification of the control of the cell cycle, which leads to disturb balance between proliferation and apoptosis. The MDM2 protein suppresses the ability of p53 to activate genes responsible for repairing or apoptosis, but also promotes p53 degradation by ubiquitination. MDM2 inhibits tumor suppressor property of pRb, by releasing E2F1, which stimulates DNA synthesis in S-phase. MDM2 influences on the neuronal and muscle differentiation. Quantity and stability of the MDM2 protein is regulated by p73, p53, TSG101, p14ARF and Ras-Raf-MEK-ERK pathway. Changes of the level of the MDM2 can disturb control of cell cycle and contribute to oncogenesis.     

A homozygous nonsense mutation (428G-->A) in the human secretor (FUT2) gene provides resistance to symptomatic norovirus (GGII) infections

Noroviruses (formerly Norwalk-like viruses) are a major cause of acute gastroenteritis worldwide and are associated with a significant number of nosocomial and food-borne outbreaks. In this study we show that the human secretor FUT2 gene, which codes for an alpha(1,2)-fucosyltransferase synthesizing the H-type 1 antigen in saliva and mucosa, is associated with susceptibility to norovirus infections. Allelic polymorphism characterization at nucleotide 428 for symptomatic (n = 53) and asymptomatic (n = 62) individuals associated with nosocomial and sporadic norovirus outbreaks revealed that homozygous nonsense mutation (428G-->A) in FUT2 segregated with complete resistance for the disease. Of all symptomatic individuals, 49% were homozygous (SeSe) and 51% heterozygous (Sese428) secretors, and none were secretor negative (se428se428), in contrast to 20% nonsecretors (se428se428) among Swedish blood donors (n = 104) (P < 0.0002) and 29% for asymptomatic individuals associated with nosocomial outbreaks (P < 0.00001). Furthermore, saliva from secretor-positive and symptomatic patients but not from secretor-negative and asymptomatic individuals bound the norovirus strain responsible for that particular outbreak. This is the first report showing that the FUT2 nonsecretor (se428se428) genotype is associated with resistance to nosocomial and sporadic outbreaks with norovirus.     

Role of the septin ring in the asymmetric localization of proteins at the mother-bud neck in Saccharomyces cerevisiae

In the yeast Saccharomyces cerevisiae, septins form a scaffold in the shape of a ring at the future budding site that rearranges into a collar at the mother-bud neck. Many proteins bind asymmetrically to the septin collar. We found that the protein Bni4-CFP was located on the exterior of the septin ring before budding and on the mother side of the collar after budding, whereas the protein kinase Kcc4-YFP was located on the interior of the septin ring before budding and moved into the bud during the formation of the septin collar. Unbudded cells treated with the actin inhibitor latrunculin-A assembled cortical caps of septins on which Bni4-CFP and Kcc4-YFP colocalized. Bni4-CFP and Kcc4-YFP also colocalized on cortical caps of septins found in strains deleted for the genes encoding the GTPase activating proteins of Cdc42 (RGA1, RGA2, and BEM3). However, Bni4-CFP and Kcc4-YFP were still partially separated in mutants (gin4, elm1, cla4, and cdc3-1) in which septin morphology was severely disrupted in other ways. These observations provide clues to the mechanisms for the asymmetric localization of septin-associated proteins.     

The unique N-terminus of R-ras is required for Rac activation and precise regulation of cell migration

The Ras family GTPase, R-Ras, elicits important integrin-dependent cellular behaviors such as adhesion, spreading and migration. While oncogenic Ras GTPases and R-Ras share extensive sequence homology, R-Ras induces a distinct set of cellular behaviors. To explore the structural basis for these differences, we asked whether the unique N-terminal 26 amino acid extension of R-Ras was responsible for R-Ras-specific signaling events. Using a 32D mouse myeloid cell line, we show that full-length R-Ras activates Rac and induces Rac-dependent cell spreading. In contrast, truncated R-Ras lacking its first 26 amino acids fails to activate Rac, resulting in reduced cell spreading. Truncated R-Ras also stimulates more beta3 integrin-dependent cell migration than full-length R-Ras, suggesting that the N-terminus may negatively regulate cell movement. However, neither the subcellular localization of R-Ras nor its effects on cell adhesion are affected by the presence or absence of the N-terminus. These results indicate that the N-terminus of R-Ras positively regulates specific R-Ras functions such as Rac activation and cell spreading but negatively regulates R-Ras-mediated cell migration.     

The PCH family member MAYP/PSTPIP2 directly regulates F-actin bundling and enhances filopodia formation and motility in macrophages

Macrophage actin-associated tyrosine phosphorylated protein (MAYP) belongs to the Pombe Cdc15 homology (PCH) family of proteins involved in the regulation of actin-based functions including cell adhesion and motility. In mouse macrophages, MAYP is tyrosine phosphorylated after activation of the colony-stimulating factor-1 receptor (CSF-1R), which also induces actin reorganization, membrane ruffling, cell spreading, polarization, and migration. Because MAYP associates with F-actin, we investigated the function of MAYP in regulating actin organization in macrophages. Overexpression of MAYP decreased CSF-1-induced membrane ruffling and increased filopodia formation, motility and CSF-1-mediated chemotaxis. The opposite phenotype was observed with reduced expression of MAYP, indicating that MAYP is a negative regulator of CSF-1-induced membrane ruffling and positively regulates formation of filopodia and directional migration. Overexpression of MAYP led to a reduction in total macrophage F-actin content but was associated with increased actin bundling. Consistent with this, purified MAYP bundled F-actin and regulated its turnover in vitro. In addition, MAYP colocalized with cortical and filopodial F-actin in vivo. Because filopodia are postulated to increase directional motility by acting as environmental sensors, the MAYP-stimulated increase in directional movement may be at least partly explained by enhancement of filopodia formation.     

