Transcriptional Activation by NFκB Increases Perlecan/HSPG2 Expression in the Desmoplastic Prostate Tumor Microenvironment

Perlecan/HSPG2, a heparan sulfate proteoglycan typically found at tissue borders including those separating epithelia and connective tissue, increases near sites of invasion of primary prostatic tumors as previously shown for other proteins involved in desmoplastic tissue reaction. Studies of prostate cancer cells and stromal cells from both prostate and bone, the major site for prostate cancer metastasis, showed that cancer cells and a subset of stromal cells increased production of perlecan in response to cytokines present in the tumor microenvironment. In silico analysis of the HSPG2 promoter revealed two conserved NFκB binding sites, in addition to the previously reported SMAD3 binding sites. By systematically transfecting cells with a variety of reporter constructs including sequences up to 2.6 kb from the start site of transcription, we identified an active cis element in the distal region of the HSPG2 promoter, and showed that it functions in regulating transcription of HSPG2. Treatment with TNF-α and/or TGFβ1 identified TNF-α as a major cytokine regulator of perlecan production. TNF-α treatment also triggered p65 nuclear translocation and binding to the HSPG2 regulatory region in stromal cells and cancer cells. In addition to stromal induction of perlecan production in the prostate, we identified a matrix-secreting bone marrow stromal cell type that may represent the source for increases in perlecan in the metastatic bone marrow environment. These studies implicate perlecan in cytokine-mediated, innate tissue responses to cancer cell invasion, a process we suggest reflects a modified wound healing tissue response co-opted by prostate cancer cells

Cryo-Electron Tomography Elucidates the Molecular Architecture of Treponema pallidum, the Syphilis Spirochete

Cryo-electron tomography (CET) was used to examine the native cellular organization of Treponema pallidum, the syphilis spirochete. T. pallidum cells appeared to form flat waves, did not contain an outer coat and, except for bulges over the basal bodies and widening in the vicinity of flagellar filaments, displayed a uniform periplasmic space. Although the outer membrane (OM) generally was smooth in contour, OM extrusions and blebs frequently were observed, highlighting the structure’s fluidity and lack of attachment to underlying periplasmic constituents. Cytoplasmic filaments converged from their attachment points opposite the basal bodies to form arrays that ran roughly parallel to the flagellar filaments along the inner surface of the cytoplasmic membrane (CM). Motile treponemes stably attached to rabbit epithelial cells predominantly via their tips. CET revealed that T. pallidum cell ends have a complex morphology and assume at least four distinct morphotypes. Images of dividing treponemes and organisms shedding cell envelope-derived blebs provided evidence for the spirochete’s complex membrane biology. In the regions without flagellar filaments, peptidoglycan (PG) was visualized as a thin layer that divided the periplasmic space into zones of higher and lower electron densities adjacent to the CM and OM, respectively. Flagellar filaments were observed overlying the PG layer, while image modeling placed the PG-basal body contact site in the vicinity of the stator–P-collar junction. Bioinformatics and homology modeling indicated that the MotB proteins of T. pallidum, Treponema denticola, and Borrelia burgdorferi have membrane topologies and PG binding sites highly similar to those of their well-characterized Escherichia coli and Helicobacter pylori orthologs. Collectively, our results help to clarify fundamental differences in cell envelope ultrastructure between spirochetes and gram-negative bacteria. They also confirm that PG stabilizes the flagellar motor and enable us to propose that in most spirochetes motility results from rotation of the flagellar filaments against the PG

The MUC1 Ectodomain: A Novel and Efficient Target for Gold Nanoparticle Clustering and Vapor Nanobubble Generation

MUC1 is a large, heavily glycosylated transmembrane glycoprotein that is proposed to create a protective microenvironment in many adenocarcinomas. Here we compare MUC1 and the well studied cell surface receptor target, EGFR, as gold nanoparticle (AuNP) targets and their subsequent vapor nanobubble generation efficacy in the human epithelial cell line, HES. Although EGFR and MUC1 were both highly expressed in these cells, TEM and confocal images revealed MUC1 as a superior target for nanoparticle intracellular accumulation and clustering. The MUC1-targeted AuNP intracellular clusters also generated significantly larger vapor nanobubbles. Our results demonstrate the promising opportunities MUC1 offers to improve the efficacy of targeted nanoparticle based approaches

MUC1: A multifunctional cell surface component of reproductive tissue epithelia

MUC1 is a large, transmembrane mucin glycoprotein expressed at the apical surface of a variety of reproductive tract epithelia. Functions attributed to MUC1 include those generally associated with mucins such as lubrication and hydration of cell surfaces as well as protection from microorganisms and degradative enzymes. In addition, MUC1 is an effective inhibitor of both cell-cell and cell-extracellular matrix interactions in both normal and malignant contexts. Moreover, a series of recent studies has shown that the highly conserved cytoplasmic tail of MUC1 interacts specifically with a series of important signal transducing molecules including β-catenin, Grb2 and erbB family members. MUC1 expression in normal epithelia can be quite dynamic, varying in response to steroid hormone or cytokine influences. Following malignant transformation, MUC1 often becomes highly overexpressed, loses its apical restriction, and displays aberrant glycosylation and altered mRNA splice variants. Regulation of MUC1 expression can occur at the transcriptional level. In addition, post-translational regulation of cell surface expression occurs via the activity of cell surface proteases or "sheddases" that release soluble forms of the large ectodomains. This review will briefly summarize studies of MUC1 expression and function in reproductive tissues with particular emphasis on the uterus. In addition, current knowledge of the mechanisms of MUC1 gene regulation, metabolic processing and potential signal transducing functions will be presented

