Small molecules as research tools for studying the biology of the tumour suppressor p53

p53 is a potent tumour suppressor that is inactivated in the majority, if not all human cancers. In about 50% of the cases, the gene is mutated and in the rest it is rendered inactive mainly by deregulation of its negative regulators such as Hdm2 and HdmX. Targeting p53 has been shown to be a promising strategy to fight cancer and the first compounds reactivating p53 have reached the stage of clinical trials. In addition to immediate clinical use, compounds that activate p53 are valuable tools to increase the knowledge about fundamental p53 biology. RITA is an example for such a compound activating p53. We found that it induces decreased protein levels of the negative regulator of p53, HdmX, in a p53-dependent manner. This is mediated by ATM-induced phosphorylation. In addition expression of Wip1 is inhibited. Wip1 depletion is an important event for HdmX decrease, as Wip1 reverses ATM mediated phosphorylation and thus can prevent ATM-induced HdmX decline. The biological significance of HdmX and Wip1 inhibition is highlighted by the fact that a knockdown of either of them enhances cell killing by the p53-activating drugs RITA and nutlin3a. To get insight into mechanisms of p53 mediated transcriptional regulation, we compared genome-wide chromatin occupancy by p53 upon its activation by three different compounds, RITA, nutlin3a and 5-FU that cause different biological outcomes. We compared genome-wide chromatin occupancy by p53. Surprisingly, the regions that were bound by p53 to the highest extent were the same after all three treatments, despite their different biological outcomes. Comparison of the p53- occupied sites with gene expression changes upon p53 activation by nutlin3a allowed identifying 280 previously unknown target genes. The common p53 binding motif is present much more frequently in the promoter region of induced genes than in that of repressed genes. This suggests different mechanisms for gene induction versus repression by p53, presumably distinguished by the involvement of cofactors. This is in line with our finding that binding sites for cofactors in the proximity of p53 sites are distinct for induced and repressed genes. We identified AURKA (Aurora kinase A) as a novel p53-repressed target gene, and found that STAT3 also regulates it, antagonising p53. Finally we found, that expression of our newly identified panel of p53 target genes correlated with the p53 status, the grade of tumours and the long-term survival in a set of 250 breast cancer patients. Malignant melanomas have very poor prognosis with extremely low long-term survival once the metastatic stage is reached. It has been shown that in 3-dimensional collagen type I matrix that mimics the microenvironment of the human dermis, melanoma cells induce integrin-dependent inactivation of p53, rescuing the cells from apoptosis. Thus, reactivation of p53 might be a promising strategy to kill melanoma cells. To test this, we treated melanoma cells that were grown in 3D conditions with the p53 reactivating compound PRIMA-1 MET/APR-246. Indeed, this induced apoptosis in the cells in a p53 dependent manner. Consistently, the growth of melanoma xenografts in mice was suppressed in a p53-dependent manner after PRIMA-1 MET treatment. In summary, this thesis demonstrates examples for both the use of p53 activating compounds as research tools to uncover new details of p53 biology as well as their application for therapy, exemplified by effects of PRIMA-1 MET in malignant melanomas in vitro and in vivo

Degradation of III-V quantum dot lasers grown directly on silicon substrates

Initial age-related degradation mechanisms for InAs quantum dot lasers grown on silicon substrates emitting at 1.3-μm are investigated. The rate of degradation is observed to increase for devices operated at higher carrier densities and is therefore dependent on gain requirement, or cavity length. While carrier localisation in quantum dots minimises degradation, an increase in the number of defects in the early stages of ageing can increase the internal optical-loss which, can initiate rapid degradation of laser performance due to the rise in threshold carrier density. Population of the 2-D states is considered the major factor for determining the rate of degradation, which can be significant for lasers requiring high threshold carrier densities. This is demonstrated by operating lasers of different cavity lengths with a constant current and measuring the change in threshold current at regular intervals. A segmented-contact device, which can be used to measure the modal absorption and also operate as a laser, is used to determine how the internal optical-loss changes in the early stages of degradation. Structures grown on silicon show an increase in internal optical-loss whereas the same structure grown on GaAs show no signs of increase in internal optical-loss when operated under the same conditions

