In vitro and in vivo studies on protease-activated receptor-2

Protease-activated receptor-2 (PAR2) is a G protein coupled receptor (GPCR) that is activated by proteolytic cleavage of its amino terminal domain by trypsin-like serine proteases. Cleavage of this receptor exposes a neoepitope, termed the tethered ligand (TL), which binds intramolecularly within the receptor to stimulate signal transduction via coupled G proteins. PAR2-mediated signal transduction is also experimentally stimulated by hexapeptides (agonist peptides; APs) that are homologous to the TL sequence. Due to the irreversible nature of PAR2 proteolysis, downstream signal transduction is tightly regulated. Following activation, PAR2 is rapidly uncoupled from downstream signalling by the post-translational modifications phosphorylation and ubiquination which facilitate interactions with â- arrestin. This scaffolding protein couples PAR2 to the internalisation machinery initiating its desensitisation and trafficking through the early and late endosomes followed by receptor degradation. PAR2 is widely expressed in mammalian tissues with key roles for this receptor in cardiovascular, respiratory, nervous and musculoskeletal systems. This receptor has also been linked to pathological states with aberrant expression and signalling noted in several cancers. In prostate cancer, PAR2 signalling induces migration and proliferation of tumour derived cell lines, while elevated receptor expression has been noted in malignant tissues. Importantly, a role for this receptor has also been suggested in prostate cancer bone metastasis as coexpression of PAR2 and a proteolytic activator has been demonstrated by immunohistochemical analysis. Based on these data, the primary focus of this project has been on two aspects of PAR2 biology. The first is characterisation of cellular mechanisms that regulate PAR2 signalling and trafficking. The second aspect is the role of this receptor in prostate cancer bone metastasis. In addition, to permit these studies, it was first necessary to evaluate the specificity of the commercially available anti-PAR2 antibodies SAM11, C17, N19 and H99. The evaluation of the four commercially available antibodies was assessed using four techniques: immunoprecipitation; Western blot analysis; immunofluorescence; and flow cytometry. These approaches demonstrated that three of the antibodies efficiently detect ectopically expressed PAR2 by each of these techniques. A significant finding from this study was that N19 was the only antibody able to specifically detect N-glycosylated endogenous PAR2 by Western blot analysis. This analysis was performed on lysates from prostate cancer derived cell lines and tissue derived from wildtype and PAR2 knockout mice. Importantly, further evaluation demonstrated that this antibody also efficiently detects endogenous PAR2 at the cell surface by flow cytometry. The anti-PAR2 antibody N19 was used to explore the in vitro role of palmitoylation, the post-translational addition of palmitate, in PAR2 signalling, trafficking, cell surface expression and desensitization. Significantly, use of the palmitoylation inhibitor 2-bromopalmitate indicated that palmitate addition is important in trafficking of PAR2 endogenously expressed by prostate cancer cell lines. This was supported by palmitate labelling experiments using two approaches which showed that PAR2 stably expressed by CHO cells is palmitoylated and that palmitoylation occurs on cysteine 361. Another key finding from this study is that palmitoylation is required for optimal PAR2 signalling as Ca2+ flux assays indicated that in response to trypsin agonism, palmitoylation deficient PAR2 is ~9 fold less potent than wildtype receptor with a reduction of about 33% in the maximum signal induced via the mutant receptor. Confocal microscopy, flow cytometry and cell surface biotinylation analyses demonstrated that palmitoylation is required for efficient cell surface expression of PAR2. Importantly, this study also identified that palmitoylation of this receptor within the Golgi apparatus is required for efficient agonist-induced rab11amediated trafficking of PAR2 to the cell surface. Interestingly, palmitoylation is also required for receptor desensitization, as agonist-induced â-arrestin recruitment and receptor degradation were markedly reduced in CHO-PAR2-C361A cells compared with CHO-PAR2 cells. Collectively, these data provide new insights on the life cycle of PAR2 and demonstrate that palmitoylation is critical for efficient signalling, trafficking, cell surface localization and degradation of this receptor. This project also evaluated PAR2 residues involved in ligand docking. Although the extracellular loop (ECL)2 of PAR2 is known to be required for agonist-induced signal transduction, the binding pocket for receptor agonists remains to be determined. In silico homology modelling, based on a crystal structure for the prototypical GPCR rhodopsin, and ligand docking were performed to identify PAR2 transmembrane (TM) amino acids potentially involved in agonist binding. These methods identified 12 candidate residues that were mutated to examine the binding site of the PAR2 TL, revealed by trypsin cleavage, as well as of the soluble ligands 2f-LIGRLO-NH2 and GB110, which are both structurally based on the AP SLIGRLNH2. Ligand binding was evaluated from the impact of the mutated residues on PAR2-mediated calcium mobilisation. An important finding from these experiments was that mutation of residues Y156 and Y326 significantly reduced 2f-LIGRLO-NH2 and GB110 agonist activity. L307 was also important for GB110 activity. Intriguingly, mutation of PAR2 residues did not alter trypsin-induced signalling to the same extent as for the soluble agonists. The reason for this difference remains to be further examined by in silico and in vitro experimentation and, potentially, crystal structure studies. However, these findings identified the importance of TM domains in PAR2 ligand docking and will enhance the design of both PAR2 agonists and potentially agents to inhibit signalling (antagonists). The potential importance of PAR2 in prostate cancer bone metastasis was examined using a mouse model. In patients, prostate cancer bone metastases cause bone growth by disrupting bone homeostasis. In an attempt to mimic prostate cancer growth in bone, PAR2 responsive 22Rv1 prostate cancer cells, which form mixed osteoblastic and osteolytic lesions, were injected into the proximal aspect of mouse tibiae. A role for PAR2 was assessed by treating these mice with the recently developed PAR2 antagonist GB88. As controls, animals bearing intra-tibial tumours were also treated with vehicle (olive oil) or the prostate cancer chemotherapeutic docetaxel. The effect of these treatments on bone was examined radiographically and by micro-CT. Consistent with previous studies, 22Rv1 tumours caused osteoblastic periosteal spicule formation and concurrent osteolytic bone loss. Significantly, blockade of PAR2 signalling reduced the osteoblastic and osteolytic phenotype of 22Rv1 tumours in bone. No bone defects were detected in mice treated with docetaxel. These qualitative data will be followed in the future by quantitative micro-CT analysis as well as histology and histomorphometry analysis of already collected tissues. Nonetheless, these preliminary experiments highlight a potential role for PAR2 in prostate cancer growth in bone. In summary, in vitro studies have defined mechanisms regulating PAR2 activation, downstream signalling and trafficking and in vivo studies point to a potential role for this receptor in prostate cancer bone metastasis. The outcomes of this project are that a greater understanding of the biology of PAR2 may lead to the development of strategies to modulate the function of this receptor in disease.