Measuring the mass of the central black hole in the bulgeless galaxy ngc 4395 from gas dynamical modeling

NGC 4395 is a bulgeless spiral galaxy, harboring one of the nearest known type 1 Seyfert nuclei. Although there is no consensus on the mass of its central engine, several estimates suggest it is one of the lightest massive black holes (MBHs) known. We present the first direct dynamical measurement of the mass of this MBH from a combination of two-dimensional gas kinematic data, obtained with the adaptive optics assisted near-infrared integral field spectrograph Gemini/NIFS and high-resolution multiband photometric data from Hubble Space Telescope's Wide Field Camera 3. We use the photometric data to model the shape and stellar mass-to-light ratio of the nuclear star cluster (NSC). From the Gemini/NIFS observations, we derive the kinematics of warm molecular hydrogen gas as traced by emission through the H2 1–0 S(1) transition. These kinematics show a clear rotational signal, with a position angle orthogonal to NGC 4395's radio jet. Our best-fitting tilted ring models of the kinematics of the molecular hydrogen gas contain a black hole with mass M={4}-3+8× {10}5 M⊙ (3σ uncertainties) embedded in an NSC of mass M=2× {10}6 M⊙. Our black hole mass measurement is in excellent agreement with the reverberation mapping mass estimate of Peterson et al. but shows some tension with other mass measurement methods based on accretion signals

A heparin binding synthetic peptide from human HIP / RPL29 fails to specifically differentiate between anticoagulantly active and inactive species of heparin

Despite extensive progress in determining structures within heparin and heparan sulfate (Hp/HS) and the discovery of numerous proteinaceous binding partners for Hp/HS so far; the only detailed characterization of a specific protein-glycosaminoglycan interaction is antithrombin III (ATIII) binding to a Hp pentasaccharide containing a unique 3-O-sulfated glucosamine residue. Previously, it was reported from our laboratories that a 16 amino acid synthetic peptide derived from the C-terminus of human HIP/RPL29 (HIP peptide-1) enriched for ATIII-dependent anticoagulant activity, presumably by specifically binding the ATIII pentasaccharide. Herein, we demonstrate that HIP peptide-1 cannot enrich ATIII-dependent anticoagulant activity from a starting pool of porcine intestinal mucosa Hp through a bio-specific interaction. However, a HIP peptide-1 column can be used to enrich for anticoagulantly active Hp from a diverse pool of glycosaminoglycans known as Hp byproducts by a mechanism of nonspecific charge interactions. Thus, HIP peptide-1 cannot recognize Hp via bio-specific interactions but binds glycosaminoglycans by non-specific charge interactions

Reverberation Mapping of the Kepler-Field AGN KA1858+4850

KA1858+4850 is a narrow-line Seyfert 1 galaxy at redshift 0.078 and is among the brightest active galaxies monitored by the Kepler mission. We have carried out a reverberation mapping campaign designed to measure the broad-line region size and estimate the mass of the black hole in this galaxy. We obtained 74 epochs of spectroscopic data using the Kast Spectrograph at the Lick 3-m telescope from February to November of 2012, and obtained complementary V-band images from five other ground-based telescopes. We measured the Hbeta light curve lag with respect to the V-band continuum light curve using both cross-correlation techniques (CCF) and continuum light curve variability modeling with the JAVELIN method, and found rest-frame lags of CCF = 13.53+2.03 2.32 days and JAVELIN = 13.15+1.08 1.00 days. The Hbeta root-mean-square line profile has a width of sigma line = 770 +/- 49 km s(exp -1). Combining these two results and assuming a virial scale factor of f = 5.13, we obtained a virial estimate of M(sub BH) = 8.06+1.59 1.72 10(exp 6) solar mass for the mass of the central black hole and an Eddington ratio of L/L(sub Edd) (is) approx. 0.2. We also obtained consistent but slightly shorter emission-line lags with respect to the Kepler light curve. Thanks to the Kepler mission, the light curve of KA1858+4850 has among the highest cadences and signal-to-noise ratios ever measured for an active galactic nucleus; thus, our black hole mass measurement will serve as a reference point for relations between black hole mass and continuum variability characteristics in active galactic nuclei

Acute cycling exercise and hippocampal subfield function and microstructure in healthy older adults

Aging is associated with deterioration in dentate gyrus (DG) and CA3, both crucial hippocampal subfields for age susceptible memory processes such as mnemonic discrimination (MD). Meanwhile, a single aerobic exercise session alters DG/CA3 function and neural activity in both rats and younger adults and can elicit short-term microstructural alterations in the hippocampus of older adults. However, our understanding of the effects of acute exercise on hippocampal subfield integrity via function and microstructure in older adults is limited. Thus, a within subject-design was employed to determine if 20-min of moderate to vigorous aerobic exercise alters bilateral hippocampal subfield function and microstructure using high-resolution functional magnetic resonance imaging (fMRI) during an MD task (n = 35) and high angular resolution multi-shell diffusion imaging (n = 31), in healthy older adults, compared to seated rest. Following the exercise condition, participants exhibited poorer MD performance, particularly when their perception of effort was higher. Exercise was also related to lower MD-related activity within the DG/CA3 but not CA1 subfield. Finally, after controlling for whole brain gray matter diffusion, exercise was associated with lower neurite density index (NDI) within the DG/CA3. However, exercise-related differences in DG/CA3 activity and NDI were not associated with differences in MD performance. Our results suggest moderate to vigorous aerobic exercise may temporarily inhibit MD performance, and suppress DG/CA3 MD-related activity and NDI, potentially through neuroinflammatory/glial processes. However, additional studies are needed to confirm whether these short-term changes in behavior and hippocampal subfield neurophysiology are beneficial and how they might relate to long-term exercise habits