Basic molecular events in wild type and mutant staphylococcal nuclease folding

1998 ◽  
Author(s):  
Zhengding Su
Keyword(s):  
Staphylococcal Nuclease ◽  
Wild Type ◽  
Molecular Events

scholarly journals Phosphoinositide 3′-Kinase γ Facilitates Polyomavirus Infection

Viruses ◽  
2020 ◽  
Vol 12 (10) ◽  
pp. 1190
Author(s):  
Paul Clark ◽  
Gretchen V. Gee ◽  
Brandon S. Albright ◽  
Benedetta Assetta ◽  
Ying Han ◽  
...  
Keyword(s):  
Cellular Proteins ◽  
Regulatory Subunit ◽  
Wild Type ◽  
Jc Polyomavirus ◽  
Knock Out ◽  
Molecular Events ◽  
Tumor Viruses ◽  
Phosphoinositide 3 Kinase ◽  
Serious Disease ◽  
Human Polyomaviruses

Polyomaviruses are small, non-enveloped DNA tumor viruses that cause serious disease in immunosuppressed people, including progressive multifocal leukoencephalopathy (PML) in patients infected with JC polyomavirus, but the molecular events mediating polyomavirus entry are poorly understood. Through genetic knockdown approaches, we identified phosphoinositide 3′-kinase γ (PI3Kγ) and its regulatory subunit PIK3R5 as cellular proteins that facilitate infection of human SVG-A glial cells by JCPyV. PI3Kα appears less important for polyomavirus infection than PI3Kγ. CRISPR/Cas9-mediated knockout of PIK3R5 or PI3Kγ inhibited infection by authentic JCPyV and by JC pseudovirus. PI3Kγ knockout also inhibited infection by BK and Merkel Cell pseudoviruses, other pathogenic human polyomaviruses, and SV40, an important model polyomavirus. Reintroduction of the wild-type PI3Kγ gene into the PI3Kγ knock-out SVG-A cells rescued the JCPyV infection defect. Disruption of the PI3Kγ pathway did not block binding of JCPyV to cells or virus internalization, implying that PI3Kγ facilitates some intracellular step(s) of infection. These results imply that agents that inhibit PI3Kγ signaling may have a role in managing polyomavirus infections.

Gene fusions evidence in a KIT/PDGFRA wild-type GIST without mutations in SDH units identified by a whole transcriptome study.

2013 ◽  
Vol 31 (15_suppl) ◽  
pp. e21523-e21523
Author(s):  
Milena Urbini ◽  
Annalisa Astolfi ◽  
Valentina Indio ◽  
Maristella Saponara ◽  
Margherita Nannini ◽  
...  
Keyword(s):  
Gene Fusion ◽  
Surgical Removal ◽  
Erk Pathway ◽  
Gene Fusions ◽  
Rna Seq ◽  
Wild Type ◽  
Rna Pol Ii ◽  
Molecular Events ◽  
Frame Fusion ◽  
Whole Transcriptome

