NFX1-123 is highly expressed in cervical cancer and increases growth and telomerase activity in HPV 16E6 expressing cells

A significant contributor to women’s cancer mortality worldwide is cervical cancer, which is caused by high-risk human papillomavirus (HR HPV). The two viral oncoproteins of HR HPV, E6 and E7, partner with host cell proteins to target oncogenic proteins and pathways. Previously, we have shown HR HPV type 16 E6 (16E6) interacts with the host protein NFX1-123 to target telomerase and cellular immortalization, requiring NFX1-123 to fully upregulate telomerase activity. We now report that NFX1-123 is highly expressed in primary cervical cancers. In vitro, cells expressing 16E6 and overexpressing NFX1-123 have extended active growth, decreased senescence marker staining, and more rapid cell cycling compared to 16E6 expressing cells with endogenous amounts of NFX1-123. These findings were associated with increased telomerase activity and augmented expression of its catalytic subunit, hTERT. In complement, HPV 16 positive cervical cancer cell lines with knocked down NFX1-123 had slowed growth and reduced hTERT over time. In cells that express HR HPV E6, greater expression of NFX1-123 can modify active cellular growth and augment hTERT expression and telomerase activity over time, potentially supporting the initiation and progression of HPV-associated cancers

HPV type 16 E6 and NFX1-123 Augment JNK Signaling to Mediate Keratinocyte Differentiation and L1 Expression

The HPV life cycle is differentiation-dependent, with cellular differentiation driving initiation of the late, productive stage of the viral life cycle. Here, we identify a role for the protein NFX1-123 in regulating keratinocyte differentiation and events of the late HPV life cycle. NFX1-123 itself increased with differentiation of epithelial cells. Greater NFX1-123 augmented differentiation marker expression and JNK phosphorylation in differentiating 16E6-expressing human foreskin keratinocytes (16E6 HFKs). This was associated with altered expression of MKK4 and MKK7, upstream kinase regulators of JNK phosphorylation. Modulating levels of NFX1-123 in HPV16-positive W12E cells recapitulated the effects on differentiation markers, JNK phosphorylation, and MKK4/7 seen in 16E6 HFKs. Crucially, levels of NFX1-123 also correlated with expression of L1, the capsid protein of HPV. Altogether, these studies define a role for NFX1-123 in mediating epithelial differentiation through the JNK signaling pathway, potentially linking expression of cellular genes and HPV genes during differentiation

Structural insights into what glycan arrays tell us about how glycan-binding proteins interact with their ligands

Screening of glycan arrays represents a powerful, high-throughput approach to defining oligosaccharide ligands for glycan-binding receptors, commonly referred to as lectins. Correlating results from such arrays with structural analysis of receptor–ligand complexes provide one way to validate the arrays. Using examples drawn from the family of proteins that contain C-type carbohydrate-recognition domains, this review illustrates how information from the arrays reflects the way that selectivity and affinity for glycan ligands is achieved. A range of binding profiles is observed, from very restricted binding to a small set of structurally similar ligands to binding of broad classes of ligands with related terminal sugars and even to failure to bind any of the glycans on an array. These outcomes provide insights into the importance of multiple factors in defining the selectivity of these receptors, including the presence of conformationally defined units in some oligosaccharide ligands, local and extended interactions between glycans and the surfaces of receptors, and steric factors that exclude binding of some ligands

