B/B Section Bank #38

[663697]

I'm having trouble understanding what the use of "total Rac" is here. The answer says that it is important to know if the total amount of Rac is affected by the strain used. The figure shows that total Rac is the same regardless of the treatment though, so how does it let us determine if VopC affected Rac expression levels?

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[aldol16]

Okay, so you knocked out VopC. What if VopC is co-expressed independently with Rac? In other words, say VopC and Rac both act on the same substrate and both need to be there for there to be any useful result. If VopC is suddenly knocked out, you'd also expect Rac to be down-regulated in that case because the cell has no need for Rac to be around without VopC as well. So in that case, you wouldn't see any GTP-Rac at all but it wouldn't be due to VopC activating Rac. It'd be a totally independent event.

 

[663697]

Okay, so you knocked out VopC. What if VopC is co-expressed independently with Rac? In other words, say VopC and Rac both act on the same substrate and both need to be there for there to be any useful result. If VopC is suddenly knocked out, you'd also expect Rac to be down-regulated in that case because the cell has no need for Rac to be around without VopC as well. So in that case, you wouldn't see any GTP-Rac at all but it wouldn't be due to VopC activating Rac. It'd be a totally independent event.

So based on the results, since total Rac levels didn't change regardless of VopC activation/inactivation, this indicates that Rac expression isn't dependent on VopC? And in the cases where there is GTP-Rac, we see that each case also has VopC present, but since the total Rac is unchanged, we can only conclude that VopC activates Rac -> GTP-Rac, but doesn't induce Rac expression? Just trying to see if this is the correct reasoning.

 

[rlaboss]

So based on the results, since total Rac levels didn't change regardless of VopC activation/inactivation, this indicates that Rac expression isn't dependent on VopC? And in the cases where there is GTP-Rac, we see that each case also has VopC present, but since the total Rac is unchanged, we can only conclude that VopC activates Rac -> GTP-Rac, but doesn't induce Rac expression? Just trying to see if this is the correct reasoning.

Yes. If they were looking at levels of Rac activation by VopC, they wouldn't want a cofounding variable causing the observed differences in Rac activation. If for some reason VopC caused a difference in expression levels of Rac, then a portion of the observed difference in Rac activity would be attributable to differences in Rac expression levels. By confirming that Rac expression doesn't change, we've eliminated the possibility of this confounding variable contributing to the observed differences in Rac activity level.