A predominantly nuclear protein affecting cytoplasmic localization of β-actin mRNA in fibroblasts and neurons

The localization of β-actin mRNA to the leading lamellae of chicken fibroblasts and neurite growth cones of developing neurons requires a 54-nt localization signal (the zipcode) within the 3′ untranslated region. In this study we have identified and isolated five proteins binding to the zipcode. One of these we previously identified as zipcode binding protein (ZBP)1, a 4-KH domain protein. A second is now investigated in detail: a 92-kD protein, ZBP2, that is especially abundant in extracts from embryonic brain. We show that ZBP2 is a homologue of the human hnRNP protein, KSRP, that appears to mediate pre-mRNA splicing. However, ZBP2 has a 47–amino acid (aa) sequence not present in KSRP. Various portions of ZBP2 fused to GFP indicate that the protein most likely shuttles between the nucleus and the cytoplasm, and that the 47-aa insert promotes the nuclear localization. Expression of a truncated ZBP2 inhibits the localization of β-actin mRNA in both fibroblast and neurons. These data suggest that ZBP2, although predominantly a nuclear protein, has a role in the cytoplasmic localization of β-actin mRNA

Regulation of the G2–M cell cycle progression by the ERK5–NFκB signaling pathway

Elucidation of mechanisms regulating cell cycle progression is of fundamental importance for cell and cancer biology. Although several genes and signaling pathways are implicated in G1–S regulation, less is known regarding the mechanisms controlling cell cycle progression through G2 and M phases. We report that extracellular signal–regulated kinase 5 (ERK5), a member of the mitogen-activated protein kinases, is activated at G2–M and required for timely mitotic entry. Stimulation of ERK5 activated nuclear factor κB (NFκB) through ribosomal S6 kinase 2 (RSK2)-mediated phosphorylation and degradation of IκB. Furthermore, selective inhibition of NFκB at G2–M phases substantially delayed mitotic entry and inhibited transcription of G2–M–specific genes, including cyclin B1, cyclin B2, Plk-1, and cdc25B. Moreover, inhibition of NFκB at G2–M diminished mitosis induced by constitutive activation of ERK5, providing a direct link between ERK5, NFκB, and regulation of G2–M progression. We conclude that a novel ERK5–NFκB signaling pathway plays a key role in regulation of the G2–M progression

S100A4 Knockout Sensitizes Anaplastic Thyroid Carcinoma Cells Harboring BRAFV600E/Mt to Vemurafenib

Background/Aims: Anaplastic thyroid cancer (ATC), with 25% BRAFV600E mutation, is one of the most lethal human malignancies that currently has no effective therapy. Vemurafenib, a BRAFV600E inhibitor, has shown promise in clinical trials, including ATC patients, but is being hampered by the acquisition of drug resistance. Therefore, combination therapy that includes BRAFV600E inhibition and avoids resistance is a clinical need. Methods: ATC cell lines 8505C (BRAFV600E/mt), SW1736 (BRAFV600E/mt), KAT18 (BRAFV600E/wt) and Cal-62(BRAFV600E/wt) cells were used in the study. The ability of S100A knockout or /and in combination with the BRAF inhibitor vemurafenib on growth, apoptosis, invasion and apoptosis in ATC cells in vitro was demonstrated by MTT and BrdUrd incorporation assay, Annexin-V-FITC staining analyzed by flow cytometry, Transwell migration and Matrigel invasion assay. S100A4,pERK1/2, pAKT and pROCK1/2 protein was detected by western blot assay; Small molecule inhibitors of Y27632, U0126, MK-2206 and constitutively active forms of pCDNA-Myc-pERK, pCMV6-HA-Akt, pCMV-RhoA were employed, and the mechanistic studies were performed. We assessed the efficiency of in vivo combination treatment with S100A4 knockout and Vemurafenib on tumors. Results: S100A4 knockout induced apoptosis and reduced proliferation by inactivation of pAKT and pERK signals, and inhibited invasion and migration by inactivation of pAKT and RhoA/ROCK1/2 signals in 8505C or Cal-62 cells in vitro, and vice versa in SW1736 and KAT18 cells. Vemurafenib did not affect apoptosis of both 8505C and SW1736 cells, but reduced proliferation via arresting cell cycle, and promoted cell migration and invasion in vitro. Combination treatment with S100A4 knockdown and vemurafenib reduced cell proliferation, migration and invasion in vitro compared to the S100A4 knockdown or Vemurafenib alone. Vemurafenib treatment resulted in a transient inhibition of pERK expression and gradually activation of pAKT expression, but quickly recovery from ERK1/2 activation inhibition by vemurafenib treatment in 4 h for SW1736 and 8505C cells. Combined treatment completely inhibited ERK1/2 and AKT activation during 48 h. In an in vivo mouse model of SW1736 and 8505C, vemurafenib treatment alone did not significantly inhibit tumor growth in both of the tumors, but inhibited tumor growth in combined groups. Conclusion: Our results show S100A4 knockout alone inhibits ATC cells (rich endogenous S100A4) survival and invasion, regardless of the BRAFV600E status, and potentiates the effect of vemurafenib on tumor regression in vitro and in vivo. In addition, S100A4 knockout potently inhibits the recovery from ERK1/2 activation inhibition and the AKT activation following vemurafenib treatment and reversed the vemurafenib resistance. This therapeutic combination may be of benefit in patients with ATC