'It had to be my choice' Indigenous smoking cessation and negotiations of risk, resistance and resilience

While Australia is considered a world leader in tobacco control, smoking rates within the Aboriginal and Torres Strait Islander population have not declined at the same rate. This failure highlights an obvious shortcoming of mainstream anti-smoking efforts to effectively understand and engage with the socio-cultural context of Indigenous smoking and smoking cessation experiences. The purpose of this article is to explore the narrative accounts of 20 Indigenous ex-smokers within an urban community and determine the motivators and enablers for successful smoking cessation. Our findings indicated that health risk narratives and the associated social stigma produced through anti-smoking campaigns formed part of a broader apparatus of oppression among Indigenous people, often inspiring resistance and resentment rather than compliance. Instead, a significant life event and supportive relationships were the most useful predictors of successful smoking cessation acting as both a motivator and enabler to behavioural change. Indigenous smoking cessation narratives most commonly involved changing and reordering a person’s life and identity and autonomy over this process was the critical building block to reclaiming control over nicotine addiction. Most promisingly, at an individual level, we found the important role that individual health professionals played in encouraging and supporting Indigenous smoking cessation through positive rather than punitive interactions. More broadly, our findings highlighted the central importance of resilience, empowerment, and trust within health promotion practice.

Digital futures for cultural and media studies

An ambitious rendering of the digital future from a pioneer of media and cultural studies, a wise and witty take on a changing field, and our orientation to it Investigates the uses of multimedia by creative and productive citizen-consumers to provide new theories of communication that accommodate social media, participatory action, and user-creativity Leads the way for new interdisciplinary engagement with systems thinking, complexity and evolutionary sciences, and the convergence of cultural and economic values Analyzes the historical uses of multimedia from print, through broadcasting to the internet Combines conceptual innovation with historical erudition to present a high-level synthesis of ideas and detailed analysis of emergent forms and practices Features an international focus and global reach to provide a basis for students and researchers seeking broader perspectives

A simulation-based training system for surgical wound debridement

A simulation-based training system for surgical wound debridement was developed and comprises a multimedia introduction, a surgical simulator (tutorial component), and an assessment component. The simulator includes two PCs, a haptic device, and mirrored display. Debridement is performed on a virtual leg model with a shallow laceration wound superimposed. Trainees are instructed to remove debris with forceps, scrub with a brush, and rinse with saline solution to maintain sterility. Research and development issues currently under investigation include tissue deformation models using mass-spring system and finite element methods; tissue cutting using a high-resolution volumetric mesh and dynamic topology; and accurate collision detection, cutting, and soft-body haptic rendering for two devices within the same haptic space.