Spin effects in single electron tunneling

An important consequence of the discovery of giant magnetoresistance in metallic magnetic multilayers is a broad interest in spin dependent effects in electronic transport through magnetic nanostructures. An example of such systems are tunnel junctions -- single-barrier planar junctions or more complex ones. In this review we present and discuss recent theoretical results on electron and spin transport through ferromagnetic mesoscopic junctions including two or more barriers. Such systems are also called ferromagnetic single-electron transistors. We start from the situation when the central part of a device has the form of a magnetic (or nonmagnetic) metallic nanoparticle. Transport characteristics reveal then single-electron charging effects, including the Coulomb staircase, Coulomb blockade, and Coulomb oscillations. Single-electron ferromagnetic transistors based on semiconductor quantum dots and large molecules (especially carbon nanotubes) are also considered. The main emphasis is placed on the spin effects due to spin-dependent tunnelling through the barriers, which gives rise to spin accumulation and tunnel magnetoresistance. Spin effects also occur in the current-voltage characteristics, (differential) conductance, shot noise, and others. Transport characteristics in the two limiting situations of weak and strong coupling are of particular interest. In the former case we distinguish between the sequential tunnelling and cotunneling regimes. In the strong coupling regime we concentrate on the Kondo phenomenon, which in the case of transport through quantum dots or molecules leads to an enhanced conductance and to a pronounced zero-bias Kondo peak in the differential conductance.Comment: topical review (36 figures, 65 pages), to be published in J. Phys.: Condens. Matte

Morphology and dynamics of inflated subaqueous basaltic lava flows

International audienceDuring eruptions onto low slopes, basaltic Pahoehoe lava can form thin lobes that progressively coalesce and inflate to many times their original thickness, due to a steady injection of magma beneath brittle and viscoelastic layers of cooled lava that develop sufficient strength to retain the flow. Inflated lava flows forming tumuli and pressure ridges have been reported in different kinds of environments, such as at contemporary subaerial Hawaiian-type volcanoes in Hawaii, La Réunion and Iceland, in continental environments (states of Oregon, Idaho, Washington), and in the deep sea at Juan de Fuca Ridge, the Galapagos spreading center, and at the East Pacific Rise (this study). These lava have all undergone inflation processes, yet they display highly contrasting morphologies that correlate with their depositional environment, the most striking difference being the presence of water. Lava that have inflated in subaerial environments display inflation structures with morphologies that significantly differ from subaqueous lava emplaced in the deep sea, lakes, and rivers. Their height is 2-3 times smaller and their length being 10-15 times shorter. Based on heat diffusion equation, we demonstrate that more efficient cooling of a lava flow in water leads to the rapid development of thicker (by 25%) cooled layer at the flow surface, which has greater yield strength to counteract its internal hydrostatic pressure than in subaerial environments, thus limiting lava breakouts to form new lobes, hence promoting inflation. Buoyancy also increases the ability of a lava to inflate by 60%. Together, these differences can account for the observed variations in the thickness and extent of subaerial and subaqueous inflated lava flows

Bioenergy and climate change mitigation: an assessment

Acknowledgements The authors are indebted to Julia Römer for assisting with editing several hundred references. Helmut Haberl gratefully acknowledges funding by the Austrian Academy of Sciences (Global Change Programme), the Austrian Ministry of Science and Research (BMWF, proVision programme) as well as by the EU-FP7 project VOLANTE. Carmenza Robledo-Abad received financial support from the Swiss State Secretariat for Economic Affairs.Peer reviewedPostprin

Cytoplasmic Poly(A) Binding Proteins Regulate Telomerase Activity and Cell Growth in Human Papillomavirus Type 16 E6-Expressing Keratinocytes▿

The high-risk human papillomavirus (HPV) E6 and E7 oncoproteins are critical to the immortalization of keratinocytes. HPV type 16 (HPV16) E6 interacts with endogenous proteins to activate hTERT, the catalytic subunit of telomerase, thus avoiding cellular senescence signals. NFX1-123, the longer splice variant of NFX1, interacts with HPV16 E6, as well as cytoplasmic poly(A) binding proteins 1 and 4 (PABPC1 and PABPC4). HPV16 E6 affects hTERT expression posttranscriptionally through NFX1-123, as NFX1-123 interacts with hTERT mRNA and stabilizes it, leading to greater telomerase activity. The PAM2 motif of NFX1-123, with which it binds PABPCs, is required for the posttranscriptional regulation of hTERT by HPV16 E6 and NFX1-123. There is increasing evidence that RNA and DNA viruses utilize RNA-processing proteins, and specifically PABPCs, in the normal virus life cycle, and there is also evidence that RNA-processing proteins are perturbed in cancers. Here, we show that PABPCs are critical in hTERT regulation by HPV16 E6. Although the amount and cellular localization of PABPCs were largely unchanged in cervical cancer cell lines with or without HPV16 and in human foreskin keratinocytes (HFKs) with or without HPV16 E6, knockdown of PABPCs decreased hTERT mRNA and telomerase activity and overexpression of PABPC4 increased these in HPV16 E6-expressing HFKs. In contrast, knockdown of PABPCs in C33A cells had no effect on hTERT mRNA or telomerase activity. Additionally, overexpression of PABPC4 and hTERT led to greater growth of cultured HPV16 E6-expressing HFKs. This is the first evidence that PABPCs have a targeted role in hTERT regulation leading to a growth advantage in cells expressing HPV16 E6

