Using online freeware Cell Collective, you will observe the processes of cell cycle as you perform computer simulations to investigate the cell cycle check points
associated activities are designed to help you work through the cell cycle and stages of mitotic division. Using online freeware Cell Collective, you will observe the processes of cell cycle as you perform computer simulations to investigate the cell cycle check points. Additionally, you will also use an online database of cell images to identify the mitotic cell division stages. Key Learning Objectives: Identify the stages of mitotic division #UNDERSTAND Outline the processes of the cell cycle #UNDERSTAND Describe what happens in a cell cycle simulation #CREATE Design a graph to communicate results of number of cells in mitotic division stages #CREATE Lab Exercise #1 PART A – Cell Collective: Cell Cycle Regulation – Investigation 1 The cell divides by mitosis for several reasons including growth, repair, and reproduction. The cell cycle includes mitotic division and interphase (G1, S, G2). There are checkpoints the cell uses to control when each process can proceed to ensure equal distribution of genetic material. The eukaryotic cell will go through interphase prior to mitotic division. The cell cycle is dependent upon the concentrations of cyclin within the cell and how much cyclin is bound to CDK (protein complex). When cyclin and CDK combine they form MPF (Maturation Promoting Factor). Figure 1. The typical eukaryotic cell cycle including interphase and the mitotic phase as diagrammed in Cell Collective (Helikar et al. 2015). 1 BIOL 1020 Fall 2020 Lab 7: Cell Cycle & Mitosis Interphase includes three stages, G1 (the first gap phase), S (synthesis phase), and G2 (the second gap phase). Progression through these phases is regulated by checkpoints. First Gap Phase (G1): This is phase is defined by cell growth and the presence of proteins required for cellular division. The G1 checkpoint restricts cells from passing unless they have all the requirements for complete division. Growth factors are proteins that are need in higher concentrations to promote the growth of the cell to a sufficient size. Synthesis (S): In this phase is when the DNA is duplicated, called synthesis (for which is where the phase gets the name S). Second Gap Phase (G2): In this phase the cell continues to grow and prepare for mitotic division and requires a certain amount of proteins that will allow for nuclear envelop to fragment and centrosomes to develop microtubules. The Mitotic phase includes the five stages where DNA material is being divided into two genetically identical daughter cells and will be further outlined in Lab Exercise #2. Lab Exercise #1 PART B – Cell Collective: Cell Cycle Regulation – Investigation 1 In your small working groups, you will simulate the cell cycle activity in two different scenarios. The model that you will be working with in Cell Collective represents a simplified version of the G1-S phase transition pathway. Progress of real cells through the cell cycle is triggered by specific environmental proteins (e.g., growth factors). There is a point in G1 where the cell no longer depends on the presence of these environmental signals in order to progress through the cell cycle. This commitment is driven by a positive feedback loop in the G1 phase which amplifies and sustains the activity of the proteins needed to drive progression from G1 to S phase even when the environmental signals are removed. Link to Cell Collective Module – https://learn.cellcollective.org/?dashboard=true#module/36647:1/cellcycle-regulation-investigation-1/1 Page 1 & 2 These pages are designed to provide you with an overview of the cell cycle and how the model has been constructed to simulate the regulation of the checkpoints. Your group should read through these pages to familiarize yourselves with the model set-up (it has been pre-built for you), you will only need to run simulations. 2 BIOL 1020 Fall 2020 Lab 7: Cell Cycle & Mitosis Page 3 On this page your group will fill in the table below to record your predictions when the only one external component, growth factor, is included in the model. Record your predictions for the three components when growth factor is initially fully available to cell, but then growth factor activity level is set to zero or is unavailable to the cell. Be sure to include a statement to explain your prediction. Cell Cycle Component Activity will (remain on, turn off, switch between on and off) Growth factor receptor G1 Protein G1 – S Transition Page 4 Now, you will run the simulation outlined above to test your predictions. • • • • • • Within the Simulation Control panel, made sure the Sliding Window Size is set to 1 Within the External Components panel set the activity for Growth Factor to 100 Within the Activity Network panel click on the Growth Factor Receptor, G1 Protein, and G1-S