SemGen: a tool for semantics-based annotation and composition of biosimulation models.

SUMMARY: As the number and complexity of biosimulation models grows, so do demands for tools that can help users understand models and compose more comprehensive and accurate systems from existing models. SemGen is a tool for semantics-based annotation and composition of biosimulation models designed to address this demand. A key SemGen capability is to decompose and then integrate models across existing model exchange formats including SBML and CellML. To support this capability, we use semantic annotations to explicitly capture the underlying biological and physical meanings of the entities and processes that are modeled. SemGen leverages annotations to expose a model's biological and computational architecture and to help automate model composition. AVAILABILITY AND IMPLEMENTATION: SemGen is freely available at https://github.com/SemBioProcess/SemGen. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Virtual Reality-based Methods for Training Novice Electrophysiology Trainees-A Pilot Study.

Developing an accurate and detailed 3-dimensional (3D) mental model of cardiac anatomy is critical for electrophysiology (EP) trainees. Due to its immersive nature, virtual reality (VR) may provide a better learning environment than traditional teaching methods for assimilating 3D cardiac anatomy. The purpose of this pilot study was to evaluate the technical feasibility of an interactive, remote VR-based method for teaching cardiac anatomy to novice EP trainees. We created a shared, remote VR environment that allows the shared viewing of high-resolution 3D cardiac models. Eighteen trainees accepted for pediatric and adult EP fellowships were recruited. We performed a cohort study comparing the traditional teaching methods with the VR learning environment. Participants completed a demographic questionnaire and a satisfaction survey. The adult EP trainees were given a multiple-choice pre- and post-test exam to assess their anatomical knowledge. Both the adult and pediatric EP trainee cohorts rated the VR experience positively and preferred the VR environment to the more traditional teaching method. All the participants expressed interest in incorporating the VR learning environment into the EP fellowship curriculum. The usability of the system was relatively low, with approximately one-third of participants rating the system as hard to use. The impact of the VR session on exam performance was mixed among the adult cohort. We demonstrated the feasibility of gathering geographically dispersed EP fellows in training with a shared VR-based environment to teach cardiac anatomy. Although we were not able to demonstrate a learning benefit over the traditional lecture format in the adult cohort, the training environment was favorably received by all the participants.

Visualization of Replisome Encounters with an Antigen Tagged Blocking Lesion.

Considerable insight is present into the cellular response to double strand breaks (DSBs), induced by nucleases, radiation, and other DNA breakers. In part, this reflects the availability of methods for the identification of break sites, and characterization of factors recruited to DSBs at those sequences. However, DSBs also appear as intermediates during the processing of DNA adducts formed by compounds that do not directly cause breaks, and do not react at specific sequence sites. Consequently, for most of these agents, technologies that permit the analysis of binding interactions with response factors and repair proteins are unknown. For example, DNA interstrand crosslinks (ICLs) can provoke breaks following replication fork encounters. Although formed by drugs widely used as cancer chemotherapeutics, there has been no methodology for monitoring their interactions with replication proteins. Here, we describe our strategy for following the cellular response to fork collisions with these challenging adducts. We linked a steroid antigen to psoralen, which forms photoactivation dependent ICLs in nuclei of living cells. The ICLs were visualized by immunofluorescence against the antigen tag. The tag can also be a partner in the Proximity Ligation Assay (PLA) which reports the close association of two antigens. The PLA was exploited to distinguish proteins that were closely associated with the tagged ICLs from those that were not. It was possible to define replisome proteins that were retained after encounters with ICLs and identify others that were lost. This approach is applicable to any structure or DNA adduct that can be detected immunologically.

Replication of the Mammalian Genome by Replisomes Specific for Euchromatin and Heterochromatin.

Replisomes follow a schedule in which replication of DNA in euchromatin is early in S phase while sequences in heterochromatin replicate late. Impediments to DNA replication, referred to as replication stress, can stall replication forks triggering activation of the ATR kinase and downstream pathways. While there is substantial literature on the local consequences of replisome stalling-double strand breaks, reversed forks, or genomic rearrangements-there is limited understanding of the determinants of replisome stalling vs. continued progression. Although many proteins are recruited to stalled replisomes, current models assume a single species of "stressed" replisome, independent of genomic location. Here we describe our approach to visualizing replication fork encounters with the potent block imposed by a DNA interstrand crosslink (ICL) and our discovery of an unexpected pathway of replication restart (traverse) past an intact ICL. Additionally, we found two biochemically distinct replisomes distinguished by activity in different stages of S phase and chromatin environment. Each contains different proteins that contribute to ICL traverse.

Visualizing replication fork encounters with DNA interstrand crosslinks.

