BBA - Molecular Cell Research (v.1813, #10)

Phosphorylation of dynamin II at serine-764 is associated with cytokinesis by Megan Chircop; Boris Sarcevic; Martin R. Larsen; Chandra S. Malladi; Ngoc Chau; Michael Zavortink; Charlotte M. Smith; Annie Quan; Victor Anggono; Peter G. Hains; Mark E. Graham; Phillip J. Robinson (1689-1699).
Calcineurin is a phosphatase that is activated at the last known stage of mitosis, abscission. Among its many substrates, it dephosphorylates dynamin II during cytokinesis at the midbody of dividing cells. However, dynamin II has several cellular roles including clathrin-mediated endocytosis, centrosome cohesion and cytokinesis. It is not known whether dynamin II phosphorylation plays a role in any of these functions nor have the phosphosites involved in cytokinesis been directly identified. We now report that dynamin II from rat lung is phosphorylated to a low stoichiometry on a single major site, Ser-764, in the proline-rich domain. Phosphorylation on Ser-764 also occurred in asynchronously growing HeLa cells and was greatly increased upon mitotic entry. Tryptic phospho-peptides isolated by TiO2 chromatography revealed only a single phosphosite in mitotic cells. Mitotic phosphorylation was abolished by roscovitine, suggesting the mitotic kinase is cyclin-dependent kinase 1. Cyclin-dependent kinase 1 phosphorylated full length dynamin II and Glutathione-S-Transferase-tagged–dynamin II–proline-rich domain in vitro, and mutation of Ser-764 to alanine reduced proline-rich domain phosphorylation by 80%, supporting that there is only a single major phosphosite. Ser-764 phosphorylation did not affect clathrin-mediated endocytosis or bulk endocytosis using penetratin-based phospho-deficient or phospho-mimetic peptides or following siRNA depletion/rescue experiments. Phospho-dynamin II was enriched at the mitotic centrosome, but this targeting was unaffected by the phospho-deficient or phospho-mimetic peptides. In contrast, the phospho-mimetic peptide displaced endogenous dynamin II, but not calcineurin, from the midbody and induced cytokinesis failure. Therefore, phosphorylation of dynamin II primarily occurs on a single site that regulates cytokinesis downstream of calcineurin, rather than regulating endocytosis or centrosome function.► DynII is phosphorylated at the onset of mitosis by Cdk1 at Ser-764. ► Ser-764 phosphorylation does not play a role in clathrin-mediated and bulk endocytosis. ► DynII mitotic centrosome localisation is not dependent on Ser-764 phosphorylation. ► Ser-764 phosphorylation regulates dynII midbody localisation. ► Phospho-dynII–Ser-764 is associated with completion of cytokinesis.
Keywords: Centrosome; Cytokinesis; Dynamin II; Mitosis; Phosphorylation;

Molecular and biochemical characterization of nitric oxide synthase isoforms and their intracellular distribution in human peripheral blood mononuclear cells by Rohit Saluja; Anupam Jyoti; Madhumita Chatterjee; Saman Habib; Anupam Verma; Kalyan Mitra; Manoj Kumar Barthwal; Virendra K. Bajpai; Madhu Dikshit (1700-1707).
Nitric oxide synthase (NOS) expression and catalytic status in human peripheral blood mononuclear cells (PBMCs) is debatable, while its sub-cellular distribution remains unascertained. The present study characterizes NOS transcripts by real time PCR, NOS protein by immunoprecipitation (IP)/Western blot (WB), nitric oxide (NO) generation by DAF-2DA and NOS sub-cellular distribution by immunogold electron microscopy in resting PBMCs, monocytes and lymphocytes obtained from healthy donors. We observed constitutive expression of full length NOS isoforms (nNOS, iNOS and eNOS) in PBMCs: with the highest expression of iNOS in comparison to nNOS and eNOS. Isolated monocytes expressed more eNOS transcript and protein as compared to nNOS and iNOS. Lymphocytes however had more iNOS transcripts and protein than nNOS and eNOS. NOS was catalytically active in PBMCs, monocytes as well as in lymphocytes as evident by NO generation in the presence of substrate and cofactors, which was significantly reduced in the presence of NOS inhibitor. Immunogold electron microscopy and morphometric analysis revealed the distinct pattern of NOS distribution in monocytes and lymphocytes and also exhibited differences in the nuclear–cytoplasmic ratio. nNOS localization was much more in the cytosol than in the nucleus among both monocytes and lymphocytes. Interestingly, iNOS distribution was comparable in both cytosol and nucleus among monocytes, but in lymphocytes iNOS was predominantly localized to the cytosol. The present study exhibits constitutive presence of all the NOS isoforms in PBMCs and reports the distinct pattern of NOS distribution among monocytes and lymphocytes.► PBMCs, monocytes and lymphocytes express all the three NOS isoforms (nNOS, iNOS, eNOS) at protein as well as transcript levels. ► The expression of iNOS and eNOS was predominant over other NOS isoforms in lymphocytes and monocytes respectively. ► NOS is catalytically active in PBMCs, monocytes and lymphocytes ► NOS catalysis is attributed to both calcium dependent and independent isoforms. ► Monocytes demonstrated more cytoplasmic predominance for NOS than nuclear distribution while lymphocytes exhibited almost equal distribution in both cellular compartments.
Keywords: Nitric oxide synthase; PBMC; Monocyte; Lymphocyte; Intracellular localization; Immunogold electron microscopy;

