Analyze the Data Chapter 4: FACS of Glucose Transport In unstimulated muscle cel
ID: 196523 • Letter: A
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Analyze the Data Chapter 4: FACS of Glucose Transport
In unstimulated muscle cells, the glucose transporter Glut4 is present in the membrane of endosomes. Upon stimulation of these cells with insulin, the endosomes fuse with the plasma membrane, thereby depositing Glut4 in the plasma membrane, where it serves to transport glucose into the cells (see Figure 15-34 on page 659). In order for insulin to elicit this change in Glut4 localization, insulin must bind to its receptor, present in the plasma membrane of some cells. To determine if a stem cell line can be induced in culture to differentiate into muscle cells in which Glut4 is expressed and exhibits this insulin-dependent change in behavior, the following studies were undertaken.
A stem cell line was incubated under culture conditions in which stem cells can differentiate into various specific cell types. The cells were then incubated in the presence and absence of insulin. To determine if either Glut4 or the insulin receptor were present on the cell surface of cells within this population, antibodies that specifically recognize each protein were used to label the cells. Glut4 on the cell surface was detected by an anti-Glut4 antibody, which fluoresces green; the insulin receptor on the cell surface was detected by an anti-insulin receptor antibody, which fluoresces red. These cells were then placed in a fluorescence activated cell sorter FACS machine (see Figure 9-29 in the text) and sorted based on both red and green fluorescence. The following data were obtained. Each • on the graphs represents 20 cells. An increased value of fluorescence intensity indicates that more antibody has bound to those cells.
A. Do all cells in the population express insulin receptor on their surface? Is this expression dependent on the presence of insulin? Explain.
(a) Insulin 10 104 10 b) Insulin 107 104 100 104 107 10° 104 10 Red fluorescence Red fluorescence.Explanation / Answer
A. Yes from the FACS data fig (a) it can be deduced that all the cells in the population express insulin receptor on their surface. The red fluorescence is very high due to the binding of anti-insulin receptor antibody.
Yes, the expression is dependant on the presence of insulin, in presence of insulin endosome containing the GLUT-4 protein fuses with the plasma membrane thereby transporting glucose inside the cell.
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