Willy Wonka had all kinds of treasured secrets in his factory. The list includes, but is not limited to: a chocolate waterfall, orange skinned/green haired midgets, everlasting gobstoppers, and geese that laid eggs full of the goodness that is both chocolate and gold. Harboring these kinds of objects also lent itself to a great deal of paranoia on Wonka’s part. Arthur Slugworth was lurking outside the walls, desperate to get inside and steal the secrets for his own candy factory. Wonka should have really invested in some sort of protection that wasn’t his own madness.
Luckily, our cells (which run something like factories or, you know, bakeries) do have a police force. Aside from viruses breaking in (see Spanish Influenza Parts 1 – 3 posts), cells are susceptible to all sorts of problems, such as DNA damage, misfolded proteins or improper protein function. Several proteins within our cells have the sole job to maintain order, trust, and well being to our cellular processes. If things go awry, they have orders to suspend operations (also referred to as cell cycle arrest) or simply destroy all evidence of the cell (known as apoptosis).
One of the key “policing” proteins is called p53. It hangs around in the cell with its ear to the phone, waiting for word of some problem to reach it. A base of DNA changes from an A to a T? p53 is called in. A cell is breaking through its checkpoints in growth? Someone get p53 on the phone. Hemoglobin’s latest shipment of oxygen did not come in? p53 will take care of it.
p53 is kind of a big deal.
After receiving information that problems are afoot, p53 goes into the nucleus and binds the DNA in very specific places. This binding of DNA leads to cell cycle arrest or apoptosis (for this post, it is not important how - just understand that it has to bind DNA properly to fulfill its function.)
These problems that arise in the cell are serious and need to be taken care of swiftly. Think back to the Central Dogma post. What would happen if an A was changed to a T in DNA? Well… if the A to T mutation was in an area that coded for a protein, the mutation would be passed on to the RNA and the wrong amino acid would be incorporated into the growing protein at the ribosome. It may not seem like a big deal that one amino acid is wrong in a string of 300 amino acids, but it can be a huge deal. Wrong amino acids in key places of proteins can lead to disease, the most recognizable one being cancer.
I’m now going to use p53 as an example to explain how a single amino acid change can have such deadly consequences.
Cells are constantly growing and dividing. A series of checkpoints exist to ensure that the cell is preparing itself appropriately to divide into two cells. This is called the cell cycle (Figure 11.1). It is comprised of four phases: G1 = growth, S = synthesis, G2 = growth, M = mitosis. In the growth phases, the cell is getting bigger so that when it splits in half, the two daughter cells are not incredibly small. The S phase is when the entire DNA molecule is faithfully copied so that two exact genomes exist in the cell. The M phase splits the cell down the middle and ensures that each new cell gets one complete copy of DNA. This entire process is highly regulated (isn’t everything in a cell?). If the cell does not go through all the steps correctly, it is p53’s job to stop that cell from dividing by either arresting its growth or simply killing it. Better to kill a poorly run cell than allow it to split into two poorly run cells.
Cancer is defined as unregulated cell growth. Cancerous cells (also called transformed cells) don’t give a crap about the cell cycle and all its checkpoints. They just want to grow, divide, keep moving and wreak havoc. Who cares if the DNA is copied well? Let’s just go. Oxygen hasn’t come along in awhile? Let’s just build a blood vessel here to carry oxygen to us! Who cares that this human’s body wouldn’t normally grow a blood vessel here? Go, go, go!
It doesn’t take a genius to realize that one of the first proteins knocked in cancer is p53. Remove the roadblock to growth and division quickly and early on. Imagine that one cell loses p53, then it divides into two cells. Those two new cells don’t have p53 and they each divide into two more cells without p53. On and on… it is exponential growth of cells without p53. Any cell without p53 can’t stop growing, can’t fix DNA problems, can’t do anything but g-r-o-w. These transformed cells are factories gone completely off the rails (bakeries selling dough or everlasting gobstoppers laced with poison!) and are difficult to stop. But, I’m sure most people appreciate the difficulty of stopping cancer growth.
So how do you knock out p53’s function? I told you earlier that in order for p53 to tell the cell to stop growth or just die, it needs to be able to bind DNA.
A landmark paper was published in 1994 by Nikola Pavletich’s laboratory. It was the tertiary structure of p53 bound to DNA. This work was able to show which amino acids in p53 were necessary for proper DNA binding. Just as fingers need to be placed correctly on a baseball to throw curves, fastballs, or sliders… (Go Phillies!), the correct amino acids of p53 must be in the correct places to grab DNA and hold on.
The paper went on to look at various amino acid mutations of p53 found in cancerous cells. Figure 11.2 is directly from their Science article. The p53 primary sequence lies along the X axis and the number of times a particular amino acid is mutated in cancer is represented by the vertical black bar. The larger the bar, the more often that particular amino acid is found mutated in cancer.
Before this paper, no one could exactly explain why the amino acid arginine was so essential at position 273. But like fingers gripping a baseball, this structure showed that arginine was necessary for p53 to grab the DNA properly. Another amino acid simply wouldn’t do. Imagine trying to throw a knuckleball without your index finger. It would be difficult or nearly impossible. Sure, you still have a hand, but if the hand’s job is to throw knuckleballs, it’s going to need that index finger in the correct place. A p53 without certain amino acids is crippled in its function in much the same way. It can’t grab hold of the DNA properly and thus it can’t tell the cell to stop growing. Without this protein working properly, a sick cell continues to grow, continues to divide, and can, in time, lead to a tumor.
How does a cell obtain a mutation in their p53 protein to begin with? It can happen at the level of DNA, RNA or protein. The ribosome could simply incorporate the wrong amino acid. It happens. Or maybe the RNA was not transcribed faithfully from the DNA. However, so many p53s are floating around the cell that one wrong one will quickly be compensated for by all the correct ones. Proteins are also degraded after a certain period of time and replaced with freshly made proteins. This turnover will also dilute out any small errors in transcription or translation.
However, DNA mutations are a bigger deal. In the previous two cases, the DNA was correct so going back to the beginning fixes the problem. But, what if the beginning is wrong to begin with? We pick up DNA mutations throughout our entire lives. A mutation in the p53 gene will be carried on to daughter cells, which will make incorrect p53. We also could have inherited a mutated copy of p53 from our parents. All our cells from the very beginning could have 50% of its p53 proteins wrong. It’s hard to fix mutated DNA or incorrect p53 molecules, but many scientists are working on that very type of research.
This topic will lead us down a fruitful road of learning about cancer, cancer therapies, new avenues of research, and so much more about p53. We’ll get there all in good time.
Cell cycle – the organized growth and division of cells
Cell cycle arrest – the pausing of the cell cycle while repairs are made inside the cell
Apoptosis – organized cell death; the process a cell undergoes when it needs to commit suicide
Transformed cells – term used to describe cells that have gained cancerous characteristics
REFERENCES
Dahl, Roald. “Charley and the Chocolate Factory.” (1964) Penguin Group, New York, New York.
Stuart, Mel. “Willy Wonka and the Chocolate Factory.” (1971)
Suryadinata, R. et al. “Control of cell cycle progression by phosphorylation of cyclin-dependent kinase (CDK) substrates.” (2010) Bioscience Reports 30, pgs 243 – 255.
Cho et al. “Crystal Structure of a p53 Tumor Suppressor-DNA Complex: Understanding Tumorigenic Mutations.” (1994) Science 265, pgs 346 – 355.
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