Parcs/Gpn3 is required for the nuclear accumulation of RNA polymerase II by Mónica R. Calera; Cristina Zamora-Ramos; Minerva G. Araiza-Villanueva; Carlos A. Moreno-Aguilar; Sonia G. Peña-Gómez; Fabiola Castellanos-Terán; Angélica Y. Robledo-Rivera; Roberto Sánchez-Olea (1708-1716).
Parcs/Gpn3 is a putative GTPase that is conserved in eukaryotic cells from yeast to humans, suggesting that it plays a fundamental, but still unknown, cellular function. Suppression of Parcs/Gpn3 expression by RNAi completely blocked cell proliferation in MCF-12A cells and other mammary epithelial cell lines. Unexpectedly, Parcs/Gpn3 knockdown had a more modest effect in the proliferation of the tumorigenic MDA-MB-231 and SK-BR3 cells. RNA polymerase II (RNAP II) co-immunoprecipitated with Parcs/Gpn3. Parcs/Gpn3 depletion caused a reduction in overall RNA synthesis in MCF-12A cells but not in MDA-MB-231 cells, demonstrating a role for Parcs/Gpn3 in transcription, and pointing to a defect in RNA synthesis by RNAP II as the possible cause of halted proliferation. The absence of Parcs/Gpn3 in MCF-12A cells caused a dramatic change in the sub-cellular localization of Rpb1, the largest subunit of RNAP II. As expected, Rpb1 was present only in the nucleus of MCF-12A control cells, whereas in Parcs/Gpn3-depleted MCF-12A cells, Rpb1 was detected exclusively in the cytoplasm. This effect was specific, as histones remained nuclear independently of Parcs/Gpn3. Rpb1 protein levels were markedly increased in Parcs/Gpn3-depleted MCF-12A cells. Interestingly, Rpb1 distribution was only marginally affected after knocking-down Parcs/Gpn3 in MDA-MB-231 cells. In conclusion, we report here, for the first time, that Parcs/Gpn3 plays a critical role in the nuclear accumulation of RNAP II, and we propose that this function explains the relative importance of Parcs/Gpn3 in cell proliferation. Intriguingly, at least some tumorigenic mammary cells have evolved mechanisms that allow them to proliferate in a Parcs/Gpn3-independent manner.► Silencing of Parcs/Gpn3 completely inhibited proliferation in MCF-12A but affected less in MDA-MB-231 cells. ► RNA polymerase II co-immunoprecipitated with Parcs/Gpn3. ► Parcs/Gpn3 depletion inhibited overall transcription in MCF-12A but not in MDA-MB-231 cells. ► Parcs/Gpn3 depletion caused a cytoplasmic localization of Rpb1 in MCF-12A but not in MDA-MB-231 cells. ► Parcs/Gpn3 depletion significantly increased Rpb1 protein levels in MCF-12A cells.
Keywords: Parcs/Gpn3; nuclear accumulation of RNA polymerase II; cell proliferation; interference RNA;

Estrogen-induced upregulation of Sftpb requires transcriptional control of neuregulin receptor ErbB4 in mouse lung type II epithelial cells by Katja Zscheppang; Mirja Konrad; Melanie Zischka; Verena Huhn; Christiane E.L. Dammann (1717-1727).
Estrogen is known for its positive stimulatory effects on surfactant proteins. ErbB4 receptor and its ligand neuregulin (NRG) positively stimulate lung development. ErbB receptors interact with nuclear receptors and ErbB4 co-regulates estrogen receptor (ER)α expression in breast cells. ERβ is highly expressed in pneumocytes and its deletion leads to fewer alveoli and reduced elastic recoil. A similar picture was seen in ErbB4-deleted lungs. We hypothesized that estrogen signals its effect on surfactant protein B (Sftpb) expression through interactions of ERβ and ErbB4. Estrogen and NRG treatment decreased cell numbers and stimulated Sftpb expression in type II cells. Estrogen and NRG both stimulated phosphorylation of ERβ and co-localization of both receptors. Overexpression of ERβ increased the cell number and Sftpb expression, which was further augmented by estrogen and NRG. Finally, estrogen and NRG stimulated ERβ and ErbB4 binding to the Sftpb promoter. Overexpression of these receptors stimulated Sftpb promoter activation, which was further enhanced by estrogen and NRG. The stimulatory effect of estrogen and NRG was abolished in ErbB4 deletion and reconstituted by re-expression of full-length ErbB4 in fetal ErbB4-deleted type II cells. Estrogen-induced nuclear translocation of ErbB4 required the intact γ-secretase cleavage site but not the nuclear localization sequence of the ErbB4 receptor, suggesting that ERβ might function as a nuclear chaperone for ErbB4. These studies demonstrate that estrogen effects on Sftpb expression require an interaction of ERβ and ErbB4. We speculate that the stimulatory effects of estrogen on Sftpb are under transcriptional control of ErbB4.► We demonstrated that estrogen and NRG treatment stimulated Sftpb expression. ► The expression of ERβ and ErbB4 stimulated Sftpb promoter activity. ► Estrogen induced nuclear translocation of ErbB4. ► Estrogen effects on Sftpb expression require interaction of ERβ and ErbB4. ► Stimulatory effects on Sftpb might be under transcriptional control of ErbB4.
Keywords: Estrogen receptor β; Lung; Tyrosine phosphorylation; Sftpb promoter;

DYRK1A is encoded in the Down's syndrome critical region on human chromosome 21, and plays an important role in the functional and developmental regulation of many types of cells, including neuronal cells. Here we have identified WDR68, an evolutionarily conserved protein with WD40-repeat domains, as a cellular binding partner of DYRK1A. WDR68 was originally identified in petunia as AN11 that controls the pigmentation of flowers by stimulating the transcription of anthocyanin biosynthetic genes. Experiments with RNA interference showed that WDR68 was indispensable for the optimal proliferation and survival of mammalian cultured cell, and WDR68 depletion induced cell apoptosis. DYRK1A and DYRK1B, but not DYRK2, DYRK3, or DYRK4, bound to endogenous and expressed WDR68. The N-terminal domain, but not the catalytic kinase domain or the C-terminal domain of DYRK1A, was responsible for the WDR68 binding. Deletions in the N-terminal or C-terminal region outside of the central WD40-repeats of WDR68 abolished its binding to DYRK1A, suggesting that WD40 repeats are not sufficient for the association with DYRK1A. Immunofluorescent staining revealed that WDR68 was distributed throughout the cell. Importantly, nuclear accumulation of WDR68 was observed upon co-expression of the wild type and a kinase-dead mutant of DYRK1A. Taken together, these results suggest that DYRK1A binds specifically to WDR68 in cells, and that the binding, but not the phosphorylation event, induces the nuclear translocation of WDR68.Display Omitted► We identified WDR68 as a cellular binding partner of DYRK1A. ► WDR68 was indispensable for the proliferation and survival of mammalian cells. ► DYRK1A and DYRK1B, but not DYRK2, DYRK3, or DYRK4, bound to WDR68. ► The N-terminal domain of DYRK1A was responsible for the WDR68 binding. ► DYRK1A overexpression (as in Down’s syndrome) induced nuclear translocation of WDR68.
Keywords: DYRK1A; WDR68; WD40-repeat; Down's syndrome; Nuclear translocation;

