Interaction of a Dimeric Single-Stranded DNA-Binding Protein (G5P) with DNA Hairpins. A Molecular Beacon Study

Gene-V protein (G5P/GVP) is a single-stranded (ss)DNA-binding protein (SBP) of bacteriophage f1 that is required for DNA synthesis and repair. In solution, it exists as a dimer that binds two antiparallel ssDNA strands with high affinity in a cooperative manner, forming a left-handed helical protein–DNA filament. Here, we report on fluorescence studies of the interaction of G5P with different DNA oligonucleotides having a hairpin structure (molecular beacon, MB) with a seven base-pair stem (dT24-stem7, dT18-stem7), as well as with DNA oligonucleotides (dT38, dT24) without a defined secondary structure. All oligonucleotides were end-labeled with a Cy3-fluorophore and a BHQ2-quencher. In the case of DNA oligonucleotides without a secondary structure, an almost complete quenching of their strong fluorescence (with about 5% residual intensity) was observed upon the binding of G5P. This implies an exact alignment of the ends of the DNA strand(s) in the saturated complex. The interaction of the DNA hairpins with G5P led to the unzipping of the base-paired stem, as revealed by fluorescence measurements, fluorescence microfluidic mixing experiments, and electrophoretic mobility shift assay data. Importantly, the disruption of ssDNA’s secondary structure agrees with the behavior of other single-stranded DNA-binding proteins (SBPs). In addition, substantial protein-induced fluorescence enhancement (PIFE) of the Cy3-fluorescence was observed

PRKACA Mediates Resistance to HER2-Targeted Therapy in Breast Cancer Cells and Restores Anti-Apoptotic Signaling

Targeting HER2 with antibodies or small molecule inhibitors in HER2-positive breast cancer leads to improved survival, but resistance is a common clinical problem. To uncover novel mechanisms of resistance to anti-HER2 therapy in breast cancer, we performed a kinase open reading frame (ORF) screen to identify genes that rescue HER2-amplified breast cancer cells from HER2 inhibition or suppression. In addition to multiple members of the MAPK and PI3K signaling pathways, we discovered that expression of the survival kinases PRKACA and PIM1 rescued cells from anti-HER2 therapy. Furthermore, we observed elevated PRKACA expression in trastuzumab-resistant breast cancer samples, indicating that this pathway is activated in breast cancers that are clinically resistant to trastuzumab-containing therapy. We found that neither PRKACA nor PIM1 restored MAPK or PI3K activation after lapatinib or trastuzumab treatment, but rather inactivated the pro-apoptotic protein BAD, thereby permitting survival signaling through BCL-XL. Pharmacological blockade of BCL-XL/BCL-2 partially abrogated the rescue effects conferred by PRKACA and PIM1, and sensitized cells to lapatinib treatment. These observations suggest that combined targeting of HER2 and the BCL-XL/BCL-2 anti-apoptotic pathway may increase responses to anti-HER2 therapy in breast cancer and decrease the emergence of resistant disease

Efficacy of Carbonic Anhydrase Inhibitors on Cystoid Fluid Collections and Visual Acuity in Patients with X-Linked Retinoschisis

Purpose: To date, there is no standard treatment regimen for carbonic anhydrase inhibitors (CAIs) in X-linked retinoschisis (XLRS) patients. This retrospective study aims to evaluate the efficacy of CAIs on visual acuity and cystoid fluid collections (CFC) in XRLS patients in Dutch and Belgian tertiary referral centers. Design: Retrospective cohort study. Participants: Forty-two patients with XLRS. Methods: In total, 42 patients were enrolled. To be included, patients had to have previous treatment with an oral CAI (acetazolamide), a topical CAI (brinzolamide/dorzolamide), or a combination of an oral and a topical CAI for at least 4 consecutive weeks. We evaluated the effect of the CAI on best-corrected visual acuity (BCVA) and central foveal thickness (CFT) on OCT. Main Outcome Measures: Central foveal thickness and BCVA. Results: The median age at the baseline visit of the patients in this cohort study was 14.7 (range, 43.6) years, with a median (interquartile range [IQR]) follow-up period of 4.0 (2.2–5.2) years. During the follow-up period, 25 patients were treated once with an oral CAI (60%), 24 patients were treated once with a topical CAI (57%), and 11 patients were treated once with a combination of both topical and oral CAI (26%). We observed a significant reduction of CFT for oral CAI by 14.37 μm per 100 mg per day (P &lt; 0.001; 95% confidence interval [CI], −19.62 to −9.10 μm) and for topical CAI by 7.52 μm per drop per day (P = 0.017; 95% CI, −13.67 to −1.32 μm). The visual acuity changed significantly while on treatment with oral CAI by −0.0059 logMAR per 100 mg (P = 0.008; 95% CI, −0.010 to −0.0013 logMAR). Seven patients (17%) had side effects leading to treatment discontinuation. Conclusions: Our data indicate that treatment with (oral) CAI may be beneficial for short-term management of CFC in patients with XLRS. Despite a significant reduction in CFT, the change in visual acuity was modest and not of clinical significance. Nonetheless, the anatomic improvement of the central retina in these patients may be of value to create an optimal retinal condition for future potential treatment options such as gene therapy. Financial Disclosure(s): The authors have no proprietary or commercial interest in any materials discussed in this article.</p