Low-Noise GaAs Quantum Dots for Quantum Photonics

Quantum dots are both excellent single-photon sources and hosts for single spins. This combination enables the deterministic generation of Raman-photons -- bandwidth-matched to an atomic quantum-memory -- and the generation of photon cluster states, a resource in quantum communication and measurement-based quantum computing. GaAs quantum dots in AlGaAs can be matched in frequency to a rubidium-based photon memory, and have potentially improved electron spin coherence compared to the widely used InGaAs quantum dots. However, their charge stability and optical linewidths are typically much worse than for their InGaAs counterparts. Here, we embed GaAs quantum dots into an $n$-$i$-$p$-diode specially designed for low-temperature operation. We demonstrate ultra-low noise behaviour: charge control via Coulomb blockade, close-to lifetime-limited linewidths, and no blinking. We observe high-fidelity optical electron-spin initialisation and long electron-spin lifetimes for these quantum dots. Our work establishes a materials platform for low-noise quantum photonics close to the red part of the spectrum.Comment: (19 pages, 12 figures, 1 table

Optically driving the radiative Auger transition

In a radiative Auger process, optical decay is accompanied by simultaneous excitation of other carriers. The radiative Auger process gives rise to weak red-shifted satellite peaks in the optical emission spectrum. These satellite peaks have been observed over a large spectral range: in the X-ray emission of atoms; close to visible frequencies on donors in semiconductors and quantum emitters; and at infrared frequencies as shake-up lines in two-dimensional systems. So far, all the work on the radiative Auger process has focussed on detecting the spontaneous emission. However, the fact that the radiative Auger process leads to photon emission suggests that the transition can also be optically excited. In such an inverted radiative Auger process, excitation would correspond to simultaneous photon absorption and electronic de-excitation. Here, we demonstrate optical driving of the radiative Auger transition on a trion in a semiconductor quantum dot. The radiative Auger and the fundamental transition together form a $\Lambda$-system. On driving both transitions of this $\Lambda$-system simultaneously, we observe a reduction of the fluorescence signal by up to $70\%$. Our results demonstrate a type of optically addressable transition connecting few-body Coulomb interactions to quantum optics. The results open up the possibility of carrying out THz spectroscopy on single quantum emitters with all the benefits of optics: coherent laser sources, efficient and fast single-photon detectors. In analogy to optical control of an electron spin, the $\Lambda$-system between the radiative Auger and the fundamental transitions allows optical control of the emitters' orbital degree of freedom.Comment: 8 pages, 6 figure

Charge Tunable GaAs Quantum Dots in a Photonic n-i-p Diode

In this submission, we discuss the growth of charge-controllable GaAs quantum dots embedded in an n-i-p diode structure, from the perspective of a molecular beam epitaxy grower. The QDs show no blinking and narrow linewidths. We show that the parameters used led to a bimodal growth mode of QDs resulting from low arsenic surface coverage. We identify one of the modes as that showing good properties found in previous work. As the morphology of the fabricated QDs does not hint at outstanding properties, we attribute the good performance of this sample to the low impurity levels in the matrix material and the ability of n- and p-doped contact regions to stabilize the charge state. We present the challenges met in characterizing the sample with ensemble photoluminescence spectroscopy caused by the photonic structure used. We show two straightforward methods to overcome this hurdle and gain insight into QD emission properties

Novel Allosteric Mechanism of Dual p53/MDM2 and p53/MDM4 Inhibition by a Small Molecule

Restoration of the p53 tumor suppressor for personalised cancer therapy is a promising treatment strategy. However, several high-affinity MDM2 inhibitors have shown substantial side effects in clinical trials. Thus, elucidation of the molecular mechanisms of action of p53 reactivating molecules with alternative functional principle is of the utmost importance. Here, we report a discovery of a novel allosteric mechanism of p53 reactivation through targeting the p53 N-terminus which promotes inhibition of both p53/MDM2 (murine double minute 2) and p53/MDM4 interactions. Using biochemical assays and molecular docking, we identified the binding site of two p53 reactivating molecules, RITA (reactivation of p53 and induction of tumor cell apoptosis) and protoporphyrin IX (PpIX). Ion mobility-mass spectrometry revealed that the binding of RITA to serine 33 and serine 37 is responsible for inducing the allosteric shift in p53, which shields the MDM2 binding residues of p53 and prevents its interactions with MDM2 and MDM4. Our results point to an alternative mechanism of blocking p53 interaction with MDM2 and MDM4 and may pave the way for the development of novel allosteric inhibitors of p53/MDM2 and p53/MDM4 interactions