Mitochondria adjust Ca2+ signaling regime to a pattern of stimulation in salivary acinar cells by Olga Kopach; Ilya Kruglikov; Tatyana Pivneva; Nana Voitenko; Alexei Verkhratsky; Nataliya Fedirko (1740-1748).
The salivary acinar cells have unique Ca2+ signaling machinery that ensures an extensive secretion. The agonist-induced secretion is governed by Ca2+ signals originated from the endoplasmic reticulum (ER) followed by a store-operated Ca2+ entry (SOCE). During tasting and chewing food a frequency of parasympathetic stimulation increases up to ten fold, entailing cells to adapt its Ca2+ machinery to promote ER refilling and ensure sustained SOCE by yet unknown mechanism. By employing a combination of fluorescent Ca2+ imaging in the cytoplasm and inside cellular organelles (ER and mitochondria) we described the role of mitochondria in adjustment of Ca2+ signaling regime and ER refilling according to a pattern of agonist stimulation. Under the sustained stimulation, SOCE is increased proportionally to the degree of ER depletion. Cell adapts its Ca2+ handling system directing more Ca2+ into mitochondria via microdomains of high [Ca2+] providing positive feedback on SOCE while intra-mitochondrial tunneling provides adequate ER refilling. In the absence of an agonist, the bulk of ER refilling occurs through Ca2+-ATPase-mediated Ca2+ uptake within subplasmalemmal space. In conclusion, mitochondria play a key role in the maintenance of sustained SOCE and adequate ER refilling by regulating Ca2+ fluxes within the cell that may represent an intrinsic adaptation mechanism to ensure a long-lasting secretion.Display Omitted► Extensive secretion requires an adaption of acinar cell Ca2+-signaling machinery. ► We model two types of cell stimulation: short and prolonged presence of an agonist. ► Upon short stimulation mitochondria slightly regulate Ca2+ entry and ER refilling. ► Upon prolonged stimulation mitochondria ensure sustained Ca2+ entry and ER refilling. ► Mitochondria take up and release Ca2+ within Ca2+ microdomains.
Keywords: Store-operated calcium entry; Mitochondria; Endoplasmic reticulum; Ca2+ microdomains; Submandibular acinar cells;

Parathyroid hormone and the regulation of cell cycle in colon adenocarcinoma cells by Natalia Calvo; Claudia Gentili; Ana Russo de Boland (1749-1757).
Parathyroid hormone (PTH) functions as a major mediator of bone remodeling and as an essential regulator of calcium homeostasis. In this study, we investigated the role of PTH in the regulation of the cell cycle in human colon adenocarcinoma Caco-2 cells. Flow cytometry analysis revealed that PTH (10− 8  M, 12–24 h) treatment increases the number of cells in the G0/G1 phase and diminishes the number in both phases S and G2/M. In addition, analysis by Western blot showed that the hormone increases the expression of the inhibitory protein p27Kip1 and diminishes the expression of cyclin D1, cyclin D3 and CDK6. However, the amounts of CDK4, p21Cip1, p15INK4B and p16INK4A were not different in the absence or presence of PTH. Inhibitors of PKC (Ro-318220, bisindolylmaleimide and chelerythine), but not JNK (SP600125) and PP2A (okadaic acid and calyculin A), reversed PTH response in Caco-2 cells. Taken together, our results suggest that PTH induces G0/G1 phase arrest of Caco-2 intestinal cells and changes the expression of proteins involved in cell cycle regulation via the PKC signaling pathway.► PTH induces G0/G1 phase arrest of Caco-2 intestinal cells. ► The hormone changes the expression of proteins involved in cell cycle regulation. ► The effects are dependent of the PKC signalling pathway.
Keywords: PTH; Caco-2 cells; Cell cycle; PKC;

The retinal pigment epithelium (RPE) plays an essential role in the survival and function of the neural retina. RPE uncontrolled proliferation leads to the development of proliferative ocular pathologies, among which proliferative vitreoretinopathy (PVR) is the main cause of retinal surgery failure. Upon the breakdown of the BRB due to trauma or metabolic imbalance the contact of RPE with serum-contained thrombin has been shown to stimulate the proliferation of otherwise quiescent RPE cells. Although the molecular mechanisms involved in this effect are still undetermined, thrombin proteolytic activation of protease-activated G protein coupled receptor-1 (PAR-1) activates PI3K and Akt, known to play an essential role in proliferation. The present study demonstrates that: 1) thrombin stimulates Ser 473 Akt phosphorylation without affecting Thr 308 basal phosphorylation in RPE cells; 2) thrombin-induced Akt stimulation promotes cyclin D1 accumulation through the phosphorylation/ inhibition of GSK-3β, thus preventing Thr 286 cyclin D1 phosphorylation, nuclear export and degradation; 3) Akt signaling requires the upstream activation of PI3K and PLC. Since the pharmacological inhibition of these pathways or the silencing of cyclin expression prevent thrombin-induced RPE cell proliferation, these results contribute relevant evidence for establishing the mechanism involved in the development of proliferative eye diseases.► Thrombin induces Akt activation through Ser 473 phosphorylation in RPE cells. ► Akt signaling requires PI3K and PLC activation in thrombin-stimulated RPE cells. ► Thrombin promotes cyclin D1 accumulation through Akt-dependent GSK-3β inhibition. ► Akt activation is required for thrombin-induced RPE cell proliferation.
Keywords: Akt; Thrombin; Cyclin D1; Proliferation;

