Endoplasmic Reticulum

ID #1268

In the Pulse-Chase Experiment, I don't understand what the purpose of the Chase is? Why can't we just throw in a bunch of labeled AAs and then see where the proteins (that they make up) go? Why do we have to throw in unlabeled AAs after that?

Good question; let's think about the consequences of not performing the chase. If you're looking at cells specializing in secreting proteins, as Palade was, let's say you pulse with labeled methionine. You now begin "marking" every protein made. If you don't close that "window" by doing the chase, you will continue to be marking new proteins until you take your last time point, which could be hours later. In your first time points, will you still see your label where the proteins are actually synthesized? Yes, this part of the experiment would be no different than the "right" way. But now let's jump ahead by even ten minutes. If you take a time point now, will you see the original proteins in a different location? Yes. Will you see the latest proteins in the location that they were synthesized? Also yes. So let's interpret the data. You have radioactivity detected in the Golgi apparatus, and radioactivity detected in the endoplasmic reticulum. As a scientist, can you say for sure that the proteins you're detecting in the ER didn't start in the Golgi and make their way to the ER? We know (because of his experiments) that this isn't the case, but if you're the first one doing the experiment, you can't make that assertion. Maybe some of the detected proteins made it from the ER to the Golgi and back. Again, you can't say for sure. Coming back a few hours later, you'll again find some label in the ER, some in the Golgi, some in vesicles, and some outside the cell. How do you interpret this "mess?" Frankly, you couldn't. The bottom line is, if you want to follow a set of proteins, you need a way to distinguish that set from all the other proteins that are around -- those that came before as well as those that came later. So you have to close that "window" that you opened by pulsing, and that's the chase.

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