e21523 Background: A subset of KIT/PDGFRA wild-type GIST (WT) harbour mutations in SDH units. In the majority of the remaining cases of WT GIST no other molecular events are identified.We performed a RNA-seq in a WT GIST without mutations in SDH genes using next generation approach to discover molecular events in this GIST population. Methods: In 2003, a 63-year old woman underwent surgery for an ileal GIST (size 6 cm, MI 6/50HPF).After 6 years, she developed a recurrence with a single hepatic lesion. The KIT and PDGFRA analysis of the lesion did not show mutations. Therefore, she did not receive imatinib but she underwent a surgical removal. The analysis of all SDH units did not show mutations. So paired-end RNA-seq (75X2) was performed with Illumina HiScanSQ platform. After mapping the short reads on the human genome(HG19), SNVs and InDels were called by SNVMix2 with an accurate filtering procedures including predictors of mutations effect at protein level. Gene fusions discovery was done considering the agreement between DeFuse, ChimeraScan and FusionMap tools and validated by SangerSequencing using primers spanning the mRNA breakpoints. Results: Four different gene fusions and 206 non-synonymous SNVs were discovered, of which 62 were called deleterious by at least one predictor, and they are undergoing further validation. SPRED2-NELFCD gene fusion originated from an interchromosomal translocation-inversion between chr 20 and 2. The event involved exon1 of SPRED2 and exon11 of NELFCD, probably leading to inactivation of both genes. NELFCD encodes a component of the NELF complex that negatively regulates transcription elongation by RNA pol II, while SPRED2 is a member of the Sprouty /SPRED family that repress growth factor-induced activation of the MAPK/ERK pathway. The other three events were intrachromosomal aberrations: MARK2-PPFIA1 and PLA2G16-ATL3 on chr 11 and ASCC1-C10orf11 on chr 10. Only the first event led to an in-frame fusion (MARK2 ex1- PPFIA1 ex2) probably dysregulating the expression of the downstream gene. Conclusions: This is the first evidence of gene fusions in GIST. The oncogenetic role and the tumor frequency of these events deserve to be studied.

Myogenesis in Wilms' Tumors is Associated with Mutations of the WT1 Gene and Activation of Bcl-2 and the Wnt Signaling Pathway

2004 ◽  
Vol 7 (2) ◽  
pp. 125-137 ◽  
Author(s):  
Ryuji Fukuzawa ◽  
Rosemary W. Heathcott ◽  
Makoto Sano ◽  
Ian M. Morison ◽  
Kankatsu Yun ◽  
...  
Keyword(s):  
Wnt Signaling ◽  
Signaling Pathway ◽  
Cytoplasmic Membrane ◽  
Wnt Signaling Pathway ◽  
Nuclear Accumulation ◽  
Beta Catenin ◽  
Wild Type ◽  
Molecular Events ◽  
Epithelial Development

Wilms tumors with WT1 mutations [ WT1(—)] have a stromal-predominant histology with varying extents of rhabdomyogenesis. These tumors also frequently have mutations in the beta-catenin gene (CTNNB1). We have investigated the molecular events that may explain the origins of rhabdomyogenesis in WT1(-) tumors. Of 35 Wilms tumors, we identified 12 with WT1 mutations, of which 9 carried CTNNB1 mutations. We compared WT1 wild-type tumors [ WT1(+)] with WT1(-) tumors for histological features, localization of beta-catenin, Bcl-2 expression, and apoptosis using an in-situ end-labeling technique. WT1(+) tumors showed triphasic and blast-emal- and epithelial predominant-histology. Expression of WT1, beta-catenin, and Bcl-2 recapitulated those of normal kidney epithelial development. Localization of beta-catenin was observed in the cytoplasm and cytoplasmic membrane of early glomerular epithelial structures. Bcl-2 is also expressed in condensing blastema and early glomerular epithelial structures which had little apoptosis. WT1(-) tumors, regardless of whether CTNNB1 mutations were detected or not, showed a stromal-rich phenotype with abundant expression of beta-catenin in the nucleus of the rhabdomyoblasts. Bcl-2 was expressed in rhabdomyoblasts, but not in blastemal cells undergoing apoptosis, suggesting that WT1 regulates Bcl-2 positively in the epithelial pathway, but negatively in the myogenic pathway. These data indicate that mutations in WT1 might alter the Wnt signaling pathway and Bcl-2 related-apoptosis. In WT1(-) tumors, the nuclear accumulation of beta-catenin and Bcl-2 expression are associated with rhabdomyogenesis, and dysregulation of Bcl-2 may be a mechanism by which the histogenesis (loss of blastemal component, muscle differentiation) may be explained.

scholarly journals Altered cell cycle kinetics, gene expression, and G1 restriction point regulation in Rb-deficient fibroblasts.