Comparative role of acetylation along c-SRC/ETS1 signaling pathway in bone metastatic and invasive mammary cell phenotypes by Paola Bendinelli; Paola Maroni; Emanuela Matteucci; Maria Alfonsina Desiderio (1767-1776).
Metastatic cells switch between different modes of migration through supramolecular plasticity mechanism(s) still largely unknown. The aim of the present paper was to clarify some molecular aspects of the epigenetic control of migration of 1833-bone metastatic cells compared to MDA-MB231-parental mammary carcinoma cells. Active c-Src overexpression enhanced 1833-cell spontaneous migration and CXCR4-mediated chemoinvasion toward CXCL12 ligand. Only in metastatic cells, in fact, c-Src seemed to stabilize nuclear CXCR4-protein receptor possibly due to tyrosine phosphorylation, by impairing protein-degradative smear and causing instead an electrophoretic-mobility shift; the cytosolic steady-state level of CXCR4 was enhanced, and the protein appeared also phosphorylated. These findings suggested the triggering of unique signaling pathways in metastasis for homing of breast-cancer cells to congenial environment of specific organs. Microenvironmental stimuli activating c-Src might influence Ets1 binding to CXCR4 promoter and consequent transactivation, as well as CXCR4 post-translational regulatory mechanisms such as phosphorylation. Enhancement of Ets1 activity and CXCR4 induction by c-Src overexpression were prevented by histone deacetylase (HDAC) blockade. In contrast, HDAC inhibition with trichostatin A increased cytosolic phosphorylated CXCR4 expression in MDA-MB231 cells, but Ets1 involvement was practically unneeded. c-Src might be suggested as a bio-marker predicting metastasis sensitivity patterns to HDAC inhibitors. Rationally designed and individualized therapy may become possible as more is learned about the target molecules of HDAC's inhibitory agents and their roles, as undertaken for CXCR4 that is likely to be crucial for homing, angiogenesis and survival in a c-Src-dependent manner in bone-metastatic mammary cells.► c-Src-dependent activation of Ets1 was regulated by HDACs in bone-metastatic 1883 cells. ► c-Src/Ets1 pathway transactivated CXCR4 and caused nuclear-tyrosine phosphorylation only in 1833 clone. ► CXCR4-molecular changes due to c-Src had a functional role for metastasis chemoinvasion. ► c-Src expression, as occurs in response to microenvironmental stimuli, might influence migratory dissemination and gene expression profile. ► c-Src may be a predictive biomarker to define metastasis sensitivity patterns to HDAC inhibitors.
Keywords: c-Src; HDACs; CXCR4; Breast cancer; Bone metastasis;

Targeted disruption of Mcm10 causes defective embryonic cell proliferation and early embryo lethality by Han Jeong Lim; Yoon Jeon; Chang Hwan Jeon; Jong Hyun Kim; Ho Lee (1777-1783).
Minichromosome maintenance 10 (MCM10) is a conserved, abundant nuclear protein, which plays a key role in the initiation of eukaryotic chromosomal DNA replication and elongation. To elucidate the physiological importance of MCM10 in vivo, we generated conventional knockout mice. No MCM10-null embryos were recovered after E8.5, and the mutation was found to be lethal before the implantation stage. Mutant embryos showed apparently normal growth until the morula stage, but growth defects after this stage. The dramatic reduction of 5-bromo-2-deoxyuridine (BrdU) incorporation in the mutant embryo, followed by cell death, suggests that defective cell proliferation may underlie this developmental failure. Taken together, these findings provide the first unequivocal genetic evidence for an essential and non-redundant physiological role of MCM10 during murine peri-implantation development.► MCM10-deficient mouse embryos die at the peri-implantation stage and none can be recovered after E8.5. ► MCM10-null mouse embryos show defective cell proliferation, followed by cell death. ► Mouse Mcm10 is highly expressed during early embryogenesis and in tissue enriched with proliferative cells.
Keywords: MCM10; Peri-implantation development; Embryo lethality; Mouse;

TRIM8 regulates Nanog via Hsp90β-mediated nuclear translocation of STAT3 in embryonic stem cells by Fumihiko Okumura; Akiko J. Okumura; Masaki Matsumoto; Keiichi I. Nakayama; Shigetsugu Hatakeyama (1784-1792).
TRIM8 is a member of a protein family defined by the presence of a common domain structure composed of a tripartite motif including a RING-finger, one or two B-box domains and a coiled-coil motif. Here, we show that TRIM8 interacts with Hsp90β, which interacts with STAT3 and selectively downregulates transcription of Nanog in embryonic stem cells. Knock-down of TRIM8 increased phosphorylated STAT3 in the nucleus and also enhanced transcription of Nanog. These findings suggest that TRIM8 modulates translocation of phosphorylated STAT3 into the nucleus through interaction with Hsp90β and consequently regulates transcription of Nanog in embryonic stem cells.► TRIM8 interacted with Hsp90β, which selectively downregulates transcription of Nanog. ► Knock-down of TRIM8 increased transcription of Nanog. ► TRIM8 may modulate translocation of STAT3 into the nucleus through Hsp90β.
Keywords: TRIM8; Hsp90β; STAT3; Nanog; Ubiquitin;

Protein kinase C α regulates nuclear pri-microRNA 15a release as part of endothelin signaling by Melanie von Brandenstein; Reinhard Depping; Ekaterine Schäfer; Hans-Peter Dienes; Jochen W.U. Fries (1793-1802).
Endothelin-1 induced signaling is characterized by an early induction of a nuclear factor-kappa B p65/mitogen-activated phosphokinase p38 transcription complex via its A-receptor versus a late induction via diacylglycerol, and protein kinase C. A possible interaction between these two pathways and a potential function for protein kinase C in this context has not previously been elucidated. Here we report that in Caki-1 tumor cells, protein kinase C α is a part of the transcription complex. With importin α4 and α5 as chaperones, the transcription complex transmigrates into the nucleus. Protein kinase C α blocks the nuclear release of pri-microRNA 15a by direct binding shown by electrophoretic mobility shift assay and Duolink immune histology. The expression levels of miRNA 15a can be further manipulated by transfection of si-protein kinase C α, or an expression vector containing protein kinase C α or miRNA 15. The miRNA 15a regulation by protein kinase C α is detectable in different malignant human tumor cell lines (renal cell carcinoma, breast carcinoma, and melanoma). Furthermore, all three cell lines harbor both endothelin receptors (ETAR/ETBR). Specific blockage of each receptor leads to major reduction of miRNA 15a expression due to increased nuclear protein kinase C α translocation. We conclude that the nuclear binding of pri-microRNA 15a is a novel function of protein kinase C α, which plays an important role in endothelin-1 mediated signaling. Since several endothelin-sensitive, malignant tumor cell lines harbor this regulation, it could indicate a more general role in tumor biology.► PKC α translocates into the nucleus as part of the NF-κB transcription complex. ► PKC α binds to pri-miRNA 15a, prevents its nuclear release and maturation to miRNA. ► Levels of PKC α and miRNA 15a are inversely related in different human cell lines. ► This signalling occurs via endothelin-1 binding to its surface receptors. ► miRNA 15a expression is dependent on expression of both endothelin receptors.
Keywords: Endothelin-1; MAPK p38; NF-κB; PKC α; miRNA 15a; Tumor cell;

