McArdle Blog L7 ~ 6/23/2014
Research Snapshot, Dr. Wei Xu: Same DNA, Cancer Cell
Proteins are the “workers” within our cells and bodies. Different proteins have different jobs; antibodies help protect us against infection from bacteria and viruses, structural proteins, like actin, serve as building blocks, and enzymes, such as polymerases, carry out the biochemical reactions necessary to keep us alive.
Some proteins, such as methyltransferases, can modify other proteins by adding small chemical tags. These tags can change where the modified protein goes within our cells or what functions it has. But sometimes tagging a protein can have unexpected consequences. Here at the McArdle Laboratory for Cancer Research Dr. Wei Xu and her colleagues have discovered that aberrant chemical tagging of one protein, BAF155, leads to its binding to unique regions of our DNA and contributing to more aggressive forms of breast cancer.
Following the trail: From CARM1 to BAF155
BAF155 is one component of a multi-protein complex called SWI/SNF (pronounced switch-sniff). A recent study from the Howard Hughes Medical Institute in Maryland found that protein subunits that make up this complex are mutated in close to 20% of the cancers they surveyed. But how these mutations in the SWI/SNF subunits are linked to cancer development has remained unclear.
Dr. Xu and her colleagues didn’t set out to study BAF155 or even the SWI/SNF complex. They were looking at an entirely different protein, called CARM1. Emerging evidence has established links between increased levels of CARM1 in cells and different cancers such as breast cancer. CARM1 – you can also call it Coactivator-Associated aRginine Methyltransferase 1 if you wish – modifies other proteins by tagging them with a methyl group (and this process is called methylation).
But we don’t know very much about which specific proteins are targeted for methylation by CARM1. Dr. Xu and her team set out to change that. Determining the targets of CARM1 methylation could help explain how increased amounts of CARM1 may be contributing to various cancers.
Finding CARM1 targets
To identify proteins that are methylated by CARM1 Dr. Xu’s lab used a technique called zinc-finger nuclease excision, which is much like using a pair of biochemically programmed scissors, to literally cut out a specific part of the CARM1 gene. The cells that had the damaged CARM1gene were unable to make any CARM1 protein.
This allowed the Xu lab access to some cells that make CARM1 and other cells that don’t. But how does that help identify CARM1’s target proteins? In the cells that don’t make any CARM1, its target proteins are not methylated. But these target proteins will be methylated in the cells that continue to make CARM1.
Dr. Xu then used a method called immunoprecipitation to separate out the methylated proteins from cells with or without CARM1. Think of a ball pit filled with balls of many different colors. You want to remove only the green balls. You have a special kind of sticky pad that attaches only to the green balls. You can tie iron nuggets to the sticky pads and mix them in with the balls. The sticky pads will attach to the green balls and you can then use a magnet to pull out iron+sticky pad+green ball complexes. Some balls of other colors might get pulled out but you will have enriched the proportion of green balls you have tremendously.
Similary, Dr. Xu used an antibody that specifically binds to proteins tagged with a methyl group. They took cells that make CARM1 and cells that don’t, broke them open, mixed in an antibody that binds to methylated proteins and pulled out the antibody+methylated protein complexes. By testing which proteins were enriched in the pool derived from cells with CARM1, they could determine which proteins were methylated when CARM1 was present but not when CARM1 was absent.
BAF155 was one such protein targeted for methylation by CARM1. The discovery of BAF155 as a target for methylation by CARM1 was exciting. Dr. Xu explained that BAF155 has been found over expressed in various types of cancer, but it has remained unclear how it actually contributes to these cancers. Could the methylation of BAF155 by CARM1 explain this enigma?
Methylated BAF155 leads to more aggressive cancers
Indeed, the Xu lab found that there was more methylated BAF155 in breast tumors compared to normal tissue from the same patient. Exploring further they found that patients whose breast tumors did not have methylated BAF155 were more likely to enjoy recurrence-free survival than patients whose breast tumors did have methylated BAF155. But what was methylated BAF155 doing in these tumors?
To answer this question the Xu lab engineered a mutant version of BAF155 that cannot be methylated by CARM1. They found BAF155 methylated by CARM1 can bind to different regions of a cell’s DNA compared to BAF155 that remained unmethylated. Even more surprisingly, it appeared that BAF155 was binding to these new DNA regions without being part of the entire SWI/SNF complex as it usually is.
Finding methylated BAF155 bound to unexpected DNA segments may be a clue as to how it contributes to cancer development. All your cells contain essentially the same DNA. But clearly you have many different kinds of cells, such as nerve cells, blood cells, liver cells and so on. What happens is that different parts of your DNA are used in different cells, much like you would use different parts of a recipe book depending on whether you were planning to make roast chicken or apple pie.
The process of controlling which parts of a cell’s DNA is used to make RNA and proteins is called epigenetics. There is an army of proteins that help determine which portions of your DNA are used in a given kind of cell. Any change in how these proteins function can cause devastating changes in the cell and even lead to the development of cancer. The SWI/SNF complex, and BAF155, help regulate which parts of DNA are usable in different cells. That’s partly how all your cells that contain the same DNA can behave differently.
By binding to unexpected DNA segments, methylated BAF155 was causing the genes in these regions to become active or more active than would be normal. Some of these genes contain the information the cell needs to make proteins with known links to cancer development when they are present in excess.
Summing it all up
Dr. Xu’s research is a wonderful exhibition of how basic research can open up previously unknown aspects of disease and potential treatment. By asking open-ended questions about how CARM1 is linked to cancer development, the Xu lab became the first to discover that methylation of BAF155 by CARM1 can contribute to making breast cancers more aggressive and likely to metastasize.
But the journey doesn’t end here. Dr. Xu points out that “breast cancer is often a multi-gene disease, and only genetics cannot give all the answers”. Studying epigenetic changes, such as the methylation of BAF155 by CARM1, is one important way to understand how tumor cells may be different compared to normal cells, even without obvious changes in DNA sequence between them.
Ultimately the goal is to understand these cancers to devise ways to treat them. Dr. Xu is passionate about her research; “I am personally motivated to help people (suffering from breast cancers) by studying and understanding the mechanisms. You can learn more about Dr. Xu’s work at http://mcardle.wisc.edu/bio/xu_w.html
This blog post is based on the paper “CARM1 methylates chromatin remodeling factor BAF155 to enhance tumor progression and metastasis” published in January, 2014 in the journal Cancer Cell.