Architecture post mortem : the diastolic architecture of decline, dystopia, and death

Architecture Post Mortem surveys architecture’s encounter with death, decline, and ruination following late capitalism. As the world moves closer to an economic abyss that many perceive to be the death of capital, contraction and crisis are no longer mere phases of normal market fluctuations, but rather the irruption of the unconscious of ideology itself. Post mortem is that historical moment wherein architecture’s symbolic contract with capital is put on stage, naked to all. Architecture is not irrelevant to fiscal and political contagion as is commonly believed; it is the victim and penetrating analytical agent of the current crisis. As the very apparatus for modernity’s guilt and unfulfilled drives-modernity’s debt-architecture is that ideological element that functions as a master signifier of its own destruction, ordering all other signifiers and modes of signification beneath it. It is under these conditions that architecture theory has retreated to an “Alamo” of history, a final desert outpost where history has been asked to transcend itself. For architecture’s hoped-for utopia always involves an apocalypse. This timely collection of essays reformulates architecture’s relation to modernity via the operational death-drive: architecture is but a passage between life and death. This collection includes essays by Kazi K. Ashraf, David Bertolini, Simone Brott, Peggy Deamer, Didem Ekici, Paul Emmons, Donald Kunze, Todd McGowan, Gevork Hartoonian, Nadir Lahiji, Erika Naginski, and Dennis Maher. Contents: Introduction: ‘the way things are’, Donald Kunze; Driven into the public: the psychic constitution of space, Todd McGowan; Dead or alive in Joburg, Simone Brott; Building in-between the two deaths: a post mortem manifesto, Nadir Lahiji; Kant, Sade, ethics and architecture, David Bertolini; Post mortem: building deconstruction, Kazi K. Ashraf; The slow-fast architecture of love in the ruins, Donald Kunze; Progress: re-building the ruins of architecture, Gevork Hartoonian; Adrian Stokes: surface suicide, Peggy Deamer; A window to the soul: depth in the early modern section drawing, Paul Emmons; Preliminary thoughts on Piranesi and Vico, Erika Naginski; architectural asceticism and austerity, Didem Ekici; 900 miles to Paradise, and other afterlives of architecture, Dennis Maher; Index.

Balancing values and imperatives : a study of nursing service in an ICU

There has been significant attention from the managers and purchasers of health services regarding the economic advantages that result from changes to the patterns of health care delivery in the acute hospital setting. The impact of these changes, whilst often rendering advantage at the economic management level of health care, can have different consequences for the people who deliver and the people who receive health service. This paper reports on a study that was conducted with a group of nurses to investigate the practice milieu of a critical care unit in the context of changes to health service management. Interpretive methods were used to capture the perspective of the nurses and the way they interpret the multiple factors that influence their practice and their practice environment. The findings indicate that the nurses in the study setting interpret these factors according to the influences they have on the structure, the geography and the value of their work. Explication of these findings provides a research base to inform recommendations relating to improving the practice milieu of the critical care environment.

The use of CCTV to police public spaces : a case of big brother or big friend?

This paper considers the role of CCTV (closed circuit television) in the surveillance, policing and control of public space in urban and rural locations, specifically in relation to the use of public space by young people. The use of CCTV technology in public spaces is now an established and largely uncontested feature of everyday life in a number of countries and the assertion that they are essentially there for the protection of law abiding and consuming citizens has broadly gone unchallenged. With little or no debate in the U.K. to critique the claims made by the burgeoning security industry that CCTV protects people in the form of a ‘Big Friend’, the state at both central and local levels has endorsed the installation of CCTV apparatus across the nation. Some areas assert in their promotional material that the centre of the shopping and leisure zone is fully surveilled by cameras in order to reassure visitors that their personal safety is a matter of civic concern, with even small towns and villages expending monies on sophisticated and expensive to maintain camera systems. It is within a context of monitoring, recording and control procedures that young people’s use of public space is constructed as a threat to social order, in need of surveillance and exclusion which forms a major and contemporary feature in shaping thinking about urban and rural working class young people in the U.K. As Loader (1996) notes, young people’s claims on public space rarely gain legitimacy if ‘colliding’ with those of local residents, and Davis (1990) describes the increasing ‘militarization and destruction of public space’, while Jacobs (1965) asserts that full participation in the ‘daily life of urban streets’ is essential to the development of young people and beneficial for all who live in an area. This paper challenges the uncritical acceptance of widespread use of CCTV and identifies its oppressive and malevolent potential in forming a ‘surveillance gaze’ over young people (adapting Foucault’s ‘clinical gaze’c. 1973) which can jeopardise mental health and well being in coping with the ‘metropolis’, after Simmel, (1964).