Efficacy of Carbonic Anhydrase Inhibitors on Cystoid Fluid Collections and Visual Acuity in Patients with X-Linked Retinoschisis

Purpose: To date, there is no standard treatment regimen for carbonic anhydrase inhibitors (CAIs) in X-linked retinoschisis (XLRS) patients. This retrospective study aims to evaluate the efficacy of CAIs on visual acuity and cystoid fluid collections (CFC) in XRLS patients in Dutch and Belgian tertiary referral centers. Design: Retrospective cohort study. Participants: Forty-two patients with XLRS. Methods: In total, 42 patients were enrolled. To be included, patients had to have previous treatment with an oral CAI (acetazolamide), a topical CAI (brinzolamide/dorzolamide), or a combination of an oral and a topical CAI for at least 4 consecutive weeks. We evaluated the effect of the CAI on best-corrected visual acuity (BCVA) and central foveal thickness (CFT) on OCT. Main Outcome Measures: Central foveal thickness and BCVA. Results: The median age at the baseline visit of the patients in this cohort study was 14.7 (range, 43.6) years, with a median (interquartile range [IQR]) follow-up period of 4.0 (2.2–5.2) years. During the follow-up period, 25 patients were treated once with an oral CAI (60%), 24 patients were treated once with a topical CAI (57%), and 11 patients were treated once with a combination of both topical and oral CAI (26%). We observed a significant reduction of CFT for oral CAI by 14.37 μm per 100 mg per day (P &lt; 0.001; 95% confidence interval [CI], −19.62 to −9.10 μm) and for topical CAI by 7.52 μm per drop per day (P = 0.017; 95% CI, −13.67 to −1.32 μm). The visual acuity changed significantly while on treatment with oral CAI by −0.0059 logMAR per 100 mg (P = 0.008; 95% CI, −0.010 to −0.0013 logMAR). Seven patients (17%) had side effects leading to treatment discontinuation. Conclusions: Our data indicate that treatment with (oral) CAI may be beneficial for short-term management of CFC in patients with XLRS. Despite a significant reduction in CFT, the change in visual acuity was modest and not of clinical significance. Nonetheless, the anatomic improvement of the central retina in these patients may be of value to create an optimal retinal condition for future potential treatment options such as gene therapy. Financial Disclosure(s): The authors have no proprietary or commercial interest in any materials discussed in this article.</p

Precision Top-Quark Mass Measurements at CDF

We present a precision measurement of the top-quark mass using the full sample of Tevatron $\sqrt{s}=1.96$ TeV proton-antiproton collisions collected by the CDF II detector, corresponding to an integrated luminosity of 8.7 $fb^{-1}$. Using a sample of $t\bar{t}$ candidate events decaying into the lepton+jets channel, we obtain distributions of the top-quark masses and the invariant mass of two jets from the $W$ boson decays from data. We then compare these distributions to templates derived from signal and background samples to extract the top-quark mass and the energy scale of the calorimeter jets with {\it in situ} calibration. The likelihood fit of the templates from signal and background events to the data yields the single most-precise measurement of the top-quark mass, \mtop = 172.85 \pm$0.71 (stat)$\pm$0.85 (syst) GeV/c^{2}.$Comment: submitted to Phys. Rev. Let

Evidence for t\bar{t}\gamma Production and Measurement of \sigma_t\bar{t}\gamma / \sigma_t\bar{t}

Using data corresponding to 6.0/fb of ppbar collisions at sqrt(s) = 1.96 TeV collected by the CDF II detector, we present a cross section measurement of top-quark pair production with an additional radiated photon. The events are selected by looking for a lepton, a photon, significant transverse momentum imbalance, large total transverse energy, and three or more jets, with at least one identified as containing a b quark. The ttbar+photon sample requires the photon to have 10 GeV or more of transverse energy, and to be in the central region. Using an event selection optimized for the ttbar+photon candidate sample we measure the production cross section of, and the ratio of cross sections of the two samples. Control samples in the dilepton+photon and lepton+photon+\met, channels are constructed to aid in decay product identification and background measurements. We observe 30 ttbar+photon candidate events compared to the standard model expectation of 26.9 +/- 3.4 events. We measure the ttbar+photon cross section to be 0.18+0.08 pb, and the ratio of the cross section of ttbar+photon to ttbar to be 0.024 +/- 0.009. Assuming no ttbar+photon production, we observe a probability of 0.0015 of the background events alone producing 30 events or more, corresponding to 3.0 standard deviations.Comment: 9 pages, 3 figure