Exploiting phonon and coulomb interactions in semiconductor quantum dots

Semiconductor quantum dots have been investigated in many different aspects, from fundamental semiconductor physics to advanced quantum technologies. After many years of growth improvements, the quantum dot emits single photons of high purity and high indistinguishability. The noise in the semiconductor is reduced to a level where it starts to be negligible (compared to other measurement errors). Recently advances also have been made in controlling a single electron(hole)-spin confined by the quantum dot. This makes the quantum dot a perfect candidate for any application involving single photons and also a spin, for example, boson sampling and cluster state generation, respectively. The quantum dot can also be used to study its coupling to additional degrees of freedom, such as for example the surrounding nuclear spins or coupling to an optical cavity. In this thesis, two individual studies are presented. First, the interaction of the quantum dot with the mechanical surrounding, i.e. phonons, and second, radiative Auger processes due to Coulomb interactions. The quantum dot naturally couples to its mechanical environment by deformation-potential coupling. The interaction is exploited by engineering the mechanical environment (density of states) by patterning a mechanical resonator (mechanical cavity). The main focus lies on reaching gigahertz mechanical frequencies, the so-called resolved-sideband regime. However, this is not trivial for two reasons. First, fabricating a mechanical resonator at such high frequencies is challenging due the small size. Second, measuring such a fast modulation of the quantum dot requires a special measurement parameter set. Nonetheless, the coupling to mechanical resonators from a few megahertz to more than a gigahertz mechanical frequency is shown. Furthermore, an in-depth study of the exciton-phonon interaction is presented which includes a semi-classical master-equation description of the coupled system as well as the observation of acoustic sideband emission. The current limit, for applications such as optomechanical cooling, is the coupling rate between the two systems. However, such experiments are within reach with a five- to ten-fold increase in the coupling rate. The quantum dot itself presents a coupled system if charged with an additional electron (or hole). Then, in the excited state, three carriers (one hole and two electrons) are tightly confined inside the dot which are coupled to each other via Coulomb interactions. The effect of this coupling is studied which gives rise to the so-called radiative Auger process. During the decay of the trion, one of the carriers (the Auger electron) is promoted to a higher energy state inside the quantum dot (p- and d-shell) and the emitted photon is correspondingly red-shifted. In more detail, it is found that the wavefunction of the trion is composed of admixtures of higher shells and the emitted photon projects the state of the remaining electron to the corresponding shell. Furthermore, the radiative Auger process gives rise to an optical transition which can be addressed with a laser. In a two-laser experiment (Λ-configuration), the radiative Auger transition is optically driven. This leads to a coherent superposition of the auger carrier being in two different quantum dot shells. These measurements pose a first step toward coherent control of the orbital state of the Auger carrier

Quantum interference of identical photons from remote GaAs quantum dots

Photonic quantum technology provides a viable route to quantum communication1,2, quantum simulation3 and quantum information processing4. Recent progress has seen the realization of boson sampling using 20 single photons3 and quantum key distribution over hundreds of kilometres2. Scaling the complexity requires architectures containing multiple photon sources, photon counters and a large number of indistinguishable single photons. Semiconductor quantum dots are bright and fast sources of coherent single photons5,6,7,8,9. For applications, a roadblock is the poor quantum coherence on interfering single photons created by independent quantum dots10,11. Here we demonstrate two-photon interference with near-unity visibility (93.0 ± 0.8)% using photons from two completely separate GaAs quantum dots. The experiment retains all the emission into the zero phonon line—only the weak phonon sideband is rejected; temporal post-selection is not employed. By exploiting quantum interference, we demonstrate a photonic controlled-not circuit and an entanglement with fidelity of (85.0 ± 1.0)% between photons of different origins. The two-photon interference visibility is high enough that the entanglement fidelity is well above the classical threshold. The high mutual coherence of the photons stems from high-quality materials, diode structure and relatively large quantum dot size. Our results establish a platform—GaAs quantum dots—for creating coherent single photons in a scalable way