I found a paper on the plant journal. In BiFC, the authors fuse YFPN to C-terminal of a plasma membrane protein called RSL1 (at the outside of the cell). And they fuse YFPC to a soluble protein (in the cytosol, ABA receptor in this paper). The results show that there are interaction on plasma membrane between the two protein. The author said: "The TM domain of RSL1 is only 20 AAs large and there are a few AAs behind. We guess the GFP protein does not cross completely the membrane to the outside face of the cell." How can GFP not cross completely the membrane? What does this mean? Is this possible?
I am not entirely sure if I catch your point in your question..however, GFP/YFP is a small protein and can easily enter any cell by passive transport; that being said if the GFP/YFP whole or partially fused to any TM domain, it certainly would not cross the cell membrane. Is it possible for you to share the paper with me [email protected]
Dear Jen-i-chen,
first of all, you should address your question to the authors. As the first respondent, we may also misunderstand your question. Of course, the interaction can not occur "across" different compartments. In fact, if a complementation (and thus fluorescence) is found this means that they are in the same compartment. Thus, in the described case either both outside or still inside the cell. What this means in this particulr case, this is what is probably discussed, i.e. either only a partial export of the fusion protein or a leakage of the cytosolic partner (maybe together with yet another interactor).
By the way, "a small protein" (and GFP is an intermediated sized protein) cannot cross a membrane passively (of course as anything in biology, this is not a 100% exclusion, if you wait a long time (maybe even longer than the lifespan of your organism) there might be an instance of spontaneous penetration/ diffusion.
best wishes,
Tony Schaeffner
I completely agree with Anton, you need to ask the corresponding author of te paper. But anyhow, as far as I recall, 20 AAs are almost enough to cross a typical membrane. So, maybe the authors suggest that the TM is not membrane spanning, but rather penetrating the membrane in a way the enables the C terminal to reenter the cytosol. Another possibility to explain this would be that their soluble protein can be exocytosed from the cell and interacts actually at the apoplast.
Cheers
Simon
Hello Jen, Good question, Can you share the doi OR link for the paper
I my case, for one of my interactors, I have a nuclear export signal
Hence I see interactions sometimes in cytosol
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