1996 ◽  
Vol 16 (5) ◽  
pp. 2402-2407 ◽  
Author(s):  
R E Herrera ◽  
V P Sah ◽  
B O Williams ◽  
T P Mäkelä ◽  
R A Weinberg ◽  
...  
Keyword(s):  
Cell Cycle ◽  
Cyclin E ◽  
Growth Cycle ◽  
Wild Type ◽  
Restriction Point ◽  
Protein Levels ◽  
Selective Growth Advantage ◽  
Molecular Events ◽  
Rb Gene ◽  
Mutant Cells

Fibroblasts prepared from retinoblastoma (Rb) gene-negative mouse embryos exhibit a shorter G1 phase of the growth cycle and smaller size than wild-type cells. In addition, the mutant cells are no longer inhibited by low levels of cycloheximide at any point in G1 but do remain sensitive to serum withdrawal until late in G1. Certain cell cycle-regulated genes showed no temporal or quantitative differences in expression. In contrast, cyclin E expression in Rb-deficient cells is deregulated in two ways. Cyclin E mRNA is generally derepressed in mutant cells and reaches peak levels about 6 h earlier in G1 than in wild-type cells. Moreover, cyclin E protein levels are higher in the Rb-/- cells than would be predicted from the levels of its mRNA. Thus, the selective growth advantage conferred by Rb gene deletion during tumorigenesis may be explained in part by changes in the regulation of cyclin E. In addition, the mechanisms defining the restriction point of late G1 may consist of at least two molecular events, one cycloheximide sensitive and pRb dependent and the other serum sensitive and pRb independent.

scholarly journals Absence of Rybp Compromises Neural Differentiation of Embryonic Stem Cells

2016 ◽  
Vol 2016 ◽  
pp. 1-12 ◽  
Author(s):  
Gergo Kovacs ◽  
Viktoria Szabo ◽  
Melinda K. Pirity
Keyword(s):  
Stem Cells ◽  
Embryonic Stem Cells ◽  
Neural Differentiation ◽  
Embryonic Stem ◽  
Wild Type ◽  
Altered Gene Expression ◽  
Molecular Events ◽  
Altered Gene ◽  
Polycomb Repressive Complex 1

Rybp (Ring1 and Yy1 Binding Protein) is a transcriptional regulator and member of the noncanonical polycomb repressive complex 1 with essential role in early embryonic development. We have previously described that alteration of Rybp dosage in mouse models induced striking neural tube defects (NTDs), exencephaly, and disorganized neurocortex. In this study we further investigated the role of Rybp in neural differentiation by utilising wild type (rybp+/+) andrybp nullmutant (rybp-/-) embryonic stem cells (ESCs) and tried to uncover underlying molecular events that are responsible for the observed phenotypic changes. We found thatrybp nullmutant ESCs formed less matured neurons, astrocytes, and oligodendrocytes from existing progenitors than wild type cells. Furthermore, lack ofrybpcoincided with altered gene expression of key neural markers including Pax6 and Plagl1 pinpointing a possible transcriptional circuit among these genes.

scholarly journals V600E BRAF mutations are alternative early molecular events in a subset of KIT/PDGFRA wild-type gastrointestinal stromal tumours

2009 ◽  
Vol 62 (7) ◽  
pp. 613-616 ◽  
Author(s):  
A Agaimy ◽  
L M Terracciano ◽  
S Dirnhofer ◽  
L Tornillo ◽  
A Foerster ◽  
...  
Keyword(s):  
Wild Type ◽  
Gastrointestinal Stromal Tumours ◽  
Braf Mutations ◽  
Molecular Events

Pressure-jump Relaxation Kinetics of Unfolding and Refolding Transitions of Staphylococcal Nuclease and Proline Isomerization Mutants

1996 ◽  
Author(s):  
Gedlminas J. A. Vidugiris ◽  
Raj Thomas
Keyword(s):  
Globular Proteins ◽  
Staphylococcal Nuclease ◽  
Site Directed Mutagenesis ◽  
Pressure Jump ◽  
Wild Type ◽  
Relaxation Kinetics ◽  
Signal Activity ◽  
In The Wild ◽  
Pressure Jump Relaxation ◽  
Kinetics Of

