LEDGF/p75 is critical but not essential for multiple-round HIV 1 replication

status: publishe

MicroRNA response to hypoxic stress in soft tissue sarcoma cells: MicroRNA mediated regulation of HIF3α

Background: Hypoxia is often encountered in solid tumors and known to contribute to aggressive tumor behavior, radiation- and chemotherapy resistance resulting in a poor prognosis for the cancer patient. MicroRNAs (miRNAs) play a role in the regulation of the tumor cell response to hypoxia, however, not much is known about the involvement of miRNAs in hypoxic signalling pathways in soft tissue sarcomas (STS).Method: A panel of twelve STS cell lines was exposed to atmospheric oxygen concentrations (normoxia) or 1% oxygen (hypoxia) for up to 48 h. Hypoxic conditions were verified and miRNA expression profiles were assessed by LNA™ oligonucleotide microarrays and RT-PCR after 24 h. The expression of target genes regulated by hypoxia responsive miRNAs is examined by end-point PCR and validated by luciferase reporter constructs.Results: Exposure of STS cell lines to hypoxic conditions gave rise to upregulation of Hypoxia Inducible Factor (HIF) 1α protein levels and increased mRNA expression of HIF1 target genes CA9 and VEGFA. Deregulation of miRNA expression after 24 h of hypoxia was observed. The most differentially expressed miRNAs (p < 0.001) in response to hypoxia were miR-185-3p, miR-485-5p, miR-216a-5p (upregulated) and miR-625-5p (downregulated). The well-known hypoxia responsive miR-210-3p could not be reliably detected by the microarray platform most likely for technical reasons, however, its upregulation upon hypoxic stress was apparent by qPCR. Target prediction algorithms identified 11 potential binding sites for miR-485-5p and a single putative miR-210-3p binding site in the 3'UTR of HIF3α, the least studied member of the HIF family. We showed that HIF3α transcripts, expressing a 3'UTR containing the miR-485-5p and miR-210-3p target sites, are expressed in all sarcoma cell lines and upregulated upon hypoxia. Additionally, luciferase reporter constructs containing the 3'UTR of HIF3α were used to demonstrate regulation of HIF3α by miR-210-3p and miR-485-5p.Conclusion: Here we provide evidence for the miRNA mediated regulation of HIF3α by hypoxia responsive miRNAs in STS, which may help to tightly regulate and fine-tune the hypoxic response. This provides a better insight into the mechanisms underlying the hypoxic response in STS and may ultimately yield information on novel prognostic and predictive markers or targets for treatment

Constraints on large extra dimensions from the MINOS experiment

We report new constraints on the size of large extra dimensions from data collected by the MINOS experiment between 2005 and 2012. Our analysis employs a model in which sterile neutrinos arise as Kaluza-Klein states in large extra dimensions and thus modify the neutrino oscillation probabilities due to mixing between active and sterile neutrino states. Using Fermilab\u27s Neutrinos at the Main Injector beam exposure of 10.56 x 10(20) protons on target, we combine muon neutrino charged current and neutral current data sets from the Near and Far Detectors and observe no evidence for deviations from standard three-flavor neutrino oscillations. The ratios of reconstructed energy spectra in the two detectors constrain the size of large extra dimensions to be smaller than 0.45 mu m at 90% C.L. in the limit of a vanishing lightest active neutrino mass. Stronger limits are obtained for nonvanishing masses

Search for Sterile Neutrinos Mixing with Muon Neutrinos in MINOS

We report results of a search for oscillations involving a light sterile neutrino over distances of 1.04 and 735 km in a nu(mu)-dominated beam with a peak energy of 3 GeV. The data, from an exposure of 10.56 x 10(20) protons on target, are analyzed using a phenomenological model with one sterile neutrino. We constrain the mixing parameters theta(24) and Delta m(41)(2) and set limits on parameters of the four-dimensional Pontecorvo-Maki-Nakagawa-Sakata matrix, vertical bar U-mu 4 vertical bar(2) and vertical bar U-tau 4 vertical bar(2), under the assumption that mixing between nu(e) and nu(s) is negligible (vertical bar U-e4 vertical bar(2) = 0). No evidence for nu(mu) -\u3e nu(s) transitions is found and we set a world-leading limit on theta(24) for values of Delta m(41)(2) less than or similar to 1 eV(2)

Measurement of single pi(0) production by coherent neutral-current nu Fe interactions in the MINOS Near Detector

Forward single pi(0) production by coherent neutral-current interactions, vA - \u3e vA pi(0), is investigated using a 2.8 x 10(20) protons-on-target exposure of the MINOS Near Detector. For single-shower topologies, the event distribution in production angle exhibits a clear excess above the estimated background at very forward angles for visible energy in the range 1-8 GeV. Cross sections are obtained for the detector medium comprised of 80% iron and 20% carbon nuclei with (A) = 48, the highest- \u3c A \u3e target used to date in the study of this coherent reaction. The total cross section for coherent neutral-current single pi(0) production initiated by the v(mu) flux of the NuMI low-energy beam with mean (mode) E-v of 4.9 GeV (3.0 GeV), is 77.6 +/- 5.0 (stat)(-) (+15.0)(16.8) (syst) x 10(-40) cm(2) pernucleus. The results are in good agreement with predictions of the Berger-Sehgal model