Synergistic inhibition of MEK/ERK and BRAF V600E with PD98059 and PLX4032 induces sodium/iodide symporter (NIS) expression and radioiodine uptake in BRAF mutated papillary thyroid cancer cells

Abstract Background and Aims The activating mutation BRAFV600E is a frequent genetic event in papillary thyroid carcinomas (PTC). Mutation BRAFV600E is associated with the loss of a sodium/iodine symporter (NIS), and subsequent radioiodide-refractory (RAI) metastatic disease. Use of BRAF V600E inhibitors could partly restore NIS expression and Iodide uptake by inhibition of mitogen-activated protein kinase (MAPK) pathway. Previous study has reported that the BRAF V600E inhibitors could re-activate MAPK signals. In the present study, we investigated whether the combination treatment of BRAF V600E inhibitor and MAPK signal inhibitor could more effectively increase NIS expression and RAI uptake, and explore the mechanisms. Methods BCPAP and K1 cells were exposed to increasing concentrations of BRAF V600E inhibitor PLX4032 (0.01 μM, 0.1 μM, 1 μM) or MEK/ERK inhibitor PD98059 (0.01 μM, 0.1 μM, 1 μM) or with their association or/and in the presence of 3 mM perchlorate (ClO−  4) for 0–72 h. Iodide uptake and expression of BRAF, phosphorylated (p) ERK1/2, NIS were detected. Results PLX4032 or PD98059 alone did not induce NIS expression and increase Iodide uptake in BCPAP and K1 cells. But combined treatment of PLX4032 and PD98059 significantly induce NIS expression and increase Iodide uptake in BCPAP and K1 cells. PLX4032 alone inhibited p-ERK expression at early time, and re-activated p-ERK expression at late time. However, combined treatment of PLX4032 and PD98059 completely inhibited p-ERK expression. Conclusion Simultaneously suppressing BRAF V600E and p-ERK restored NIS expression and increase Iodide uptake in PTC cells, which was associated the inhibition of p-ERK expression. The results warrants clinical trials to confirm

Assessment of Cu(II) Removal from Aqueous Solutions by Modified Pomelo Peels: Experiments and Modelling

In this study, low-cost pomelo peel wastes were used as a bio-sorbent to remove copper ions (e.g., Cu(II)) from aqueous solutions. Prior to testing its Cu(II) removal capability, the structural, physical and chemical characteristics of the sorbent were examined by scanning electron microscope (SEM), Fourier transform infrared (FTIR) spectroscopy, and Brunauer–Emmett–Teller (BET) surface area analysis. The impacts of the initial pH, temperature, contact time and Cu(II) feed concentration on the Cu(II) biosorption using modified pomelo peels were then assessed. Thermodynamic parameters associated to the biosorption clearly demonstrate that this biosorption is thermodynamically feasible, endothermic, spontaneous and entropy driven. Furthermore, adsorption kinetic data were found to fit very well with the pseudo-second order kinetics equation, highlighting that this process is driven by a chemical adsorption. Finally, an artificial neural network with a 4:9:1 structure was then established for describing the Cu(II) adsorption using modified pomelo peels with R2 values close to 0.9999 and to 0.9988 for the training and testing sets, respectively. The results present a big potential use of the as-prepared bio-sorbent for the removal of Cu(II), as well as an efficient green technology for ecological and environmental sustainability

Forecast of pain degree of lumbar disc herniation based on back propagation neural network

To further explore the pathogenic mechanism of lumbar disc herniation (LDH) pain, this study screens important imaging features that are significantly correlated with the pain score of LDH. The features with significant correlation imaging were included into a back propagation (BP) neural network model for training, including Pfirrmann classification, Michigan State University (MSU) regional localization (MSU protrusion size classification and MSU protrusion location classification), sagittal diameter index, sagittal diameter/transverse diameter index, transverse diameter index, and AN angle (angle between nerve root and protrusion). The BP neural network training model results showed that the specificity was 95 ± 2%, sensitivity was 91 ± 2%, and accuracy was 91 ± 2% of the model. The results show that the degree of intraspinal occupation of the intervertebral disc herniation and the degree of intervertebral disc degeneration are related to LDH pain. The innovation of this study is that the BP neural network model constructed in this study shows good performance in the accuracy experiment and receiver operating characteristic experiment, which completes the prediction task of lumbar Magnetic Resonance Imaging features for the pain degree of LDH for the first time, and provides a basis for subsequent clinical diagnosis