Transition component circles so they become gray/blue You can now click Play and run the simulation for ~ 60 time steps Without stopping or pausing the simulating, change the activity level of Growth Factor in the External Components panel to zero Record what happens in the table below Cell Cycle Component Observations of activity when Growth Factor activity changes from 100 -> 0. Growth factor receptor G1 Protein G1 – S Transition 3 BIOL 1020 Fall 2020 Lab 7: Cell Cycle & Mitosis What do your observations inform you about the influence of Growth Factor? #APPLY Page 5 On this page your group will fill in the table below to record your predictions when the both external components, growth factor and positive feedback loop, are included in the model. Record your predictions for the three components when growth factor is initially fully available to cell, but then growth factor activity level is set to zero or is unavailable to the cell. Be sure to include a statement to explain your prediction. Cell Cycle Component Activity will (remain on, turn off, switch between on and off) Growth factor receptor G1 Protein G1 – S Transition Page 6 Now, you will run the simulation outlined above to test your predictions. • • • • • • Within the Simulation Control panel, made sure the Sliding Window Size is set to 1 Within the External Components panel set the activity for Growth Factor and Positive Feedback Loop to 100 Within the Activity Network panel click on the Growth Factor Receptor, G1 Protein, and G1-S Transition component circles so they become gray/blue You can now click Play and run the simulation for ~ 60 time steps Without stopping or pausing the simulating, change the activity level of Growth Factor in the External Components panel to zero Record what happens in the table below 4 BIOL 1020 Fall 2020 Cell Cycle Component Lab 7: Cell Cycle & Mitosis Observations of activity when Growth Factor activity changes from 100 -> 0. Growth factor receptor G1 Protein G1 – S Transition What do your observations inform you about the influence of Positive Feedback Loop? #APPLY Lab Exercise #1 PART C – Cell Collective: Cell Cycle Regulation – Investigation 1 Back in the main Zoom session, one of your TAs will review a cell cycle simulation graph and guide you through how to construct informative quantitative descriptions of the results (graph). Describing data can either be done qualitatively or quantitively, and often both types of descriptions can be useful for communicating and interpreting results. How are the two different? For this exercise, we will focus on quantitatively describing figure 2. Figure 2. Cell Collective simulation of the cell cycle with only external component growth factor influencing the activity level of other components. 5 BIOL 1020 Fall 2020 Lab 7: Cell Cycle & Mitosis When you are describing data as a written summary, just as in graphing, it is important to think about what the main message you want to communicate is. In our example with figure 2, we have simulated the changes in growth factor receptor, G1 Protein and G1 – S Transition using the external component, growth factor. 1) What happened to growth factor activity level at time 40? 2) Do you think the growth factor activity level at time 70 – 110 is zero? 3) When is growth factor activity likely zero? 4) What is happening to growth factor activity at time 160? Using the questions above and with the TA guidance, write out 1 – 3 sentences describing the results of the simulation. Lab Exercise #2 PART A – Identifying Mitotic Stages Adapted from Shelden et al. 2019 In the main Zoom session, one of your TAs will review the mitotic division stages so you can identify the cells in the dataset that is provided on UMLearn – Lab Handouts – Lab 7 Dataset. The mitotic division phase of the cell cycle has five cellular division stages. What are they? What are the defining features to identifying which stage of mitotic division a cell is in? What types cells don’t go through cytokinesis? #UNDERSTAND 6 BIOL 1020 Fall 2020 Lab 7: Cell Cycle & Mitosis You will conduct an analysis of the stages of cell division using real data images provided by the Allen Institute for Cell Science in Seattle, Washington. Your lab instructors have set up the photographs of the cell from their website for you. We will be analyzing ten images of stained cells, to assist in identifying the mitotic stage. The cells are stained to make viewing of DNA and mitotic spindles easier. With your knowledge of the events of mitosis, you will identify and count the cells at each stage: Interphase, Prophase, Prometaphase, Metaphase, Anaphase and Telophase. Example Cell Identification & Counting Your TAs will do an example image identification and counting, use the table below to record the data. Interphase Prophase Prometaphase Metaphase Anaphase Telophase Example Image Total How would you graph this data? #ANALYZE What is the independent