Nucleation experiments in a microgravity environment

A simple experimental apparatus is described in which a wide variety of vapor phase nucleation studies of refractory materials could be performed aboard NASA's KC-135 Research Aircraft. The chief advantage of a microgravity environment for these studies is the expected absence of thermally driven convective motions in the gas. The absence of convection leads to much more accurate knowledge of both the temperature distribution in the system and the time evolution of the refractory vapor concentration as a function of distance from the crucible.The evolution of the apparatus will be described as more experience is gained with the microgravity environment. Such experiments will be used to prepare for similar ones carried out aboard either the shuttle or Space Station where considerably longer duration experiments are possible.

Seriously playful : the uses of networked spoof videos in China

This thesis examines online spoof videos in China. It shows the relationship between user-created content and change and how such videos are impacting on social memory. In the West, we are witnessing two outstanding trends in media. On the one hand, media are turning more "demotic" (Turner, 2006) and "participatory" (Jenkins, 2006), whereby lay audiences use popular media for identity formation, representation and association, reconfiguring the media and cultural landscape, and rendering invalid the old paradigm based on the dichotomy of audience and author, creator and consumer, expert and amateur. On the other hand, in both mainstream media and user-creation online there is a trend towards "silly citizenship", with comedy, send-ups and spoofs that used to reside in the margin propelled to the central stage in both pleasure and politics (Hartley, 2010), as is shown in the rising popularity of the Daily Show, Colbert Report, and spoof videos in elections ,e.g. the 2008 presidential election in US (Gray, Jones, & Thompson, 2009; Tryon, 2008). User generated content—and spoof subcultures—is now much a debated phenomenon in China. However, with different political (one party rule and censorship) and cultural (media regarded mainly as instrument for education and social stabilization instead of a critical fourth estate) configurations, will the social and cultural impacts of the two trends in the West be as the same in China? If not, what will be the specificities in the China context? The project starts with a historical review of popular culture and user-created content in China, before turning to spoof videos and looking at how they are produced and shared, travel and diffused on the Internet, and how the communities and sub-cultures forming and emerging around spoof videos are changing the overall cultural landscape in China. By acting as a participant observer in online video sharing sites and conducting face-to face as well as online interviews, I identify lead users and creators of spoof videos and the social networks emerging around them. I call these lead users "skill hubs" and their networks "liquid communities", foregrounding the fact that their appeal doesn’t come from their amicable personality, but rather from their creative skills; and that the networks surrounding them are in a permanent flux, with members coming and going as they see fit. I argue that the "liquidness" (Bauman, 2000) of these communities is what makes them constantly creative and appealing. Textual analysis of online videos, their comments and derivatives are conducted to tease out the uses that that can be made of spoof videos, namely as phatic communication, as alternative memory and as political engagement. Through these analyses I show that spoof videos constitute not only a space where young generations can engage with each other, communicate their anger and dissatisfaction, fun and hope, and where they participate in socio-cultural and political debates, but also create a space where they can experiment with their new skills, new ideas, and new citizenship. The rise of spoof videos heralds the beginning of a trend in popular culture in contemporary China towards the "canonization of the jester" and the dethroning of the establishment. I also argue that a historical perspective is needed to understand the current surge of use creativity and user activism in China, and that many forms of popular media we experience today have their antecedents in various stages of Chinese history. The entrenched "control-resistance" binary is inadequate in interpreting the rich, flux and multilayered Internet space in China.

