Reactivation of Epstein–Barr virus by a dual-responsive fluorescent EBNA1-targeting agent with Zn2+-chelating function

EBNA1 is the only Epstein–Barr virus (EBV) latent protein responsible for viral genome maintenance and is expressed in all EBV-infected cells. Zn2+ is essential for oligomerization of the functional EBNA1. We constructed an EBNA1 binding peptide with a Zn2+ chelator to create an EBNA1-specific inhibitor (ZRL5P4). ZRL5P4 by itself is sufficient to reactivate EBV from its latent infection. ZRL5P4 is able to emit unique responsive fluorescent signals once it binds with EBNA1 and a Zn2+ ion. ZRL5P4 can selectively disrupt the EBNA1 oligomerization and cause nasopharyngeal carcinoma (NPC) tumor shrinkage, possibly due to EBV lytic induction. Dicer1 seems essential for this lytic reactivation. As can been seen, EBNA1 is likely to maintain NPC cell survival by suppressing viral reactivation

Structural Variations in Carboxylated Bispidine Ligands: Influence of Positional Isomerism and Rigidity on the Conformation, Stability, Inertness and Relaxivity of their Mn 2+ Complexes

Abstract Mn 2+ complexes of 2,4‐pyridyl‐disubstituted bispidine ligands have emerged as more biocompatible alternatives to Gd 3+ ‐based MRI probes. They display relaxivities comparable to that of commercial contrast agents and high kinetic inertness, unprecedented for Mn 2+ complexes. The chemical structure, in particular the substituents on the two macrocyclic nitrogens N3 and N7, are decisive for the conformation of the Mn 2+ complexes, and this will in turn determine their thermodynamic, kinetic and relaxation properties. We describe the synthesis of four ligands with acetate substituents in positions N3, N7 or both. We evidence that the bispidine conformation is dependent on N3 substitution, with direct impact on the thermodynamic stability, kinetic inertness, hydration state and relaxivity of the Mn 2+ complexes. These results unambiguously show that (i) solely a chair‐chair conformation allows for favorable inertness and relaxivity, and (ii) in this family such chair‐chair conformation is accessible only for ligands without N3‐appended carboxylates

Substrate Sequence Determines Catalytic Activities, Domain-Binding Preferences, and Allosteric Mechanisms in Pin1

Pin1 is a unique phosphorylation-dependent peptidyl–prolyl isomerase that regulates diverse subcellular processes and an important potential therapeutic target. Functional mechanisms of Pin1 are complicated because of the two-domain structural organization: the catalytic domain both binds the specific pSer/Thr-Pro motif and catalyzes the cis/trans isomerization, whereas the WW domain can only bind the trans configuration and is speculated to be responsible for substrate-binding specificity. Numerous studies of Pin1 have led to two divergent conclusions on the functional role of the WW domain. One opinion states that the WW domain is an allosteric effector, and substrate binding to this domain modulates the binding and catalysis in the distal catalytic domain. The other opinion, however, argues that the WW domain does not have any allosteric role. Here, using molecular dynamics and binding free-energy calculations, we examine catalysis and allosteric mechanisms in Pin1 under various substrate- and WW-binding conditions. Our results reveal a strong substrate sequence dependency of catalysis, domain-binding preferences, and allosteric outputs in Pin1. Importantly, we show that the different opinions about the WW domain can be unified in one framework, in which substrate sequences determine whether a positive, negative, or neural allosteric effect will be elicited. Our work further elucidates detailed mechanisms underlying the sequence-dependent allostery of Pin1 and finds that interdomain contacts are key mediators of intraprotein allosteric communications. Our findings collectively provide new insights into the function of Pin1, which may facilitate the development of novel therapeutic drugs targeting Pin1 in the future

Energy Transfer Mechanism and Quantitative Modeling of Rate from an Antenna to a Lanthanide Ion

The excitation energy transfer (ET) pathway and mechanism from an organic antenna to a lanthanide ion has been the subject of discussion for many decades. In the case of europium (Eu3+), it has been suggested that the transfer originates from the ligand singlet state or a triplet state. Taking the lanthanide complex Eu(TTA)3(H2O)2 as an example, we have investigated the spectra and luminescence kinetics, mainly at room temperature and 77 K, to acquire the necessary experimental data. We put forward an experimental and theoretical approach to measure the energy transfer rates from the antenna to different Eu3+ levels using the Dexter formulation. We find that transfer from the ligand singlet state to Eu3+ may account for the ET pathway, by combined electric dipole–electric dipole (ED–ED) and ED-electric quadrupole (EQ) mechanisms. The contributions from the triplet state by these mechanisms are very small. An independent systems rate equation approach can effectively model the experimental kinetics results. The model utilizes the cooperative processes that take place on the metal ion and ligand and considers S0, S1, and T1 ligand states in addition to 7F0,1, 5D0, 5D1, and 5DJ (=5L6, 5D3, 5D2 combined) Eu3+ states. The triplet exchange ET rate is estimated to be of the order 107 s–1. The observation of a nanosecond risetime for the Eu3+ 5D1 level does not enable the assignment of the ET route or the mechanism. Furthermore, the 5D1 risetime may be contributed by several processes. Observation of its temperature dependence and also that of the ground-state population can supply useful information concerning the mechanism because the change in metal-ion internal conversion rate has a greater effect than changes in singlet or triplet nonradiative rates. A critical comparison is included for the model of Malta employed in the online software LUMPAC and JOYSpectra. The theoretical treatment of the exchange mechanism and its contribution are now being considered

Solution-State Cooperative Luminescence Upconversion in Molecular Ytterbium Dimers

