1. Modeling the involvement of Rpb4/7 in the mRNA biogenesis and degradation with Object-Process Methodology Dov Dori and Mordechai Choder Technion - Israel Institute of Technology August 2005 • We use Object-Process Methodology to model specific biological pathways at the molecular and cellular levels. • In this study we model biogenesis and degradation of messengerRNA (mRNA) and its transport between cellular compartments. • Our aim is to gain conceptual understanding of the role various factors play in this process, focusing on currently studied Rpb4/7. • The combination of formal yet intuitive graphics and automatically-generated equivalent natural language clarifies how experimental findings fit together and raises ideas for new research where gaps in knowledge exist.

2. OPM in a nutshell:Bread BakingandEatingexample Objects (green boxes) can have states (brown rounded corner rectangles):“Person can be hungry or satisfied.”Processes (ellipse, e.g., Baking) transform objects by: • Consuming :BakingconsumesDough. • Generating:“Baking yields Bread.” • Changing their state:“EatingchangesPersonfromhungrytosatisfied.” • Ovenis instrument forBaking: “BakingrequiresOven.” • Dough has parts: “Dough consists of Flour, Water, and Salt.” • Temperature is an attribute of water: “WaterexhibitsTemperature.” The text to the left, Object-Process Language (OPL) is a subset of English generated automatically and is fully equivalent to the OPD. The inverse action is also possible: One can write OPL sentences and the OPD will be updated automatically accordingly.

3. The basic cellular compartments The list above shows the hierarchy of Object-Process Diagrams (OPDs) describing the system. The OPD (top right) zooms into the Cytoplasm showing the structure of cellular compartments. The text below, Object-Process Language (OPL) is a subset of English generated automatically and is fully equivalent to the OPD.

4. The transcription process: mRNA synthesis The Transcription processes consumes Ribo-nucleotides to generate mRNAwithin the Chromosome using the DNA and the RNA Polymerase II as instruments. Compare the OPD with the automatically generated OPL text below.

5. Composition of RNA Polymerase II The OPD can schematically show (as done here) the spatial arrangement of the components that comprise the RNA Polymerase II. We propose to enhance OPCAT with spatial animation (see last slide) for enhanced visualization.

6. mRNA transport between cellular compartments Red links indicate that some of the proposed functions of Rpb4/7in the mRNA Transport & Decay process are hypothetical. Path names are recorded along arrows: nc = nucleus to cytoplasm cr = cytoplasm to ribosome rP = ribosometo P-body They help define the localization of mRNA-Rb4/7 Complex.

7. Our unpublished results suggest that Rpb4/7 is involved in translation. In OPL it means that in the Ribosome the mRNA Rpb4/7 Complex enables the Translation process, which consumes Amino Acid and yields Protein. Red links indicate uncertainty. We also propose that Pat1 undergoes Pat1 Recruitment by Rpb4 and Rpb7 to yield the mRNA Rpb4/7 Pat1 Complex. The red instrument links from Rpb4 and Rpb7 to Pat1 Recruitmentdenote that the involvement of Rpb4 and Rpb7 still needs to be experimentally verified. Rpb4/7 recruitment of Pat1

8. Like objects, processes can also be zoomed in. Here we zoom into the mRNA transport & decay process. Details of mRNA transport & decay Complete OPL text is in the next slide. Red indicates uncertainty as to: (1) whether Ribo-nucleotides fromDegradationof Bulk mRNA undergo Transport To Nucleus. RASING THIS QUESTION IS A DIRECT RESULT OF MODELING!(2) whether Deadenylation occurs only in P-body, and (3) whether Rpb4/7 participates in all these processes.

9. Details of mRNA transport & decay: The complete OPL text This is the complete OPL text of the OPD in the previous slide.

10. Rpb4/7-dependent mRNA degradation in the P-body Note that mRNA Transport & Decay was modeled earlier as a multi-stepprocess. Here we zoom in the P-body and the OPD shows only the relevant steps that take place in this compartment: Decapping and Degradation.

11. mRNA transport & decay: OPCAT animated simulation example Here we see mRNATransport & Decay occurringwith the sub-processes happen in a top-to-bottom order. We define Ribosome Entry as a process that changes Localization of mRNA from cytoplasm to ribosome: When OPCAT package is practiced, the red dots move along the arrows, the brown color of cytoplasm fades, while ribosome color is getting brown. This helps visualizing the process dynamics and finding “bugs” if one does not introduce data correctly. OPCAT can simulate the system by animating occurrence of processes, generation and consumption of objects, and change of their states. This cannot be performed with PowerPoint and requires OPCAT to be installed.

12. Free text definition of Adenylate cyclase from the Protein-Protein Interaction Ontology

13. The translation of the textual definition of Adenylate cyclase in previous slide to OPL script Adenylate Cyclase consists of Catalytic Domain.Adenylate Cyclase is encoded by CyaA.Ternary Complex consists of T25, T18, and Calmodulin.T18 and T25 are compelementary.Calmodulin triggers Association.CyaA is a Gene .Proteolytical Cleavage consumes Catalytic Domain .Proteolytical Cleavage yields T18 and T25.Association consumes T18, T25, and Calmodulin.

14. The automatically generated OPD (top) that is equivalent in its knowledge content to the OPL scrip (bottom), alsolisted in previous slide