Cold-plasma modification of oxide surfaces for covalent biomolecule attachment

While many processes have been developed to modify the surface of glass and other oxides for biomolecule attachment, they rely primarily upon wet chemistry and are costly and time-consuming. We describe a process that uses a cold plasma and a subsequent in vacuo vapor-phase reaction to terminate a variety of oxide surfaces with epoxide chemical groups. These epoxide groups can react with amine-containing biomolecules, such as proteins and modified oligonucleotides, to form strong covalent linkages between the biomolecules and the treated surface. The use of a plasma activation step followed by an in vacuo vapor-phase reaction allows for the precise control of surface functional groups, rather than the mixture of functionalities normally produced. By maintaining the samples under vacuum throughout the process, adsorption of contaminants is effectively eliminated. This process modifies a range of different oxide surfaces, is fast, consumes a minimal amount of reagents, and produces attachment densities for bound biomolecules that are comparable to or better than commercially available substrates.     

The family feud: do proteins with similar structures fold via the same pathway?

Theoretical and experimental studies of protein folding have suggested that the topology of the native state may be the most important factor determining the folding pathway of a protein, independent of its specific amino acid sequence. To test this concept, many experimental studies have been carried out with the aim of comparing the folding pathways of proteins that possess similar tertiary structures, but divergent sequences. Many of these studies focus on quantitative comparisons of folding transition state structures, as determined by Phi(f) value analysis of folding kinetic data. In some of these studies, folding transition state structures are found to be highly conserved, whereas in others they are not. We conclude that folds displaying more conserved transition state structures may have the most restricted number of possible folding pathways and that folds displaying low transition state structural conservation possess many potential pathways for reaching the native state.     

Parallel synthesis of pteridine derivatives as potent inhibitors for hepatitis C virus NS5B RNA-dependent RNA polymerase

From compound library screening using an HCV NS5B RNA-dependent RNA polymerase enzymatic assay, we identified a pteridine hit compound with an IC(50) of 15 microM. Our SAR studies were focused on the different groups at the 6- and 7-positions, substitutions at the 4-position, and replacement of N(1) or N(3) with carbon in the pteridine ring. We found that NH or OH at 4-position is critical for the inhibitory activity. Furthermore, a hydrophobic substituent at the 4-position may help compounds permeate through the cell membrane.     

Androgynous rex - the utility of chevrons for determining the sex of crocodilians and non-avian dinosaurs

The sex of non-avian dinosaurs has been inferred on numerous occasions using a variety of anatomical criteria, but the efficacy of none has been proven. Nearly 50 years ago Romer suggested that the cranial-most or first chevron in the tails of some reptiles, including crocodilians, is sexually dimorphic. Recent work on this subject purportedly substantiated that the female first chevron articulates in a more caudal position than in males. Furthermore, it was concluded that this element is shorter in females. These phenotypic attributes theoretically provide a broader cloacal passageway for eggs by ovipositing females and a greater attachment area for male “penile retractor muscles”. Because theropod dinosaurs such as Tyrannosaurus rex presumably show similar variation in chevron anatomy, the same criteria has been advocated for sexing dinosaurs. We tested the neontological model for the chevron sexual dimorphism hypothesis using a skeletonized growth series of American alligators (Alligator mississippiensis) of known sex. No statistical support for the hypothesis was found. Furthermore, analysis of a diversity of crocodilian taxa from museum collections revealed similar findings suggesting the alligator results are not taxon specific. Study of well-preserved tyrannosaurid dinosaurs in museum collections showed nearly invariant chevron positioning like that seen in crocodilians. This suggests the usefulness of chevron anatomy for sexing dinosaurs is tenuous.     

Phylogenetic analysis of ecomorphological divergence, community structure, and diversification rates in dusky salamanders (Plethodontidae: Desmognathus)

An important dimension of adaptive radiation is the degree to which diversification rates fluctuate or remain constant through time. Focusing on plethodontid salamanders of the genus Desmognathus, we present a novel synthetic analysis of phylogeographic history, rates of ecomorphological evolution and species accumulation, and community assembly in an adaptive radiation. Dusky salamanders are highly variable in life history, body size, and ecology, with many endemic lineages in the southern Appalachian Highlands of eastern North America. Our results show that life-history evolution had important consequences for the buildup of plethodontid-salamander species richness and phenotypic disparity in eastern North America, a global hot spot of salamander biodiversity. The origin of Desmognathus species with aquatic larvae was followed by a high rate of lineage accumulation, which then gradually decreased toward the present time. The peak period of lineage accumulation in the group coincides with evolutionary partitioning of lineages with aquatic larvae into seepage, stream-edge, and stream microhabitats. Phylogenetic simulations demonstrate a strong correlation between morphology and microhabitat ecology independent of phylogenetic effects and suggest that ecomorphological changes are concentrated early in the radiation of Desmognathus. Deep phylogeographic fragmentation within many codistributed ecomorph clades suggests long-term persistence of ecomorphological features and stability of endemic lineages and communities through multiple climatic cycles. Phylogenetic analyses of community structure show that ecomorphological divergence promotes the coexistence of lineages and that repeated, independent evolution of microhabitat-associated ecomorphs has a limited role in the evolutionary assembly of Desmognathus communities. Comparing and contrasting our results to other adaptive radiations having different biogeographic histories, our results suggest that rates of diversification during adaptive radiation are intimately linked to the degree to which community structure persists over evolutionary time.