Direct regulation of Treslin by cyclin-dependent kinase is essential for the onset of DNA replication

Treslin, a TopBP1-interacting protein, is necessary for deoxyribonucleic acid (DNA) replication in vertebrates. Association between Treslin and TopBP1 requires cyclin-dependent kinase (Cdk) activity in Xenopus laevis egg extracts. We investigated the mechanism and functional importance of Cdk for this interaction using both X. laevis egg extracts and human cells. We found that Treslin also associated with TopBP1 in a Cdk-regulated manner in human cells and that Treslin was phosphorylated within a conserved Cdk consensus target sequence (on S976 in X. laevis and S1000 in humans). Recombinant human Cdk2–cyclin E also phosphorylated this residue of Treslin in vitro very effectively. Moreover, a mutant of Treslin that cannot undergo phosphorylation on this site showed significantly diminished binding to TopBP1. Finally, human cells harboring this mutant were severely deficient in DNA replication. Collectively, these results indicate that Cdk-mediated phosphorylation of Treslin during S phase is necessary for both its effective association with TopBP1 and its ability to promote DNA replication in human cells

Ataxia-telangiectasia Mutated (ATM)-dependent Activation of ATR Occurs through Phosphorylation of TopBP1 by ATM

ATM (ataxia-telangiectasia mutated) is necessary for activation of Chk1 by ATR (ATM and Rad3-related) in response to double-stranded DNA breaks (DSBs) but not to DNA replication stress. TopBP1 has been identified as a direct activator of ATR. We show that ATM regulates Xenopus TopBP1 by phosphorylating Ser-1131 and thereby strongly enhancing association of TopBP1 with ATR. Xenopus egg extracts containing a mutant of TopBP1 that cannot be phosphorylated on Ser-1131 are defective in the ATR-dependent phosphorylation of Chk1 in response to DSBs but not to DNA replication stress. Thus, TopBP1 is critical for the ATM-dependent activation of ATR following production of DSBs in the genome

Roles of replication fork-interacting and Chk1-activating domains from claspin in a DNA replication checkpoint response

Claspin is essential for the ATR-dependent activation of Chk1 in Xenopus egg extracts containing incompletely replicated DNA. Claspin associates with replication forks upon origin unwinding. We show that Claspin contains a replication fork-interacting domain (RFID, residues 265–605) that associates with Cdc45, DNA polymerase ε, replication protein A, and two replication factor C complexes on chromatin. The RFID contains two basic patches (BP1 and BP2) at amino acids 265–331 and 470–600, respectively. Deletion of either BP1 or BP2 compromises optimal binding of Claspin to chromatin. Absence of BP1 has no effect on the ability of Claspin to mediate activation of Chk1. By contrast, removal of BP2 causes a large reduction in the Chk1-activating potency of Claspin. We also find that Claspin contains a small Chk1-activating domain (residues 776–905) that does not bind stably to chromatin, but it is fully effective at high concentrations for mediating activation of Chk1. These results indicate that stable retention of Claspin on chromatin is not necessary for activation of Chk1. Instead, our findings suggest that only transient interaction of Claspin with replication forks potentiates its Chk1-activating function. Another implication of this work is that stable binding of Claspin to chromatin may play a role in other functions besides the activation of Chk1

Resonance at the Rabi frequency in a superconducting flux qubit

We analyze a system composed of a superconducting flux qubit coupled to a transmission-line resonator driven by two signals with frequencies close to the resonator's harmonics. The first strong signal is used for exciting the system to a high energetic state while a second weak signal is applied for probing effective eigenstates of the system. In the framework of doubly dressed states we showed the possibility of amplification and attenuation of the probe signal by direct transitions at the Rabi frequency. We present a brief review of theoretical and experimental works where a direct resonance at Rabi frequency have been investigated in superconducting flux qubits. The interaction of the qubit with photons of two harmonics has prospects to be used as a quantum amplifier (microwave laser) or an attenuator.Comment: This paper is the extended version of the talk given by one of the authors at the Conference On Nuclei And Mesoscopic Physics, 5-9 May 2014, Michigan State University, East Lansing, US

Sociolinguistic Perspective on Teaching English Intonation for Adult Learners

AbstractIn this paper, we aim at identifying the efficiency of teaching English intonation at the university level. We show how sociolinguistic approach with the focus on conscious control of using intonation can enhance the communicative performance of linguistic students at Novosibirsk State Technical University. We present a teaching model based on appealing to senses, on making students’ egos more permeable and on creating favorable socio- and psychological conditions conducive to the acquisition of the second language intonation. Finally, we make some suggestions on how to integrate cognitive, emotional and psychological perspectives in the language learning process