Human papillomavirus type 16 E6 and NFX1-123 mislocalize immune signaling proteins and downregulate immune gene expression in keratinocytes

<div><p>Human papillomavirus (HPV) is the most prevalent sexually transmitted infection, affecting an estimated 11% of the world’s population. The high-risk HPV types (HR HPV) account for approximately 5% of the global burden of cancer and thus cause high morbidity and mortality. Although it is known that persistent infection with HR HPV is the greatest risk factor for developing HPV-associated cancer, and that the HPV early proteins E6 and E7 dysregulate immune detection by its host cells, the mechanisms of immune evasion by HR HPV are not well understood. Previous work in the laboratory identified the endogenous cytoplasmic host protein NFX1-123 as a binding partner of the HR HPV type 16 oncoprotein E6 (16E6). Together NFX1-123 and 16E6 affect cellular growth, differentiation, and immortalization genes and pathways. In a whole genome microarray, human foreskin keratinocytes (HFKs) stably expressing 16E6 and overexpressing NFX1-123 showed a diverse set of innate immune genes downregulated two-fold or more when compared to 16E6 cells with endogenous NFX1-123. We demonstrated that 16E6 and NFX1-123 decreased expression of pro-inflammatory cytokines and interferon-stimulated genes (ISGs) in 16E6 HFKs at the mRNA and protein level. Knock down of NFX1-123 in 16E6 HFKs resulted in a derepression of innate immune genes, pointing to the requirement of NFX1-123 for immune regulation in the context of 16E6. Studies using immunofluorescent microscopy revealed that 16E6 and NFX1-123 disturbed the normal localization of signaling proteins involved in initiating the immune response. This study identifies NFX1-123 as a critical host protein partner through which 16E6 is able to subvert the immune response and in turn permit a long-lived HR HPV infection.</p></div

Genes and pathways decreased in 16E6/FN123 HFKs.

<p>(A) A subset of the over 200 genes that were decreased in 16E6/FN123 cells compared to 16E6/control. Fold change shown is the average fold change over all 16E6 HFK cell lines in which that gene was decreased. (B) Microarray data were analyzed using GeneSpring GX11.5.1, and the significant Gene Ontology pathways for the collection of genes decreased in two out of three 16E6 HFK cell lines were found. Only pathways with a p-value ≤ 0.001 are shown.</p

Microarray analysis of genes downregulated in 16E6 HFKs with increased NFX1-123.

<p>Whole genome expression microarrays were conducted in HFKs stably expressing 16E6 and overexpressing NFX1-123 or with endogenous levels of NFX1-123. (A) Three biologically independent HFKs were transduced with 16E6 and p53 protein levels assessed. (B) 16E6 HFK lines 1, 2, and 3 were transduced with NFX1-123 overexpression construct (FN123) or vector control (control). Levels of NFX1-123 mRNA and protein expression levels were quantified and compared. All qPCRs were normalized to the housekeeping gene 36B4, and all error bars represent 95% confidence intervals from the technical replicates shown (n = 3). GAPDH = loading control for (A and C). (D) Venn diagram of genes whose average expression was decreased at least two-fold in 16E6/FN123 cells compared to 16E6/control. Box indicates 26 genes represented in (D) (E) Heat map with hierarchical clustering of the 26 genes decreased in 16E6/FN123 cells compared to 16E6/control.</p