Replication forks encounter numerous challenges as they move through eu- and hetero-chromatin during S phase in mammalian cells. These include a variety of impediments to the unwinding of DNA by the replicative helicase such as alternate DNA structures, transcription complexes and R-loops, DNA-protein complexes, and DNA chemical adducts. Much of our knowledge of these events is based on analysis of markers of the replication stress and DNA Damage Response that follow stalling of replisomes. To examine consequences for the replisomes more directly, we developed an approach for imaging collisions of replication forks with the potent block presented by an interstrand crosslink (ICL). The strategy is based on the visualization on DNA fibers of the encounter of replication tracts and an antigen tagged ICL. Our studies revealed an unexpected restart of DNA synthesis past an intact ICL. In addition, and also unexpected, we found two distinct versions of the replisome, one biased toward euchromatin and the other more prominent in heterochromatin. Here, we present details of our experimental procedures that led to these observations.

Virtual-Reality Guided Versus Fluoroscopy-Guided Transseptal Puncture in a Cardiac Phantom.

OBJECTIVES: We compared virtual-reality guided versus fluoroscopy-guided transseptal puncture by novice and experienced operators in a cardiac phantom. Outcome measures included accuracy, time, transseptal path distance, and a survey of the operator experience. METHODS: A transseptal simulator was created using a Plexiglas case and a 3D-printed cardiac phantom with a replaceable fossa ovalis, a customized support, and an electromagnetic tracking system. A precisely registered virtual-reality rendering was constructed. To display the transseptal instruments in virtual reality, we attached electromagnetic sensors to standard transseptal instruments, including the needle, dilator, and sheath. Each subject completed 6 simulated transseptal punctures (3 fluoroscopy-guided and 3 virtual-reality guided). We measured the distance traversed by the transseptal needle, accuracy, and time for each simulated transseptal puncture. Operators were then surveyed regarding their experience. RESULTS: A total of 8 subjects (6 faculty, 2 fellows) completed the trial. We found that virtual-reality guidance resulted in significantly more accurate puncture site selection and, subjectively, was more intuitive for the operator, particularly for novices. None of the participants experienced negative symptoms in virtual reality that required cessation of the procedure. CONCLUSIONS: Virtual reality compared with fluoroscopic guidance for transseptal puncture shows considerable promise, particularly for novice trainees, where it could lessen the learning curve. Current barriers to widespread implementation are discussed.

DONSON and FANCM associate with different replisomes distinguished by replication timing and chromatin domain.

Duplication of mammalian genomes requires replisomes to overcome numerous impediments during passage through open (eu) and condensed (hetero) chromatin. Typically, studies of replication stress characterize mixed populations of challenged and unchallenged replication forks, averaged across S phase, and model a single species of "stressed" replisome. Here, in cells containing potent obstacles to replication, we find two different lesion proximal replisomes. One is bound by the DONSON protein and is more frequent in early S phase, in regions marked by euchromatin. The other interacts with the FANCM DNA translocase, is more prominent in late S phase, and favors heterochromatin. The two forms can also be detected in unstressed cells. ChIP-seq of DNA associated with DONSON or FANCM confirms the bias of the former towards regions that replicate early and the skew of the latter towards regions that replicate late.

Remodeling of Interstrand Crosslink Proximal Replisomes Is Dependent on ATR, FANCM, and FANCD2.

Eukaryotic replisomes are driven by the mini chromosome maintenance (MCM [M]) helicase complex, an offset ring locked around the template for leading strand synthesis by CDC45 (C) and GINS (G) proteins. Although the CDC45 MCM GINS (CMG) structure implies that interstrand crosslinks (ICLs) are absolute blocks to replisomes, recent studies indicate that cells can restart DNA synthesis on the side of the ICL distal to the initial encounter. Here, we report that restart requires ATR and is promoted by FANCD2 and phosphorylated FANCM. Following introduction of genomic ICLs and dependent on ATR and FANCD2 but not on the Fanconi anemia core proteins or FAAP24, FANCM binds the replisome complex, with concomitant release of the GINS proteins. In situ analysis of replisomes proximal to ICLs confirms the ATR-dependent release of GINS proteins while CDC45 is retained on the remodeled replisome. The results demonstrate the plasticity of CMG composition in response to replication stress.

Semantics-Based Composition of Integrated Cardiomyocyte Models Motivated by Real-World Use Cases.

Semantics-based model composition is an approach for generating complex biosimulation models from existing components that relies on capturing the biological meaning of model elements in a machine-readable fashion. This approach allows the user to work at the biological rather than computational level of abstraction and helps minimize the amount of manual effort required for model composition. To support this compositional approach, we have developed the SemGen software, and here report on SemGen's semantics-based merging capabilities using real-world modeling use cases. We successfully reproduced a large, manually-encoded, multi-model merge: the "Pandit-Hinch-Niederer" (PHN) cardiomyocyte excitation-contraction model, previously developed using CellML. We describe our approach for annotating the three component models used in the PHN composition and for merging them at the biological level of abstraction within SemGen. We demonstrate that we were able to reproduce the original PHN model results in a semi-automated, semantics-based fashion and also rapidly generate a second, novel cardiomyocyte model composed using an alternative, independently-developed tension generation component. We discuss the time-saving features of our compositional approach in the context of these merging exercises, the limitations we encountered, and potential solutions for enhancing the approach.