Data interpretation question

[theonlytycrane]

Passage excerpt:

Genetics and cell biology based experiments link ARF1 to lipid droplet biogenesis. ARF1 function is regulated by the exchange of bound GDP for GTP via an exchange factor (e.g. GBF1) transitioning ARF1 from an inactive to an active conformation. This is then followed by hydrolysis of bound GTP to GDP transitioning ARF1 back to an inactive conformation. Due to its intrinsically low GTP hydrolysis rate, ARF1 requires an activating protein for GTP hydrolysis. In mammalian cells, three related activating proteins termed ARFGAP1-3 are linked to COPI vesicle formation through ARF of which ARFGAP1 is the prototypical protein.

Then they knockdown ARFGAP1 and measure effect on the lipid droplets (vesicles).

My interpretation:

When ARFGAP1 is knocked out, lipid droplet counts per cell decreased. This seemed weird to me though, since ARFGAP1 inactivates ARF1 which helps makes the lipid droplets. I expected droplet counts to increase when ARFGAP1 (the inactivator) was knocked out. I guess I can't really assume a biological mechanism, though. Is it possible that ARFGAP1 inactivates ARF1 and in doing so somehow completes lipid droplet synthesis or release?

Source: http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0111309

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[aldol16]

When ARFGAP1 is knocked out, lipid droplet counts per cell decreased. This seemed weird to me though, since ARFGAP1 inactivates ARF1 which helps makes the lipid droplets. I expected droplet counts to increase when ARFGAP1 (the inactivator) was knocked out. I guess I can't really assume a biological mechanism, though. Is it possible that ARFGAP1 inactivates ARF1 and in doing so somehow completes lipid droplet synthesis or release?

ARFGAP1 is an activator. You say it yourself.

n mammalian cells, three related activating proteins termed ARFGAP1-3 are linked to COPI vesicle formation through ARF of which ARFGAP1 is the prototypical protein.

This is just like a G-protein.

 

[theonlytycrane]

@aldol16 When the GTP on ARF1 is hydrolyzed, it's inactivated. By turning ARF1 off, I was thinking that ARFGAP1 would be an inactivator?

 

[aldol16]

@aldol16 When the GTP on ARF1 is hydrolyzed, it's inactivated. By turning ARF1 off, I was thinking that ARFGAP1 would be an inactivator?

Hmmm, it looks like they are not completely clear on this point, which is why they present weird data. They say that the ARFGAP proteins are activators that are linked to vesicle formation but then they say it right after they say that ARF1 needs GTP hydrolysis activators. Your interpretation is correct if the latter case is correct - that is, that ARFGAP proteins regulate GTP hydrolysis. In that case, GTP hydrolysis would be accelerated, thereby inactivating the ARF1 proteins.

 

[bobeanie95]

Hmmm, it looks like they are not completely clear on this point, which is why they present weird data. They say that the ARFGAP proteins are activators that are linked to vesicle formation but then they say it right after they say that ARF1 needs GTP hydrolysis activators. Your interpretation is correct if the latter case is correct - that is, that ARFGAP proteins regulate GTP hydrolysis. In that case, GTP hydrolysis would be accelerated, thereby inactivating the ARF1 proteins.

^ I agree. They state in the conclusion that lipid formation could be impaired due to cell stress as a result of knocking down ARFGAP. The specific mechanism is unclear as stated: "The physiological context as to why ARFGAP1 is present on lipid droplets remains to be determined. " Usually, you'd be correct to assume that but in this case it could be that knocking down the activator could inhibit maybe clathirin formation and affect lipid synthesis.