New & Noteworthy

The Brazor of Biology

January 8, 2014

The janitor on the U.S. comedy series Scrubs is always coming up with terrible inventions. One of his worst was the knife-wrench. It is what it sounds like—a tool with a knife at one end and a wrench at the other.

Just like this brush-razor, or brazor, Cet1p has two distinct, but related, functions.

Of course not all dual purpose tools have to be so useless.  Imagine a tool like the one at the right with a razor at one end and a toothbrush on the other.  Now you can easily brush your teeth and shave in the shower or at your bathroom sink (as long as you are careful not to cut your cheek).

Turns out that biology has these dual purpose tools too except that they are almost always more useful.  For example, Lahudkar and coworkers show in the most recent issue of GENETICS that Cet1p doesn’t just help out with capping mRNA.  No, these authors found that it also helps clear RNA polymerase II (RNA pol II) away from promoters.  And what’s most interesting is that this second function has little to do with its job in mRNA capping.

Basically the two functions are probably in the same protein because they both happen in the same place, at the start site of a promoter.  Just like our brazor is useful because both jobs happen in the bathroom.

The first step was to show that in the absence of Cet1p, RNA pol II was more likely to be found near the start of transcription.  The authors showed that this was the case by using a temperature sensitive mutant of Cet1p and a chromatin immunoprecipitation (ChIP) assay targeted at RNA pol II—there was more RNA pol II crowded near the promoter at the nonpermissive temperature. 

The next set of experiments showed that merely messing with the cap is not sufficient to cause the polymerase to pause. Lahudkar and coworkers found that RNA pol II occupancy was unchanged in strains carrying mutations in STO1 (also known as CBP80) or CEG1, two components of the capping machinery. Cet1p apparently has a separate, unrelated function in helping to clear polymerases away from the start site of transcription.

The final set of experiments showed that the unpausing activity of Cet1p was found in a different part of the protein from its capping function.  Cet1p be can be broadly divided into three regions—a poorly characterized N-terminal domain (amino acids 1-204), a Ceg1p interaction domain (aa 205-266), and a triphosphatase domain (aa 265-549).  The last two domains are critical to its capping function.

Lahudkar and coworkers found that deleting the 1-204 aa domain from Cet1p caused polymerase stalling at the promoter without affecting its capping ability.  And conversely, that when they impaired the ability of Cet1p to perform its capping function while retaining its 1-204 aa domain, RNA pol II escaped the promoter at the same rate as it did in the presence of wild type Cet1p.  A final experiment showed that just expressing the first 300 amino acids of Cet1p was sufficient to get the polymerases moving. 

All in all these experiments provide strong evidence that Cet1p has two separate functions—an enzymatic role in capping mRNA and an unrelated activity that helps clear RNA pol II from the regions around the promoters of genes.  Which all goes to show that even when you think you have a handle on a protein, it can still surprise you with something new.  Turn it around and you just might find a toothbrush at the end.

by D. Barry Starr, Ph.D., Director of Outreach Activities, Stanford Genetics