Paper6InClassQ&A

Questions for Team WEST Paper Discussion 6 MCDB 4650 Names of people in your group ______________________________ Lab TA______ 1. Describe the knockout mice that were used in this paper. How were they generated, and what does the data show regarding expression of FGF4 and FGF8 in the limbs? (Figure 1) 2. In Figure 2, how does this data lend credence to the pre-patterning hypothesis? Distal structures are present despite the lack of FGF8 and FGF4. If time spent proliferating (dependent on FGFs) were responsible for cell fate, only proximal elements would be present, but instead there are distal elements present in the absence of proximal elements. 3. What do the data in Fig. 3 a-h indicate? The AER is intact and survives in the double knockout situation. Other FGFs must take over the role of FGF8 in survival of the AER. Some important mesenchymal markers are not present , like BMPS, Hox 13, Ptch, etc, thus, the FGFs must be important for their expression. 4. What is shown in Figure 5 and what did it allow the authors to conclude about the function of FGF8? Here they looked at cell proliferation and found, rather surprisingly, that the cells could proliferate pretty normally in the absence of FGF4 and 8. This led them to conclude that FGFs were required for early proliferation, to allow for the right number of cells in the limb bud, but not for later proliferation. In the double KO, The hindlimbs start out really small; the forelimbs get smaller due to cell death. Questions for Team EAST Paper Discussion 6 MCDB 4650 Names of people in your group _____________________________ Lab TA______ 1. Why is there expression of FGF4 and FGF8 in the forelimb in the double knockout? Why might FGF4 be turned on early in the forelimb? Msx-2 expression comes on earlier in the hindlimb than in the forelimb, so there is some expression of the fgfs in the forelimb before cre cuts out exons 2 and 3. fgf-4 might come on early because of fgf8 expression. 2. How does the data in Figure 2 i-p correlate to the phenotypes of the double knockout limbs (Figures 2 a-h). it correlates very well—in hindlimb, nothing actually develops, but in the forelimb, the humerus is normal, while the autopod and zeugopod are truncated. 3. Why are the double knockout limb buds reduced in size, and what was shown in Fig 4? How do these data relate to the other data in the paper? These experiments looked at cell death. Apoptotic cells are in high numbers in the absence of AER, and also in the double knockout situation. Interestingly, the domain of cell death was not right underneath the AER, but rather deeper in the mesenchyme. Could this be why the stylopod is so severely affected in the hindlimbs? Why isn’t the stylopod more affected in the forelimbs? 4. Did the authors convince you that the prepattern model was more reasonable than the progress zone model? Why or why not? Which pieces of data were most convincing? Clicker?? In Figure 2 i-p: what does the Sox-9 expression tell you about the number of cells in each region (stylopod, zeugopod and autopod) of the limb bud? all three regions of the limb are affected equally (decreased) in the double knockouts in the hindlimb, stylopod population is greatly decreased, but in the forelimb, stylopod population is relatively unaffected. In the forelimb, stylopod population is greatly decreased, but in the hindlimb, stylopod population is relatively unaffected. In the forelimb, all three regions of the limb are normal B