Ventilation distribution in rats : part 2 – a comparison of electrical impedance tomography and hyperpolarised helium magnetic resonance imaging

Background: Hyperpolarised helium MRI (He3 MRI) is a new technique that enables imaging of the air distribution within the lungs. This allows accurate determination of the ventilation distribution in vivo. The technique has the disadvantages of requiring an expensive helium isotope, complex apparatus and moving the patient to a compatible MRI scanner. Electrical impedance tomography (EIT) a non-invasive bedside technique that allows constant monitoring of lung impedance, which is dependent on changes in air space capacity in the lung. We have used He3MRI measurements of ventilation distribution as the gold standard for assessment of EIT. Methods: Seven rats were ventilated in supine, prone, left and right lateral position with 70% helium/30% oxygen for EIT measurements and pure helium for He3 MRI. The same ventilator and settings were used for both measurements. Image dimensions, geometric centre and global in homogeneity index were calculated. Results: EIT images were smaller and of lower resolution and contained less anatomical detail than those from He3 MRI. However, both methods could measure positional induced changes in lung ventilation, as assessed by the geometric centre. The global in homogeneity index were comparable between the techniques. Conclusion: EIT is a suitable technique for monitoring ventilation distribution and inhomgeneity as assessed by comparison with He3 MRI.

Embedding diversity in a university justice curriculum : an exploration of the issues

This paper reports on an action-learning project conducted within the first year criminal justice curriculum in an Australian university. The project was initiated after an audit of first year units and student feedback revealed that there were gaps in the curriculum that possibly were disadvantaging certain groups of students, including mature, international, queer and disabled students, rendering them invisible. Official (university controlled student surveys and other feedback mechanisms) and anecdotal feedback found that at least some students in these groups felt disenfranchised; that is, unable to relate to either the subject mater, other students, or the university setting itself. As a school in which social justice provides the context for learning about criminal justice, first year subject coordinators as a group came to recognise the need for embedding diversity in the curriculum.

The new normativity : the innovation imperative

At a quite fundamental level, the very way in which Public Service Broadcasting (PSB) may envisage its future usually captured in the semantic shift from PSB to Public Service Media (PSM) is at stake when considering the recent history of public value discourse and the public value test. The core Reithian PSB idea assumed that public value would be created through the application of core principles of universality of availability and appeal, provision for minorities, education of the public, distance from vested interests, quality programming standards, program maker independence, and fostering of national culture and the public sphere. On the other hand, the philosophical import of the public value test is that potentially any excursion into the provision of new media services needs to be justified ex ante. In this era of New Public Management, greater transparency and accountability, and the proposition that resources for public value deliverables be contestable and not sequestered in public sector institutions, what might be the new Archimedean point around which a contemporised normativity for PSM be built? This paper will argue for the innovation imperative as an organising principle for contemporary PSM. This may appear counterintuitive, as it is precisely PSB’s predilection for innovating in new media services (in online, mobile, and social media) that has produced the constraining apparatus of the ex ante/public value/Drei-Stufen-Test in Europe, based on principles of competitive neutrality and transparency in the application of public funds for defined and limited public benefit. However, I argue that a commitment to innovation can define as complementary to, rather than as competitive ‘crowding out’, the new products and services that PSM can, and should, be delivering into a post-scarcity, superabundant all-media marketplace. The evidence presented in this paper for this argument is derived mostly from analysis of PSM in the Australian media ecology. While no PSB outside Europe is subject to a formal public value test, the crowding out arguments are certainly run in Australia, particularly by powerful commercial interests for whom free news is a threat to monetising quality news journalism. Take right wing opinion leader, herself a former ABC Board member, Judith Sloan: ‘… the recent expansive nature of the ABC – all those television stations, radio stations and online offerings – is actually squeezing activity that would otherwise be undertaken by the private sector. From partly correcting market failure, the ABC is now causing it. We are now dealing with a case of unfair competition and wasted taxpayer funds’ (The Drum, 1 August http://www.abc.net.au/unleashed/2818220.html). But I argue that the crowding out argument is difficult to sustain in Australia because of the PSB’s non-dominant position and the fact that much of innovation generated by the two PSBs, the ABC and the SBS, has not been imitated by or competed for by the commercials. The paper will bring cases forward, such as SBS’ Go Back to Where you Came From (2011) as an example of product innovation, and a case study of process and organisational innovation which also has resulted in specific product and service innovation – the ABC’s Innovation Unit. In summary, at least some of the old Reithian dicta, along with spectrum scarcity and market failure arguments, have faded or are fading. Contemporary PSM need to justify their role in the system, and to society, in terms of innovation.