We present here the first report of the pressure dependence of pressure-jump relaxation kinetics for protein folding transitions. We have studied the relaxation kinetics for the unfolding/refolding of wild-type Staphylococcal nuclease and have found that the relaxation kinetics observed at high pressure are much slower than those observed by pH or denaturant jumps at atmospheric pressure. This indicates that these processes have large, positive values for the activation volumes, most likely stemming from exclusion of solvent from a transition state that is less well packed than the native state. We examined the pressure-jump relaxation kinetics of three single-site mutations in nuclease that lead to alterations in the interactions between the two domains of the protein and changes in the equilibrium constant for isomerization of the lysine-116 to proline 117 peptide bond away from the cis form that predominates in the wild-type enzyme. At comparable pressures, the relaxation times for these mutants were significantly shorter than those observed for the wild type, indicating lower values of the activation volumes. We propose that these mutations cause a decrease in the cooperativity of the unfolding of the two domains, leading to a decrease in the degree of solvent exclusion at the rate-limiting step. The mechanism by which a particular amino acid sequence determines the fold and stability of globular proteins remains one of the most interesting and important unresolved issues in biophysical chemistry. The approaches to increasing our understanding of this phenomenon typically have involved perturbation of the proteins by chemical means or by temperature extremes. The equilibrium or time-dependent responses to these perturbations are then monitored (using a spectroscopic signal, activity, or some other observable) to extract the energetic or kinetic aspects of the unfolding or refolding transitions. Another means of perturbing the system is to modify the protein itself, either chemically or by site-directed mutagenesis, and to assess the effects of modification on the equilibrium or kinetic folding or refolding profiles. This approach has generated a great deal of information about small globular proteins that denature reversibly.

scholarly journals Endothelial cells isolated from caveolin-2 knockout mice display higher proliferation rate and cell cycle progression relative to their wild-type counterparts

2010 ◽  
Vol 298 (3) ◽  
pp. C693-C701 ◽  
Author(s):  
Leike Xie ◽  
Philippe G. Frank ◽  
Michael P. Lisanti ◽  
Grzegorz Sowa
Keyword(s):  
Cell Cycle ◽  
Cell Cycle Progression ◽  
Kinase Inhibitors ◽  
Fold Increase ◽  
S Phase ◽  
Lower Percentage ◽  
Wild Type ◽  
Cycle Progression ◽  
Molecular Events

The goal of this study was to determine whether caveolin-2 (Cav-2) is capable of controlling endothelial cell (EC) proliferation in vitro. To realize this goal, we have directly compared proliferation rates and cell cycle-associated signaling proteins between lung ECs isolated from wild-type (WT) and Cav-2 knockout (KO) mice. Using three independent proliferation assays, we have determined that Cav-2 KO ECs proliferate by ca. 2-fold faster than their WT counterparts. Cell cycle analysis by flow cytometry of propidium iodide-stained cells showed a relatively higher percentage of Cav-2 KO ECs in S and G2/M and lower percentage in Go/G1 phases of cell cycle relative to their WT counterparts. Furthermore, an over 2-fold increase in the percentage of S phase-associated Cav-2 KO relative to WT ECs was independently determined with bromodeoxyuridine incorporation assay. Mechanistically, the increase in proliferation/cell cycle progression of Cav-2 KO ECs correlated well with elevated expression levels of predominantly S phase- and G2/M phase-associated cyclin A and B1, respectively. Further mechanistic analysis of molecular events controlling cell cycle progression revealed increased level of hyperphosphorylated (inactive) form of G1 to S phase transition inhibitor, the retinoblastoma protein in hyperproliferating Cav-2 KO ECs. Conversely, the expression level of the two cyclin-dependent kinase inhibitors p16INK4 and p27Kip1 was reduced in Cav-2 KO ECs. Finally, increased phosphorylation (activation) of proproliferative extracellular signal-regulated kinase 1/2 was observed in hyperproliferating Cav-2 KO ECs. Overall, our data suggest that Cav-2 negatively regulates lung EC proliferation and cell cycle progression.

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