International audienceTwo homometallic ytterbium dimers are prepared and their solution-state photoluminescence and upconversion properties are investigated. Both complexes exhibit two-photon cooperative luminescence upconversion in the visible region (lambda(em) approximate to 510 nm) upon excitation into the near-infrared Yb F-2(5/2) <- F-2(7/2) absorption band at 980 nm. This miniaturization of the cooperative luminescence phenomenon down to just two Yb ions unequivocally proves the mechanistic origins of this process. Time-resolved measurements and excited-state modeling indicate the presence of a slow recombination of two singly excited ions Yb*Yb* into a virtual excited state, which ultimately gives rise to the observed emission at approximate to 510 nm

Upconversion in a d–f [RuYb 3 ] Supramolecular Assembly

International audienceWe have prepared a hetero-tetrametallic assembly consisting of three ytterbium ions coordinated to a central [Ru(bpm)3] 2+ (bpm = 2,2'-bipyrimidine) motif. Irradiation into the absorption band of the peripheral ytterbium ions at 980 nm engenders emission of the 3 MLCT state of the central [Ru(bpm)3] 2+ core at 636 nm, which represents the first example of f→d molecular upconversion (UC). Time-resolved measurements reveal a slow rise of the UC emission, which was modeled with a mathematic treatment of the observed kinetics according to a cooperative photosensitization mechanism using a virtual Yb centered doubly excited state followed by energy transfer to the Ru centered 1 MLCT state

Lanthanide–Cyclen–Camptothecin Nanocomposites for Cancer Theranostics Guided by Near-Infrared and Magnetic Resonance Imaging

We have devised a molecular-to-micellar strategy to incorporate a lanthanide nanoplatform for the delivery of an anticancer drug that simultaneously offers hybrid near-infrared (NIR) and magnetic resonance imaging (MRI) capabilities with defined lanthanide(III) ratio control. This cancer-selective lanthanide-based self-assembled nanocomposite (LnNPs) has been synthesized by conjugating lanthanide–cyclen complexes (cycLn) with a well-known drug–camptothecin (CPT) through a redox-sensitive disulfide bond (−ss–). By accurately controlling the ratio of Gd(III) and Yb(III) complexes, we prepared hybrid nanoparticles (Gd/YbNPs) with both NIR and MR imaging properties. The enhanced stability at ultralow critical aggregation concentrations (CACs), simultaneous optical and MR imaging, improved delivery/chemotherapeutic efficiency, and cancer cell selectivity of such nanomicellar theranostic prodrugs in vitro and in vivo have thus been achieved and validated. The work provides a blueprint combining a stimuli-activated NIR luminescence and real-time MR imaging into a safe and biocompatible nanoplatform for selective cancer treatment

Erratum: Reactivation of Epstein–Barr virus by a dual-responsive fluorescent EBNA1-targeting agent with Zn2+-chelating function (Proceedings of the National Academy of Sciences of the United States of America (2019) 116 (26614-26624) DOI: 10.1073/pnas.1915372116)

Correction for “Reactivation of Epstein–Barr virus by a dualresponsive fluorescent EBNA1-targeting agent with Zn2+- chelating function,” by Lijun Jiang, Hong Lok Lung, Tao Huang, Rongfeng Lan, Shuai Zha, Lai Sheung Chan, Waygen Thor, Tik-Hung Tsoi, Ho-Fai Chau, Cecilia Boreström, Steven L. Cobb, Sai Wah Tsao, Zhao-Xiang Bian, Ga-Lai Law, Wing-Tak Wong, William Chi-Shing Tai, Wai Yin Chau, Yujun Du, Lucas Hao Xi Tang, Alan Kwok Shing Chiang, Jaap M. Middeldorp, Kwok-Wai Lo, Nai Ki Mak, Nicholas J. Long, and Ka-Leung Wong, which was first published December 10, 2019; 10.1073/pnas.1915372116 (Proc. Natl. Acad. Sci. U.S.A. 116, 26614–26624). The authors note that Fig. 6 appeared incorrectly. Part of panel D of the published figure was inadvertently omitted. The corrected figure and its legend appear below. (Figure Presented)

Reactivation of Epstein-Barr virus by a dual-responsive fluorescent EBNA1-targeting agent with Zn2+-chelating function

Epstein-Barr nuclear antigen 1 (EBNA1) plays a vital role in the maintenance of the viral genome and is the only viral protein expressed in nearly all forms of Epstein-Barr virus (EBV) latency and EBV-associated diseases, including numerous cancer types. To our knowledge, no specific agent against EBV genes or proteins has been established to target EBV lytic reactivation. Here we report an EBNA1- and Zn2+-responsive probe (ZRL5P4) which alone could reactivate the EBV lytic cycle through specific disruption of EBNA1. We have utilized the Zn2+ chelator to further interfere with the higher order of EBNA1 self-association. The bioprobe ZRL5P4 can respond independently to its interactions with Zn2+ and EBNA1 with different fluorescence changes. It can selectively enter the nuclei of EBV-positive cells and disrupt the oligomerization and oriP-enhanced transactivation of EBNA1. ZRL5P4 can also specifically enhance Dicer1 and PML expression, molecular events which had been reported to occur after the depletion of EBNA1 expression. Importantly, we found that treatment with ZRL5P4 alone could reactivate EBV lytic induction by expressing the early and late EBV lytic genes/proteins. Lytic induction is likely mediated by disruption of EBNA1 oligomerization and the subsequent change of Dicer1 expression. Our probe ZRL5P4 is an EBV proteinspecific agent that potently reactivates EBV from latency, leading to the shrinkage of EBV-positive tumors, and our study also suggests the association of EBNA1 oligomerization with the maintenance of EBV latency