LEDGF/p75-Independent HIV-1 Replication Demonstrates a Role for HRP-2 and Remains Sensitive to Inhibition by LEDGINs

Lens epithelium–derived growth factor (LEDGF/p75) is a cellular cofactor of HIV-1 integrase (IN) that interacts with IN through its IN binding domain (IBD) and tethers the viral pre-integration complex to the host cell chromatin. Here we report the generation of a human somatic LEDGF/p75 knockout cell line that allows the study of spreading HIV-1 infection in the absence of LEDGF/p75. By homologous recombination the exons encoding the LEDGF/p75 IBD (exons 11 to 14) were knocked out. In the absence of LEDGF/p75 replication of laboratory HIV-1 strains was severely delayed while clinical HIV-1 isolates were replication-defective. The residual replication was predominantly mediated by the Hepatoma-derived growth factor related protein 2 (HRP-2), the only cellular protein besides LEDGF/p75 that contains an IBD. Importantly, the recently described IN-LEDGF/p75 inhibitors (LEDGINs) remained active even in the absence of LEDGF/p75 by blocking the interaction with the IBD of HRP-2. These results further support the potential of LEDGINs as allosteric integrase inhibitors

Search for flavor-changing nonstandard neutrino interactions using nu(e) appearance in MINOS

We report new constraints on flavor-changing nonstandard neutrino interactions from the MINOS long-baseline experiment using nu(e) and (nu) over bar (e) appearance candidate events from predominantly nu(mu) and (nu) over bar (mu) beams. We used a statistical selection algorithm to separate nu(e) candidates from background events, enabling an analysis of the combined MINOS neutrino and antineutrino data. We observe no deviations from standard neutrino mixing, and thus place constraints on the nonstandard interaction matter effect, vertical bar epsilon(e tau)vertical bar, and phase, (delta(CP) + delta(e tau)), using a 30-bin likelihood fit

Improved Constraints on Sterile Neutrino Mixing from Disappearance Searches in the MINOS, MINOS+, Daya Bay, and Bugey-3 Experiments.

Searches for electron antineutrino, muon neutrino, and muon antineutrino disappearance driven by sterile neutrino mixing have been carried out by the Daya Bay and MINOS+ collaborations. This Letter presents the combined results of these searches, along with exclusion results from the Bugey-3 reactor experiment, framed in a minimally extended four-neutrino scenario. Significantly improved constraints on the θ_{μe} mixing angle are derived that constitute the most constraining limits to date over five orders of magnitude in the mass-squared splitting Δm_{41}^{2}, excluding the 90%&nbsp;C.L. sterile-neutrino parameter space allowed by the LSND and MiniBooNE observations at 90%&nbsp;CL_{s} for Δm_{41}^{2}&lt;13  eV^{2}. Furthermore, the LSND and MiniBooNE 99%&nbsp;C.L. allowed regions are excluded at 99% CL_{s} for Δm_{41}^{2}&lt;1.6 eV^{2}

Peptides derived from the HIV-1 integrase promote HIV-1 infection and multi-integration of viral cDNA in LEDGF/p75-knockdown cells

<p>Abstract</p> <p>Background</p> <p>The presence of the cellular Lens Epithelium Derived Growth Factor p75 (LEDGF/p75) protein is essential for integration of the Human immunodeficiency virus type 1 (HIV-1) cDNA and for efficient virus production. In the absence of LEDGF/p75 very little integration and virus production can be detected, as was demonstrated using LEDGF/p75-knokdown cells.</p> <p>Results</p> <p>Here we show that the failure to infect LEDGF/p75-knockdown cells has another reason aside from the lack of LEDGF/p75. It is also due to inhibition of the viral integrase (IN) enzymatic activity by an early expressed viral Rev protein. The formation of an inhibitory Rev-IN complex in virus-infected cells can be disrupted by the addition of three IN-derived, cell-permeable peptides, designated INr (IN derived-Rev interacting peptides) and INS (IN derived-integrase stimulatory peptide). The results of the present work confirm previous results showing that HIV-1 fails to infect LEDGF/p75-knockdown cells. However, in the presence of INrs and INS peptides, relatively high levels of viral cDNA integration as well as productive virus infection were obtained following infection by a wild type (WT) HIV-1 of LEDGF/p75-knockdown cells.</p> <p>Conclusions</p> <p>It appears that the lack of integration observed in HIV-1 infected LEDGF/p75-knockdown cells is due mainly to the inhibitory effect of Rev following the formation of a Rev-IN complex. Disruption of this inhibitory complex leads to productive infection in those cells.</p

Search for flavor-changing nonstandard neutrino interactions using νe appearance in MINOS

We report new constraints on flavor-changing nonstandard neutrino interactions from the MINOS longbaseline experiment using νe and ¯νe appearance candidate events from predominantly νμ and ¯νμ beams. We used a statistical selection algorithm to separate νe candidates from background events, enabling an analysis of the combined MINOS neutrino and antineutrino data. We observe no deviations from standard neutrino mixing, and thus place constraints on the nonstandard interaction matter effect, |εeτ|, and phase, (δCP + δeτ), using a 30-bin likelihood fit