C/EBP ζ targets to neutrophil gelatinase-associated lipocalin (NGAL) as a repressor for metastasis of MDA-MB-231 cells by Lihong Wang; Huawen Li; Jian Wang; Wei Gao; Yani Lin; Weina Jin; Guoqiang Chang; Ruojun Wang; Qinghua Li; Li Ma; Tianxiang Pang (1803-1813).
Breast cancer is a leading cause of morbidity in women worldwide. neutrophil gelatinase-associated lipocalin (NGAL), a useful biomarker of ER negative (ER) breast cancer, promotes local tumor invasion and lymph node metastasis. We first identified the distinctive expression of NGAL in two breast cancer cell lines MCF7 and MDA-MB-231 cells, and then confirmed NGAL as a critical inducer of metastasis. Finally, the transcriptional factor CCAAT enhancer-binding proteins ζ (C/EBP ζ) was overexpressed in MDA-MB-231 cells. Consistent with the effect of NGAL knockdown, C/EBP ζ overexpression caused the significant changes that could prevent cell metastasis. C/EBP ζ overexpression induced a strong decrease in NGAL and matrix metalloproteinases (MMPs) expressions as determined by quantitative real time PCR and Western blotting. To identify the potential role of C/EBP ζ on regulating of NGAL in breast cancer, we established the dual-luciferase reporter assay for NGAL in MDA-MB-231 cells cotransfected with C/EBP ζ. Promoter reporter assays determined that C/EBP ζ directly repressed the human NGAL gene promoter activity by inhibiting the NGAL transcription. Taken together, this work identified that the C/EBP ζ overexpression downregulated NGAL to inhibit migration and invasion of breast cancer, which could be used as a novel strategy for breast cancer therapy.► NGAL knockdown evidently decreases MMPs expression and cell metastasis. ► C/EBP ζ overexpression significantly prevents cell metastasis. ► C/EBP ζ overexpression induces a strong decrease in NGAL and MMPs expression. ► C/EBP ζ directly represses the human NGAL gene promoter activity. ► NHE1 inhibition reduces cell metastasis via C/EBP ζ-mediated NGAL downregulation.
Keywords: Neutrophil gelatinase-associated lipocalin; Breast cancer; Metastasis; C/EBP ζ;

Glucocorticoids induce mitochondrial gene transcription in HepG2 cells by Anna-Maria G. Psarra; Constantine E. Sekeris (1814-1821).
Glucocorticoids are major regulators of a plethora of cellular functions, acting on target cells through glucocorticoid receptors (GR) and modulation of gene transcription, among other mechanisms. One main site of action of glucocorticoids is the hepatocyte, which responds to the hormonal stimulus with induction of several proteins among them enzymes of oxidative phosphorylation (OXPHOS), both nuclearly and mitochondrially encoded. The induction of OXPHOS is regarded as a result of a nuclear action of the receptor on the respective nuclear genes and on genes encoding mitochondrial transcription factors. The presence of GR in mitochondria and of sequences in the mitochondrial genome similar to glucocorticoid responsive elements, suggested a direct action of GR on mitochondrial transcription. We demonstrate in HepG2 hepatocarcinoma cells specific binding of GR to the regulatory D-loop region of the mitochondrial genome and show that dexamethasone induces the mitochondrial transcription factors A, B1, and B2, the mitochondrial ribosomal RNA, and several mitochondrially encoded OXPHOS genes. Applying α-amanitin, the specific inhibitor of DNA-dependent RNA polymerase II, the dexamethasone-induced transcription of the mitochondrial genes can still proceeds, whereas the DEX effect on transcription of the mitochondrial transcription factors is suppressed. Moreover, HepG2 cells overexpressing mitochondrial targeted GR showed increased RNA synthesis, cytrochrome oxidase subunit I protein expression, and mitochondrial ATP production. We conclude that glucocorticoids can stimulate directly mitochondrial transcription by the mitochondrially localized GR, affecting OXPHOS enzyme biosynthesis. This takes place in addition to their action on mitochondrial genes by way of induction of the nuclearly encoded mitochondrial transcription factors.► Glucocorticoid receptor binds to the D-loop of the mitochondrial DNA of HepG2 cells. ► DEX induces OXPHOS gene expression by the mitochondrial glucocorticoid receptor. ► DEX induces mitochondrial transcription factors gene expression. ► DEX-induced mitochondrial transcription maintained in the presence of α-amanitin. ► Mitochondrial GR activates OXPHOS enzyme synthesis and mitochondrial ATP production.
Keywords: Glucocorticoid receptor; Mitochondrial transcription; OXPHOS; Mitochondrial transcription factor; Mitochondrial DNA; Mitochondrial ATP production;

Lactoferrin inhibits neutrophil apoptosis via blockade of proximal apoptotic signaling events by Nigel Francis; See Heng Wong; Peter Hampson; Keqing Wang; Stephen P. Young; Hans Peter Deigner; Michael Salmon; Dagmar Scheel-Toellner; Janet M. Lord (1822-1826).
Neutrophils are the most abundant leukocyte and have a short lifespan, dying by apoptosis approximately five days after leaving the bone marrow. Their apoptosis can be delayed at sites of inflammation to extend their functional lifespan, but inappropriate inhibition of apoptosis contributes to chronic inflammatory disease. Levels of the physiological iron chelator lactoferrin are raised at sites of inflammation and we have shown previously that iron-unsaturated lactoferrin inhibited human neutrophil apoptosis, but the mechanisms involved were not determined. Here we report that the anti-apoptotic effect of lactoferrin is dependent upon its iron saturation status as iron-saturated lactoferrin did not affect neutrophil apoptosis. We also show that the effect of lactoferrin is mediated at an early stage in apoptosis as it inhibited activation of sphingomyelinase, generation of ceramide, activation of caspase 8 and Bax and cleavage of Bid. Lactoferrin did not inhibit apoptosis induced by exogenous ceramide, supporting the proposal that it acts upstream of ceramide generation. We therefore conclude that raised lactoferrin levels are likely to contribute to chronic inflammation by delaying neutrophil apoptosis and that this is achieved by inhibiting proximal apoptotic signaling events.► The anti-apoptotic effect of lactoferrin on neutrophils is dependent on its iron saturation status. ► The anti-apoptotic effect of lactoferrin is mediated at an early stage in neutrophil apoptosis. ► Lactoferrin inhibits generation of ceramide and activation of caspase 8 and Bax, as well as Bid cleavage and loss of Mcl-1.
Keywords: Neutrophil; apoptosis; iron chelator; inflammation; apoptotic signaling;