Measurement of branching ratio and Bs0 lifetime in the decay Bs0 -> J/psi f0(980) at CDF

We present a study of Bs0 decays to the CP-odd final state J/psi f0(980) with J/psi -> mu+ mu- and f0(980) -> pi+ pi-. Using ppbar collision data with an integrated luminosity of 3.8/fb collected by the CDF II detector at the Tevatron we measure a Bs0 lifetime of tau(Bs0 -> J/psi f0(980)) = 1.70 -0.11+0.12(stat) +-0.03(syst) ps. This is the first measurement of the Bs0 lifetime in a decay to a CP eigenstate and corresponds in the standard model to the lifetime of the heavy Bs0 eigenstate. We also measure the product of branching fractions of Bs0 -> J/psi f0(980) and f0(980) -> pi+ pi- relative to the product of branching fractions of Bs0 -> J/psi phi and phi -> K+ K- to be R_f0/phi = 0.257 +_0.020(stat) +-0.014(syst), which is the most precise determination of this quantity to date.Comment: accepted by Phys. Rev.

A search for resonant production of ttˉt\bar{t} pairs in $4.8\ \rm{fb}^{-1}ofintegratedluminosityof of integrated luminosity of p\bar{p}collisionsat collisions at \sqrt{s}=1.96\ \rm{TeV}$

We search for resonant production of tt pairs in 4.8 fb^{-1} integrated luminosity of ppbar collision data at sqrt{s}=1.96 TeV in the lepton+jets decay channel, where one top quark decays leptonically and the other hadronically. A matrix element reconstruction technique is used; for each event a probability density function (pdf) of the ttbar candidate invariant mass is sampled. These pdfs are used to construct a likelihood function, whereby the cross section for resonant ttbar production is estimated, given a hypothetical resonance mass and width. The data indicate no evidence of resonant production of ttbar pairs. A benchmark model of leptophobic Z \rightarrow ttbar is excluded with m_{Z'} < 900 GeV at 95% confidence level.Comment: accepted for publication in Physical Review D Sep 21, 201

Combined search for the standard model Higgs boson decaying to a bb pair using the full CDF data set

We combine the results of searches for the standard model Higgs boson based on the full CDF Run II data set obtained from sqrt(s) = 1.96 TeV p-pbar collisions at the Fermilab Tevatron corresponding to an integrated luminosity of 9.45/fb. The searches are conducted for Higgs bosons that are produced in association with a W or Z boson, have masses in the range 90-150 GeV/c^2, and decay into bb pairs. An excess of data is present that is inconsistent with the background prediction at the level of 2.5 standard deviations (the most significant local excess is 2.7 standard deviations).Comment: To be published in Phys. Rev. Lett (v2 contains minor updates based on comments from PRL

Measurements of branching fraction ratios and CP-asymmetries in suppressed B^- -> D(-> K^+ pi^-)K^- and B^- -> D(-> K^+ pi^-)pi^- decays

We report the first reconstruction in hadron collisions of the suppressed decays B^- -> D(-> K^+ pi^-)K^- and B^- -> D(-> K^+ pi^-)pi^-, sensitive to the CKM phase gamma, using data from 7 fb^-1 of integrated luminosity collected by the CDF II detector at the Tevatron collider. We reconstruct a signal for the B^- -> D(-> K^+ pi^-)K^- suppressed mode with a significance of 3.2 standard deviations, and measure the ratios of the suppressed to favored branching fractions R(K) = [22.0 \pm 8.6(stat)\pm 2.6(syst)]\times 10^-3, R^+(K) = [42.6\pm 13.7(stat)\pm 2.8(syst)]\times 10^-3, R^-(K)= [3.8\pm 10.3(stat)\pm 2.7(syst]\times 10^-3, as well as the direct CP-violating asymmetry A(K) = -0.82\pm 0.44(stat)\pm 0.09(syst) of this mode. Corresponding quantities for B^- -> D(-> K^+ pi^-)pi^- decay are also reported.Comment: 8 pages, 1 figure, accepted by Phys.Rev.D Rapid Communications for Publicatio