Enzyme Found to Control Gene Expression in a Novel Way—By Breathing New Life into Old RNA

By Jason Socrates Bardi


A recently identified enzyme from a family of enzymes that is ubiquitous in all eukaryotic life—from yeast to humans—has been found to play a novel role in regulating the expression of a gene in fission yeast Schizosaccharomyces pombe. This finding may have implications for the treatment of cancer.

The characterization of this regulatory enzyme, the result of a collaboration between the groups of investigators Paul Russell and John Yates at The Scripps Research Institute (TSRI), is described in an article in the latest issue of the journal Cell. In addition to having potential applications in medicine, the research is important from a basic science perspective because, unlike other enzymes of the same class, this new enzyme, called Cid13, works in the cytoplasm.

"Normally we think of genes that are involved in expression of mRNA operating in the nucleus," says Russell, who led the study. "This is a new class of enzyme that can control the activity of mRNA after it has been exported to the cytoplasm."

When genes are expressed in eukaryotic cells, the DNA of the genes is first transcribed into RNA "messages" in the nucleus by RNA polymerases, which read off the gene starting from the leading 5' end and terminating at a 3' tail. Then, this mRNA is transported outside the cell nucleus into the cytoplasm, but not before it undergoes splicing to remove any non-coding regions (introns) that are present and is polyadenylated by an enzyme called a poly(A) polymerase.

Poly(A) polymerases adds several adenines to the tail end of each mRNA molecule. Without this tail, the mRNA will not be translated into protein. In fact, once in the cytoplasm, the mRNA will only be translated into protein as long as the poly(A) tail is present. Eventually, nucleases in the cytoplasm can remove the tail, and then that mRNA becomes degraded.

Cid13, a type of poly(A) polymerase, rejuvinates expressed mRNA molecules that are about to be recycled by the cell by adding a new poly(A) tail to the mRNA.

"Our enzyme steps in when the poly(A) tail has been shortened but before the mRNA has been degraded," says Russell. "It resynthesizes a new poly(A) tail and breathes new life into that mRNA."

Previously it was thought that all poly(A) polymerases resided in the nucleus of somatic cells, where they did their work to freshly spliced mRNAs before they were transported out of the cell to meet ribosomes, but Cid13 works in the cytoplasm.

Interestingly, unlike other poly(A) polymerases in the nuclei, which add the tails indiscriminately, the Cid13 seems to reactivate only one particular gene—the mRNA of an important metabolic enzyme called ribonucleotide reductase.

Ribonucleotide reductase is a crucial enzyme because it supplies cells with the deoxyribonucleotides they need to build DNA. When cells begin to copy their DNA before they divide, these essential ribonucleotide reductase enzymes are upregulated in a variety of ways. One of these, apparently, is recycling used ribonucleotide reductase mRNAs and activating them before they can otherwise be degraded by the cell.

Interestingly, unlike other known poly(A) polymerases, Cid13 seems to be specific for ribonucleotide reductase mRNA, though follow up work in Russell’s lab is aimed at determining whether other mRNAs are targeted as well. Other follow-up work aims at determining how the enzymes are regulated. And given the ubiquity of this class of enzymes, they speculate that there may be specific Cid13-like enzymes in other organisms, including humans.

"The activity is possibly universal in [eukaryotes]," says Shigeaki Saitoh, who is first author on the paper.

Russell and Saitoh believe that the enzyme is a cell’s way of dealing with certain types of stress that can lead to DNA damage, like heat shock or oxidative damage. In such times of stress, the cell can use the Cid13 pathway to reduce the need to make fresh, potentially damaged, mRNA. But this type of enzyme might also be an important tool used by cancerous cells to fight chemotherapy.

In fact, the current study stemmed from Russell and Saitoh’s interest in understanding how cells survive treatment with hydroxyurea. Hydroxyurea is a U.S. Food and Drug Administration-approved cancer chemotherapy and is commonly given to AIDS and sickle-cell anemia patients. It works by inhibiting ribonucleotide reductase and choking off a cell’s supply of nucleotides.

Cells can become resistant to hydroxyurea by increasing the activity of the ribonucleotide reductase enzyme, a task which it may accomplish by using the molecule Cid13.

Saitoh found that strains with an abundance of Cid13 are resistant to hydroxyurea. Tumor cells that are resistant to hydroxyurea may be using a similar enzyme to increase the amount of ribonucleotide reductase that is translated. If this proves to be the case, then a Cid13 inhibitor in combination with hydroxyurea might be a more potent treatment than hydroxyurea alone.

This work was a collaboration between the laboratories of Paul Russell and John Yates. The Yates laboratory did mass spectroscopy on Cid13, which had already been identified but which scientists had assumed was a DNA polymerase. The mass spectroscopy allowed Russell and Saitoh to determine which other proteins Cid13 interacts with, which turned out to be proteins that bind to the poly(A) tail of mRNA. None were involved in DNA polymerization.

"Right away, this reinforced [the idea] that the enzyme was likely to be involved in RNA metabolism and poly (A) addition as opposed to DNA replication," says Russell, who stresses that the mass spectrometry technique was essential.

The article "Cid13 is a Cytoplasmic Poly(A) Polymerase that Regulates Ribonucleotide Reductase mRNA" is authored by Shigeaki Saitoh, Andrei Chabes, W. Hayes McDonald, Lars Thelander, John R. Yates III, and Paul Russell, and appears in the May 31, 2002 issue of the journal Cell.





Wild type and cid13- mutant yeast grown on a plate in the presence of 10 mM hydroxyurea. The wild type yeast (top portion) expresses Cid13 and survives. The mutant cannot express Cid13 and does not survive. (Picture courtesy of Shigeaki Saitoh.)