Pro-inflammatory cytokine-mediated beta cell apoptosis is activated through multiple signaling pathways involving mitochondria and endoplasmic reticulum. Activation of organelle-specific caspases has been implicated in the progression and execution of cell death. This study was therefore performed to elucidate the effects of pro-inflammatory cytokines on a possible cross-talk between the compartment-specific caspases 9 and 12 and their differential contribution to beta cell apoptosis. Moreover, the occurrence of ROS-mediated mitochondrial damage in response to beta cell toxic cytokines has been quantified. ER-specific caspase-12 was strongly activated in response to pro-inflammatory cytokines; however, its inhibition did not abolish cytokine-induced mitochondrial caspase-9 activation and loss of cell viability. In addition, there was a significant induction of oxidative mitochondrial DNA damage and elevated cardiolipin peroxidation in insulin-producing RINm5F cells and rat islet cells. Overexpression of the H2O2 detoxifying enzyme catalase effectively reduced the observed cytokine-induced oxidative damage of mitochondrial structures. Taken together, the results strongly indicate that mitochondrial caspase-9 is not a downstream substrate of ER-specific caspase-12 and that pro-inflammatory cytokines cause apoptotic beta cell death through activation of caspase-9 primarily by hydroxyl radical-mediated mitochondrial damage.► Caspase-9 is not a target of the ER-specific caspase-12 in insulin-producing cells. ► Activation of caspase-12 is not involved in cytokine-induced beta cell apoptosis. ► Cytokines induce beta cell death by ROS-mediated mitochondrial damage. ► Mitochondrially located catalase prevents ROS-induced damage of mitochondrial structures.
Keywords: Oxidative damage; Mitochondrial apoptosis pathway; Beta cell death; Caspase-9; ER-specific caspase-12; Cytokine;

HIV-1 Tat binds to SH3 domains: Cellular and viral outcome of Tat/Grb2 interaction by Slava Rom; Marco Pacifici; Giovanni Passiatore; Susanna Aprea; Agnieszka Waligorska; Luis Del Valle; Francesca Peruzzi (1836-1844).
The Src-homology 3 (SH3) domain is one of the most frequent protein recognition modules (PRMs), being represented in signal transduction pathways and in several pathologies such as cancer and AIDS. Grb2 (growth factor receptor-bound protein 2) is an adaptor protein that contains two SH3 domains and is involved in receptor tyrosine kinase (RTK) signal transduction pathways. The HIV-1 transactivator factor Tat is required for viral replication and it has been shown to bind directly or indirectly to several host proteins, deregulating their functions. In this study, we show interaction between the cellular factor Grb2 and the HIV-1 trans-activating protein Tat. The binding is mediated by the proline-rich sequence of Tat and the SH3 domain of Grb2. As the adaptor protein Grb2 participates in a wide variety of signaling pathways, we characterized at least one of the possible downstream effects of the Tat/Grb2 interaction on the well-known IGF-1R/Raf/MAPK cascade. We show that the binding of Tat to Grb2 impairs activation of the Raf/MAPK pathway, while potentiating the PKA/Raf inhibitory pathway. The Tat/Grb2 interaction affects also viral function by inhibiting the Tat-mediated transactivation of HIV-1 LTR and viral replication in infected primary microglia.► The SH3 domain of Grb2 interacts with HIV-1 Tat protein. ► Tat/Grb2 interaction affects cellular signaling pathways and viral functions. ► It impairs IGF-1R/ERK activation and enhances PKA/Raf inhibitory cascade in LN229. ► It inhibits viral replication in HIV-infected human primary microglia.
Keywords: HIV-1 Tat; Grb2; Signal transduction; Viral replication; HIVE; SH3 domain;

Tau inhibits tubulin oligomerization induced by prion protein by Katarzyna M. Osiecka; Hanna Nieznanska; Krzysztof J. Skowronek; Jolanta Jozwiak; Krzysztof Nieznanski (1845-1853).
In previous studies we have demonstrated that prion protein (PrP) interacts with tubulin and disrupts microtubular cytoskeleton by inducing tubulin oligomerization. These observations may explain the molecular mechanism of toxicity of cytoplasmic PrP in transmissible spongiform encephalopathies (TSEs). Here, we check whether microtubule associated proteins (MAPs) that regulate microtubule stability, influence the PrP-induced oligomerization of tubulin. We show that tubulin preparations depleted of MAPs are more prone to oligomerization by PrP than those containing traces of MAPs. Tau protein, a major neuronal member of the MAPs family, reduces the effect of PrP. Importantly, phosphorylation of Tau abolishes its ability to affect the PrP-induced oligomerization of tubulin. We propose that the binding of Tau stabilizes tubulin in a conformation less susceptible to oligomerization by PrP. Since elevated phosphorylation of Tau leading to a loss of its function is observed in Alzheimer disease and related tauopathies, our results point at a possible molecular link between these neurodegenerative disorders and TSEs.► Tau inhibits PrP-induced oligomerization of tubulin. ► Tau does not compete with PrP for binding site on tubulin. ► Tau stabilizes tubulin in a form less susceptible to PrP-induced oligomerization. ► Phosphorylation regulates Tau's ability to inhibit PrP-induced oligomerization. ► Tau phosphorylation may indirectly enhance neurotoxicity of cytosolic PrP.
Keywords: Prion protein; Tubulin; Tau; Phosphorylation; Prion disease; Alzheimer disease;

MEK5/ERK5/mef2: A novel signaling pathway affected by hepatitis C virus non-enveloped capsid-like particles by Konstantina Katsarou; Panagiota Tsitoura; Urania Georgopoulou (1854-1862).
Hepatitis C virus (HCV) is an RNA positive strand virus, member of the Flaviviridae family. The viral particle is composed of a capsid containing the genome, surrounded by E1 and E2 proteins, however different forms of viral particles have been observed including non-enveloped particles. Previous reports have proposed that hepatitis C non-enveloped capsid-like particles (HCVne) enter cells of hepatic origin via clathrin-mediated endocytosis, during which different signaling events occur. In this report we show that HCVne particles are capable of inducing the recently discovered ERK5 pathway, in a dose dependent way. The ERK5 pathway can be activated by growth factors and other extracellular signals. This specific activation occurs through a well characterized upstream kinase, MEK5, and is capable of inducing gene regulation of mef2. In contrast, when HCV core structural and NS5A non-structural proteins were expressed endogenously no activation of this pathway was detected. These cell signaling events could be of critical importance and might give clues for the elucidation of cellular manifestations associated with HCV infection.► HCVne particles as modulators of MAPK cellular signaling. ► HCVne particles can activate the MEK5–ERK5–mef2. ► Incubation with HCVne produced a perinuclear and in a few cases a nuclear distribution of ERK5. ► MEK5/ERK5/mef2 modulation is HCVne endocytosis-dependent.
Keywords: Hepatitis C virus; Non-enveloped capsid-like particle (HCVne); ERK5; mef2; Cell signaling;

