New & Noteworthy
February 25, 2014
Most people know that Incans relied on human runners to get messages across their empire. Basically they had runners stationed at various places and one runner would hand the message off to the next. This relayed message could then quickly travel across the country.
As shown in a new study by Nadal-Ribelles and coworkers, it turns out that something similar happens in yeast when the CDC28 gene is turned up in response to high salt. In this case, the runner is the stress activated protein kinase (SAPK) Hog1p and it is stationed at the 3’ end of the gene. When the cell is subjected to high salt, the message is relayed from the 3’ end of the CDC28 gene to its 5’ end by the Hog1p kinase. The end result is about a 2-fold increase in the amount of Cdc28p made, which allows the cell to enter the cell cycle more quickly after the salty insult.
Unlike the Incans who had their paths all set up in front of them, poor Hog1p has to build its own path. It does this by activating a promoter at the 3’ end of the CDC28 gene that produces an antisense long noncoding RNA (lncRNA) that is needed for the transfer of the Hog1p. It is as if our Incan runner had to build a bridge over a gorge to send his message.
This mechanism isn’t peculiar to the CDC28 gene either. The authors in this study directly show that something similar happens with a second salt sensitive gene, MMF1. And they show that a whole lot more lncRNAs are induced by high salt in yeast as well.
Nadal-Ribelles and coworkers started off by identifying coding and noncoding regions of the yeast genome that respond positively to high salt. The authors found that 343 coding regions and 173 noncoding regions were all induced at 0.4 M NaCl. Both coding and noncoding regions required the SAPK Hog1p for activation.
The authors next focused on CDC28 and its associated antisense lncRNA. After adding high salt, Nadal-Ribelles and coworkers found that Hog1p was both at the start and end of the CDC28 gene – as would be expected, since both CDC28 and the antisense lncRNA required this kinase for transcriptional activation.
Things got interesting when they were able to prevent the lncRNA from being made. When they did this, Hog1p was missing from both the 5′ and 3′ ends of the CDC28 gene and as expected, activation was compromised. But Nadal-Ribelles and coworkers showed that expressing the lncRNA from a plasmid did not allow for CDC28 activation. It appears that where the lncRNA is made is just as important as whether it is made.
Through a set of clever experiments, the authors showed that not only does the lncRNA need to be made in the right place, but it needs to be activated in the right way. When they set up a system where the lncRNA was induced in the right place using a Gal4-VP16 activator, CDC28 was not induced by high salt. A closer look showed that this was most likely due to a lack of Hog1p at the start of the CDC28 gene.
The situation was different when they activated the lncRNA with a Gal4-Msn2p activator which uses Hog1p to increase expression. In this case, CDC28 now responded to high salt and Hog1p was present at both the start and end of the CDC28 gene. But this activation went away if they added a terminator which prevented the full length lncRNA from being made.
Phew, that was a lot! What it means is that for there to be a Hog1p at the business end of the CDC28 gene, there needs to be one at the 3’ end. It also means that for the Hog1p to get to the start of the CDC28 gene, the antisense lncRNA needs to be made.
This would all make sense if maybe the lncRNA was involved in DNA looping, which could get the Hog1p from the end of CDC28 to the start where it can do some good. Nadal-Ribelles and coworkers showed that this indeed was the case, as CDC28 activation required SSU72, a key looping gene. When there was no Ssu72p in a cell, salt induction of CDC28 was severely compromised.
So it looks like an antisense lncRNA in yeast is being used as part of a looping mechanism to provide the cell with a quick way to start dividing once it has dealt with its environmental insult. The authors show that yeast that can properly induce their CDC28 gene enter the cell cycle around 20 minutes faster than yeast that cannot induce the gene. The cells are poised for a quick recovery.
And this is almost certainly not merely a yeast phenomenon. Some recent work in mammalian cells has implicated lncRNAs in recruiting proteins involved in controlling gene activity through a looping mechanism as well (reviewed here). Now that the same thing has been found in yeast, scientists can bring to bear all the powerful tools available to dissect out the mechanism(s) of lncRNA action. And that’s far from a loopy idea…