Groundwater visualisation system (GVS) : a software framework for integrated display and interrogation of conceptual hydrogeological models, data and time-series animation

Management of groundwater systems requires realistic conceptual hydrogeological models as a framework for numerical simulation modelling, but also for system understanding and communicating this to stakeholders and the broader community. To help overcome these challenges we developed GVS (Groundwater Visualisation System), a stand-alone desktop software package that uses interactive 3D visualisation and animation techniques. The goal was a user-friendly groundwater management tool that could support a range of existing real-world and pre-processed data, both surface and subsurface, including geology and various types of temporal hydrological information. GVS allows these data to be integrated into a single conceptual hydrogeological model. In addition, 3D geological models produced externally using other software packages, can readily be imported into GVS models, as can outputs of simulations (e.g. piezometric surfaces) produced by software such as MODFLOW or FEFLOW. Boreholes can be integrated, showing any down-hole data and properties, including screen information, intersected geology, water level data and water chemistry. Animation is used to display spatial and temporal changes, with time-series data such as rainfall, standing water levels and electrical conductivity, displaying dynamic processes. Time and space variations can be presented using a range of contouring and colour mapping techniques, in addition to interactive plots of time-series parameters. Other types of data, for example, demographics and cultural information, can also be readily incorporated. The GVS software can execute on a standard Windows or Linux-based PC with a minimum of 2 GB RAM, and the model output is easy and inexpensive to distribute, by download or via USB/DVD/CD. Example models are described here for three groundwater systems in Queensland, northeastern Australia: two unconfined alluvial groundwater systems with intensive irrigation, the Lockyer Valley and the upper Condamine Valley, and the Surat Basin, a large sedimentary basin of confined artesian aquifers. This latter example required more detail in the hydrostratigraphy, correlation of formations with drillholes and visualisation of simulation piezometric surfaces. Both alluvial system GVS models were developed during drought conditions to support government strategies to implement groundwater management. The Surat Basin model was industry sponsored research, for coal seam gas groundwater management and community information and consultation. The “virtual” groundwater systems in these 3D GVS models can be interactively interrogated by standard functions, plus production of 2D cross-sections, data selection from the 3D scene, rear end database and plot displays. A unique feature is that GVS allows investigation of time-series data across different display modes, both 2D and 3D. GVS has been used successfully as a tool to enhance community/stakeholder understanding and knowledge of groundwater systems and is of value for training and educational purposes. Projects completed confirm that GVS provides a powerful support to management and decision making, and as a tool for interpretation of groundwater system hydrological processes. A highly effective visualisation output is the production of short videos (e.g. 2–5 min) based on sequences of camera ‘fly-throughs’ and screen images. Further work involves developing support for multi-screen displays and touch-screen technologies, distributed rendering, gestural interaction systems. To highlight the visualisation and animation capability of the GVS software, links to related multimedia hosted online sites are included in the references.

Thermal-elastic stresses and the criticality of the continental crust

Heating or cooling can lead to high stresses in rocks due to the different thermal-elastic properties of minerals. In the upper 4 km of the crust, such internal stresses might cause fracturing. Yet it is unclear if thermal elasticity contributes significantly to critical stresses and failure deeper in Earth's continental crust, where ductile creep causes stress relaxation. We combined a heating experiment conducted in a Synchrotron microtomograph (Advanced Photon Source, USA) with numerical simulations to calculate the grain-scale stress field in granite generated by slow burial. We find that deviatoric stresses >100 MPa can be stored during burial, with relaxation times from 100's to 1000's ka, even in the ductile crust. Hence, grain-scale thermal-elastic stresses may serve as nuclei for instabilities, thus rendering the continental crust close to criticality.

ATM is required for the cellular response to thymidine induced replication fork stress

Genetically distinct checkpoints, activated as a consequence of either DNA replication arrest or ionizing radiation-induced DNA damage, integrate DNA repair responses into the cell cycle programme. The ataxia-telangiectasia mutated (ATM) protein kinase blocks cell cycle progression in response to DNA double strand breaks, whereas the related ATR is important in maintaining the integrity of the DNA replication apparatus. Here, we show that thymidine, which slows the progression of replication forks by depleting cellular pools of dCTP, induces a novel DNA damage response that, uniquely, depends on both ATM and ATR. Thymidine induces ATM-mediated phosphorylation of Chk2 and NBS1 and an ATM-independent phosphorylation of Chk1 and SMC1. AT cells exposed to thymidine showed decreased viability and failed to induce homologous recombination repair (HRR). Taken together, our results implicate ATM in the HRR-mediated rescue of replication forks impaired by thymidine treatment.