Neurotensin downregulates the pro-inflammatory properties of skin dendritic cells and increases epidermal growth factor expression by Lucília da Silva; Bruno Miguel Neves; Liane Moura; Maria Teresa Cruz; Eugénia Carvalho (1863-1871).
In the last decades some reports reveal the neuropeptide neurotensin (NT) as an immune mediator in the Central Nervous System and in the gastrointestinal tract, however its effects on skin immunity were not identified. The present study investigates the effect of NT on signal transduction and on pro/anti-inflammatory function of skin dendritic cells. Furthermore, we investigated how neurotensin can modulate the inflammatory responses triggered by LPS in skin dendritic cells. We observed that fetal-skin dendritic cells (FSDCs) constitutively express NTR1 and NTR3 (neurotensin receptors) and that LPS treatment induces neurotensin expression. In addition, NT downregulated the activation of the inflammatory signaling pathways NF-κB and JNK, as well as, the expression of the cytokines IL-6, TNF-α, IL-10 and the vascular endothelial growth factor (VEGF), while the survival pathway ERK and epidermal growth factor (EGF) were upregulated. Simultaneous dendritic cells exposure to LPS and NT induced a similar cytokine profile to that one induced by NT alone. However, cells pre-treated with NT and then incubated with LPS, completely changed their cytokine profile, upregulating the cytokines tested, without changes on growth factor expression. Overall, our results could open new perspectives in the design of new therapies for skin diseases, like diabetic wound healing, where neuropeptide exposure seems to be beneficial.Display Omitted► Skin dendritic cells (DC) constitutively express Neurotensin (NT) receptors 1 and 3. ► NT expression increases under inflammatory conditions in contrast to NTR1 and NTR3. ► NT downregulates LPS-induced signaling pathways NF-kB and JNK, while activating ERK. ► NT downregulates the expression of IL-6, TNF-α, IL-10 and VEGF, while activating EGF. ► NT downregulates the pro-inflammatory properties of DC and increases EGF expression.
Keywords: Skin diseases; Wound healing; Neuropeptides; Neurotensin; Langerhans cells; Inflammation and signaling pathways;

The enhanced host-cell permissiveness of human cytomegalovirus is mediated by the Ras signaling pathway by Harilaos Filippakis; Panagiota Dimitropoulou; Aristides G. Eliopoulos; Demetrios A. Spandidos; George Sourvinos (1872-1882).
Human cytomegalovirus utilizes cellular signal transduction pathways to activate viral or cellular transcription factors involved in the control of viral gene expression and DNA replication. In the present study, we demonstrate that Harvey-ras-transformed cells show increased permissiveness to human cytomegalovirus when compared to their parental non-transformed cells. Both the progeny viral yield and the protein levels were elevated in the human cytomegalovirus-infected Harvey-ras-transformed cells requiring active viral gene replication, as shown by the infection with UV-inactivated human cytomegalovirus. Inhibition of Ras or of key molecules of the Ras pathway, effectively suppressed viral infection in the Harvey-ras-transformed cells. On a cellular level, the human cytomegalovirus-infected Harvey-ras-transformed cells formed larger cellular foci, which were significantly higher in number, compared to the uninfected cells and preferentially recruited human cytomegalovirus virions, thereby incriminating human cytomegalovirus infection for the increased transformation of these cells. Furthermore, proliferation assays revealed a higher rate for the human cytomegalovirus-infected Harvey-ras-transformed cells compared to mock-infected cells, whereas human cytomegalovirus infection had no considerable effect on the proliferation of the non-transformed cells. Higher susceptibility to apoptosis was also detected in the human cytomegalovirus-infected ras-transformed cells, which in combination with the higher progeny virus reveals a mode by which human cytomegalovirus achieves efficient spread of infection in the cells expressing the oncogenic Harvey-ras (12 V) gene. Collectively, our data suggest that human cytomegalovirus employs the host-cell Ras signaling pathway to ensue viral expression and ultimately successful propagation. Transformed cells with an activated Ras signaling pathway are therefore particularly susceptible to human cytomegalovirus infection.► H-ras-transformed cells show increased permissiveness to HCMV compared to their parental non-transformed cells. ► Inhibition of the Ras pathway, effectively suppresses viral infection in the H-ras-transformed cells. ► HCMV promotes cellular foci formation and increased proliferation rate of the H-ras-transformed cells. ► HCMV-infected ras-transformed cells are more susceptible to apoptosis facilitating the efficient viral spread.
Keywords: Human cytomegalovirus; Ras; Transformation; Signal transduction; Cancer;

Direct interaction of the Usher syndrome 1G protein SANS and myomegalin in the retina by Nora Overlack; Dilek Kilic; Katharina Bauß; Tina Märker; Hannie Kremer; Erwin van Wijk; Uwe Wolfrum (1883-1892).
The human Usher syndrome (USH) is the most frequent cause of combined hereditary deaf-blindness. USH is genetically heterogeneous with at least 11 chromosomal loci assigned to 3 clinical types, USH1-3. We have previously demonstrated that all USH1 and 2 proteins in the eye and the inner ear are organized into protein networks by scaffold proteins. This has contributed essentially to our current understanding of the function of USH proteins and explains why defects in proteins of different families cause very similar phenotypes. We have previously shown that the USH1G protein SANS (scaffold protein containing ankyrin repeats and SAM domain) contributes to the periciliary protein network in retinal photoreceptor cells. This study aimed to further elucidate the role of SANS by identifying novel interaction partners. In yeast two-hybrid screens of retinal cDNA libraries we identified 30 novel putative interacting proteins binding to the central domain of SANS (CENT). We confirmed the direct binding of the phosphodiesterase 4D interacting protein (PDE4DIP), a Golgi associated protein synonymously named myomegalin, to the CENT domain of SANS by independent assays. Correlative immunohistochemical and electron microscopic analyses showed a co-localization of SANS and myomegalin in mammalian photoreceptor cells in close association with microtubules. Based on the present results we propose a role of the SANS-myomegalin complex in microtubule-dependent inner segment cargo transport towards the ciliary base of photoreceptor cells.► The human Usher syndrome (USH) type 1G protein SANS directly interacts with myomegalin. ► Both scaffold proteins are partially co-localized in the retinas of mouse, non-human primate and human. ► In retinal photoreceptor cells both proteins are associated with microtubule-based intracellular cargo transport. ► Present decipherment of the USH protein network composition provides novel insights into the pathology of the USH disease.
Keywords: Sensorineuronal degeneration; Photoreceptor cell function; Intracellular transport; Microtubule based transport; Phosphodiesterase 4D interacting protein (PDE4DIP);