variable? #ANALYZE What is the dependent variable? #ANALYZE Is the independent variable categorical or continuous? #ANALYZE Lab Exercise #2 PART B – Identifying Mitotic Stages In your small working groups, you will have one person share their screen with the Lab 7 Dataset PDF found in UMLearn. As a group you will discuss the following questions with the note that interphase is the longest part of the cell cycle, so for this lab exercise and associated assignment we are focusing on the stages of mitosis. Do you think all stages will have equal counts? Why or why not? # APPLY Which stage of mitosis do you think will be most common? Why? #APPLY Which stage of mitosis do you think will be least common? Why? #APPLY The questions above are not part of your assignment but will assist you in your interpretation of the results. Not all cells go through the cell cycle at the same frequency, recall from your lecture why that might be the case. 7 BIOL 1020 Fall 2020 Lab 7: Cell Cycle & Mitosis Data Collection When you are in your breakout room identifying and counting the cells use table 1 to record your observation. The dataset provided to in UMLearn lab handouts content folder to view all ten images. The first five images are part of Cell Group A, and the second set of five images are part of Cell Group B. You will treat each group separately and compare the results of each group in your summary as part of your assignment. Table 1. Counted results from the dataset provided of cells in the cell cycle. Images 1-5 represent a dataset of Cell Group A, and Images 6 – 10 represent a dataset of Cell Group B. Cell Group A B Image Interphase Prophase Prometaphase Metaphase Anaphase Telophase 1 2 3 4 5 Total (1-5) 6 7 8 9 10 Total (6-10) Which Cell Group goes through the cell cycle more rapidly? How can you tell? #ANALYZE 8 BIOL 1020 Fall 2020 Lab 7: Cell Cycle & Mitosis Lab 7 Assignment This assignment has TWO stages and will be submitted and completed in Peerceptiv. You should upload your document to Peerceptiv as either a Word or PDF document only. You will have two days for EACH stage. This lab is worth 4% of your BIOL 1020 final grade and is out of 26 marks. This assignment is designed to give you an opportunity to continue to practice your scientific writing and data presentation skills. You should be focusing on clearly and efficiently communicating your results. PART A – Mitotic Cell Division Graph In your lab you used ten images to identify and count cells going through the cell cycle. There were two datasets: Cell Group A (images 1 – 5) Cell Group B (images 6 – 10) To represent the results, you will construct a single graph to represent both datasets on the same graph. You will graph the results with each stage (not including interphase). Be sure to make sure the graph clearly displays the data for both cell groups and has all the necessary components (see the rubric on page 10 and 11 as your checklist). PART B – Mitotic Cell Division Result Summary Once you have constructed your graph, you will also write a result summary in 300 words or less. Within the summary you should summarize the trends of both cell groups that is reflected in the graph you construct. In your summary address the following questions: • What are the trends in your graph? • State how many cells in each of the two cell groups (A and B) are in interphase • Which cell group (A or B) do you think is going through the cell cycle quicker? Why or Why not? Use your results to support your answer. 9 BIOL 1020 Fall 2020 Lab 7: Cell Cycle & Mitosis RUBRIC: Part A: Graph (16 marks) Mechanics ( /8 Marks) Descriptive Caption 1 Mark – Should be: a) in the form of a statement, b) mention the subject(s), c) appropriate variables including a key to their representation (i.e., colour or pattern), d) include relevant details about the experiment essential for understanding the take home message 0.75 Mark – The caption is present but missing one of the above criteria. 0.5 Mark – The caption is present but missing two of the above criteria. 0.25 Mark – The caption is present but missing three of the above criteria. 0 Marks – Topic is not clearly identified or communicated X Axis Label 1 Mark – Should be descriptive and appropriate for the experiment. For categorical data sets, labels are also included for each dataset 0.75 Mark – The X-axis is present but not descriptive 0.5 Mark – The X-axis is present but not appropriate 0.25 Mark – The X-axis is unclear 0 Marks – There is no X-axis X Axis Units 1 Mark – Should be descriptive and appropriate for the experiment 0.75 Mark – The X-axis units are not descriptive 0.5 Mark – The X-axis units are not appropriate 0.25 Mark – The X-axis units are unclear and confusing 0 Marks – There is no X-axis unit present Y Axis Label 1 Mark – Should be descriptive and appropriate for the experiment. For manipulated data sets, the manipulation is indicated (average results vs. raw data) 0.75 Mark – The Y-axis is present but not descriptive 0.5 Mark – The Y-axis is present but not appropriate 0.25 Mark – The Y-axis is unclear 0 Marks – There is no Y-axis Y Axis Units Scale 1 Mark – Should be descriptive and appropriate for the experiment 0.75 Mark – The Y-axis units are not descriptive 0.5 Mark – The Y-axis units are not appropriate 0.25 Mark – The Y-axis units are unclear and confusing 0 Marks – There is no Y-axis unit present 1 Mark – Intervals and range should be appropriate for the data displayed. 0.5 Mark – The scale is not appropriate for one of the axes. 0 Mark -The scale is not appropriate for any axis. Continued on the Next Page 10 BIOL 1020 Fall 2020 Lab 7: Cell Cycle & Mitosis Legend 1 Mark – Should be appropriate and descriptive for the data displayed and include descriptions of different colours/patterns 0.5 Mark – The legend is not descriptive or includes errors 0 Mark – The legend is not present, and is needed Accuracy 1 Mark – The collected data is presented accurately with respect to both axes 0.75 Mark – The collected data is presented accurately to the dependent variable only 0.5 Mark – The Collected data is presented accurately to the independent variable only 0 mark – The data is not presented accurately Communication ( /4 Marks) Ease of Understanding 2 Marks – The graph is constructed in a way that is: a) clear to identify trends and b) easy to note the take home message 1 Mark – The graph is constructed in a way that data trends are difficult to observe or it is difficult to understand the take home message. 0 Marks – The graph is ineffective at communicating trends and the take home message. 2 Marks – The graph has: a) data plotted to take up sufficient room in the Cartesian plane, b) legible fonts, c) clear and legible X- and Y- axis lines, d) data displayed in the appropriate number of bars/lines, and e) no junk elements such as distracting colours, patterns, dark grid lines, … Aesthetics 1.5 Marks – The graph is missing one of the points above. 1 Mark – The graph is missing two of the points above. 0.5 Mark – The graph is missing three of the points above. 0 Marks – The graph has too many flaws that interfere with the interpretation of the data Graph Choice ( /4 Marks) 2 Marks – The data displayed is appropriate for both independent and dependent variables and data (i.e., categorical or continuous). Graph Type 1.5 Marks – One minor error in how the data are displayed 1 Mark – Two minor errors in how the data are displayed 0.5 Mark – The data displayed is not suitable for either the dependent or independent variable OR there is a better way to present the data. 0 Marks – The graph is not suitable for both experimental variables Continued on the Next Page 11 BIOL 1020 Fall 2020 Data Displayed Lab 7: Cell Cycle & Mitosis 2 Marks – The graph indicates the type of data (i.e., raw, mean, etc.) that are plotted. There should be a clear distinction between raw and manipulated data based on information in the legend and axes labels. 1.5 Marks – The graph is missing one of the points above. 1 Mark – The graph is missing two of the points above. 0.5 Mark – The graph Is missing three of the points above. 0 Marks – The graph type is in appropriate for the data type. Part B: Summary of Results (10 Marks) Summary of Results Cell Group Results 3 Marks – The results are stated clearly for both cell groups 2 Marks – The results are stated clearly for one of the cell groups 1 Mark – The results are stated for both cell groups, but not clearly 0.5 Marks – The results are stated for one cell group, but not clearly 0 Marks – The results are not stated clearly. Cell Group – Interphase Results 1 Marks – The number of cells in interphase is stated for both cell groups. 0.5 Marks – The number of cells in interphase is stated for one of the cell groups. 0 Marks – The number of cells in interphase is not stated for either cell group. Cell Group – Division Frequency 1 Marks – It is stated which cell group went through the cycle more quickly and it is clearly justified why. 0.5 Marks – It is stated which cell group went through the cycle more quickly and not clearly justified why. 0 Marks – It is not stated which cell group went through the cycle more quickly. 5 Marks – The summary is well written and clearly summarizes the graph and cell group results. Cell Group Graph Summary (Clarity) 4 Marks – The summary is well written and summarizes the graph and the cell group results but needs to be written more clearly. 3 Marks – The summary is well written and vaguely summarizes graph and includes important information about the two cell groups. 2 Marks – The summary contains numerous spelling or grammatical errors, and vaguely summarizes the graph and cell group results 1 Mark – The summary is written but does not explain the graph. 0 Marks – The summary is too challenging to read and u …
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