The endoplasmic reticulum (ER) is the biggest organelle in most cell types, but its characterization as an organelle with a continuous membrane belies the fact that the ER is actually an assembly of several, distinct membrane domains that execute diverse functions. Almost 20 years ago, an essay by Sitia and Meldolesi first listed what was known at the time about domain formation within the ER. In the time that has passed since, additional ER domains have been discovered and characterized. These include the mitochondria-associated membrane (MAM), the ER quality control compartment (ERQC), where ER-associated degradation (ERAD) occurs, and the plasma membrane-associated membrane (PAM). Insight has been gained into the separation of nuclear envelope proteins from the remainder of the ER. Research has also shown that the biogenesis of peroxisomes and lipid droplets occurs on specialized membranes of the ER. Several studies have shown the existence of specific marker proteins found on all these domains and how they are targeted there. Moreover, a first set of cytosolic ER-associated sorting proteins, including phosphofurin acidic cluster sorting protein 2 (PACS-2) and Rab32 have been identified. Intra-ER targeting mechanisms appear to be superimposed onto ER retention mechanisms and rely on transmembrane and cytosolic sequences. The crucial roles of ER domain formation for cell physiology are highlighted with the specific targeting of the tumor metastasis regulator gp78 to ERAD-mediating membranes or of the promyelocytic leukemia protein to the MAM.► This review summarizes mechanisms that lead to the targeting of ER proteins to individual membrane domains (e.g., rough ER). ► Formation of interactions between the Sec61 proteins, ribosomes and mRNAs is critical for rough ER targeting. ► The equilibrium between atlastins, reticulons and DP1/Yop1p determines the formation of smooth ER tubules. ► Targeting to the MAM depends on ER redox and on cytosolic or mitochondrial sorting motifs. ► Targeting to ERES, ERQC, and sites of peroxisome and lipid droplet biogenesis depend on COPI and COPII proteins.
Keywords: Endoplasmic reticulum (ER); Mitochondria-associated membrane (MAM); Plasma membrane-associated membrane (PAM); Russell bodies; Lipid droplet;

The first and third extracellular loops (ECL) of G protein-coupled receptors (GPCRs) have been implicated in ligand binding and receptor function. This study describes the results of an alanine/leucine scan of ECLs 1 and 3 and loop-associated transmembrane (TM) domains of the secretin-like GPCR calcitonin receptor-like receptor which associates with receptor activity modifying protein 1 to form the CGRP receptor. Leu195Ala, Val198Ala and Ala199Leu at the top of TM2 all reduced αCGRP-mediated cAMP production and internalization; Leu195Ala and Ala199Leu also reduced αCGRP binding. These residues form a hydrophobic cluster within an area defined as the “minor groove” of rhodopsin-like GPCRs. Within ECL1, Ala203Leu and Ala206Leu influenced the ability of αCGRP to stimulate adenylate cyclase. In TM3, His219Ala, Leu220Ala and Leu222Ala have influences on αCGRP binding and cAMP production; they are likely to indirectly influence the binding site for αCGRP as well as having an involvement in signal transduction. On the exofacial surfaces of TMs 6 and 7, a number of residues were identified that reduced cell surface receptor expression, most noticeably Leu351Ala and Glu357Ala in TM6. The residues may contribute to the RAMP1 binding interface. Ile360Ala impaired αCGRP-mediated cAMP production. Ile360 is predicted to be located close to ECL2 and may facilitate receptor activation. Identification of several crucial functional loci gives further insight into the activation mechanism of this complex receptor system and may aid rational drug design.► This study provides the first detailed analysis of the function of the 1st and 3rd extracellular loops of the CGRP receptor. ► A hydrophobic cluster of amino acids identified on the 2nd transmembrane helix may be required for CGRP recognition. ► A cluster of amino acids in the middle of the third transmembrane helix of the receptor are needed for receptor activation. ► Residues in the third extracellular loop and associated helices are needed for receptor expression; they may recognise RAMP1.
Keywords: CGRP; Extracellular loop; Receptor activation; Juxtamembrane domain; G protein-coupled receptor; Receptor activity-modifying protein;

PLTP regulates STAT3 and NFκB in differentiated THP1 cells and human monocyte-derived macrophages by S. Vuletic; W. Dong; G. Wolfbauer; C. Tang; J.J. Albers (1917-1924).
Phospholipid transfer protein (PLTP) plays an important role in regulation of inflammation. Previously published studies have shown that PLTP binds, transfers and neutralizes bacterial lipopolysaccharides. In the current study we tested the hypothesis that PLTP can also regulate anti-inflammatory pathways in macrophages. Incubation of macrophage-like differentiated THP1 cells and human monocyte-derived macrophages with wild-type PLTP in the presence or absence of tumor necrosis factor alpha (TNFα) or interferon gamma (IFNγ) significantly increased nuclear levels of active signal transducer and activator of transcription 3, pSTAT3Tyr705 (p < 0.01). Similar results were obtained in the presence of a PLTP mutant without lipid transfer activity (PLTPM159E), suggesting that PLTP-mediated lipid transfer is not required for activation of the STAT3 pathway. Inhibition of ABCA1 by chemical inhibitor, glyburide, as well as ABCA1 RNA inhibition, reversed the observed PLTP-mediated activation of STAT3. In addition, PLTP reduced nuclear levels of active nuclear factor kappa-B (NFκB) p65 and secretion of pro-inflammatory cytokines in conditioned media of differentiated THP1 cells and human monocyte-derived macrophages. Our data suggest that PLTP has anti-inflammatory capabilities in macrophages.Display Omitted► PLTP increases active STAT3 in human macrophages and differentiated THP1 cells. ► PLTP-mediated STAT3 activation does not depend on PLTP phospholipid transfer. ► PLTP-mediated increase in nuclear pSTAT3Tyr705 is inhibited by ABCA1 inhibition. ► PLTP reduces active NFκB p65 in human macrophages and differentiated THP1 cells. ► PLTP reduces inflammatory cytokines in macrophages and differentiated THP1 cells.
Keywords: STAT3 activation; NFκB reduction; Inflammation; Human monocyte-derived macrophage; Differentiated THP1 cell;