Archives for posts with tag: Centromere

Cancer has been described as the most common genetic disease. This doesn’t necessarily mean it’s hereditary – usually the genetic mistakes that cause cancer arise in the body’s organs or tissues and can’t be inherited. We’re continually learning of new cancer-causing genetic mistakes.

If we think of genes as words spelling out the instructions for our bodies to function, there are different types of mistakes or “typos” that can cause cancer. Some of these are like spelling mistakes – an incorrect letter or two. The mistakes I’ll be talking about here involve whole words (or lots of them). For example one or more copies of a word are added – ” very big” becomes “very very big” – extra copies of a cancer gene (we call them oncogenes) can cause or accelerate cancer growth. Or if a word is lost – “don’t grow” becomes “grow” – this illustrates loss of a tumour suppressor gene.

I described the breakage-fusion-bridge cycle a few weeks back. The BFB cycle was a theory developed from studies with maize, but it also applies to some cancers. This is an example of basic research,  inspired by curiosity but eventually being useful in ways we never imagined. If you look back at that post it shows how the BFB cycle can cause gain or loss of genes. If a cancer gene is  multiplied, or if a tumour suppressor gene is in the part that’s lost, the cell can gain a growth advantage over other cells, which is part of the process causing cancer.

Here’s an example.

Cancer cell lines are cancer cells that can be grown indefinitely in the laboratory. I’ve just published a paper on HEL, which is a leukaemia cell line. It’s popular for studying how cells make globin (molecules in red blood cells that help us process the air we breathe).

The chromosomes of HEL are very abnormal. I’ve used a combination of techniques to show how the chromosomes are reorganised as well as which parts have been lost, gained and amplified.  It’s very complicated.

One of the gene abnormalities in HEL is amplification of the JAK2 gene. JAK2 is a well-known cancer gene that is often abnormal in blood cancers. The normal gene can be mutated to become a cancer gene, for example by a “spelling mistake” in the DNA. By adding extra copies of this abnormal gene the effects can be magnified. This is known as gene amplification. There are a few cancer genes that are commonly amplified in cancers.

To cut a long story short, JAK2 is amplified in the HEL cell line. And a nearby tumour suppressor gene (CDKN2A) has been lost. But only by looking at the chromosomes does the reason become clear. Some detective work tells us that there were some breakage-fusion-bridge events. I won’t go into the detail – if you’re interested it’s in the paper. But we have chromosomes whose ancestors had two centromeres, and if we use a DNA tag for the region between the centromeres we can see “stripes”.

Here’s an example from HEL that shows DNA amplified by the BFB cycle – we can show where a gene is on the chromosomes by labelling it with a fluorescence tagged DNA “probe”. The striped pattern reminds us of the yellow dots in the modelling clay demonstration:

The red is DNA that's normally at one end of some of the chromosomes. The stripes tell us that the end of a chromosome (22) is in the middle of these chromosomes and there are extra copies. This helps us work out how these chromosomes were made. It's a strong clue that BFB cycles were involved and the ancestral chromosome had two centromeres.

The red is DNA that’s normally at one end of some of the chromosomes. The stripes tell us that the end of a chromosome (22) is in the middle of these chromosomes and there are extra copies. It’s a strong clue that BFB cycles were involved and the ancestral chromosome had two centromeres.

The new chromosome with four copies of the yellow gene courtesy of the breakage-fusion-bridge cycle.

The new chromosome with four copies of the yellow gene courtesy of the breakage-fusion-bridge cycle.

Recently JAK2 amplification was also reported in triple-negative breast cancer. Triple negative means that three well-known genetic causes of breast cancer are not present. So finding JAK2 amplification would help explain the cause of some triple-negative breast cancers, and could help work out an effective treatment. Perhaps this JAK2 amplification is sometimes caused by BFB cycles. Without looking at the layout of the abnormal chromosomes we may never know.

To end this story, here’s Bruce Mercer’s cartoon diagram showing the BFB cycle in HEL: http://emph.oxfordjournals.org/content/2013/1/225/F6.expansion.html

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Barbara McClintock published a paper describing the breakage-fusion-bridge (BFB) cycle in 1939. Many of her ideas were well before their time. Like many such profound leaps in thinking, the BFB cycle took a long time to catch on. She wrote in 1973, “I stopped publishing detailed reports long ago when I realized, and acutely, the extent of disinterest and lack of confidence in the conclusions I was drawing …One must await the right time for conceptual change.” Her work was appreciated much later and she was awarded a Nobel Prize in 1983 for her discovery of “jumping genes“.

A few weeks back I introduced this post by describing normal chromosome division. This time we’ll look at the breakage-fusion-bridge cycle. This is one way chromosome division can go wrong. Very wrong, in the sense that it can cause the chromosomes to keep changing, and this can cause cancer.

A human chromosome with two centromeres is abnormal. Chromosomes with two centromeres are not unusual in cancer cells. In fact they’re probably a lot more common than we think, because in both research and diagnostic labs the centromeres are usually not looked at.

To recap, a normal chromosome has one centromere. Before the chromosome divides, the two identical halves (chromatids) are held together at the centromere. When the chromosome divides the centromere splits into two halves, the chromatids become the new chromosomes, and the centromeres take the two new chromosomes in different directions into the two new daughter cells.

So what happens if there are two centromeres? If they’re both aligned so that they head in the same direction it’s not a problem – together they take a complete new chromosome with them. The closer the centromeres are together the more likely this is.

Now follow the pictures and their captions. These describe chromosome division in an abnormal chromosome with two centromeres. Especially follow the yellow dots.

A twist between the two centromeres when the chromosomes align ready for chromosome division.

If the two centromeres on a chromosome go in the same direction there’s no problem. But if there’s a twist between the two centromeres when the chromosomes align ready for chromosome division….

....then, when the two halves of each centromere separate they head off in different directions.

….then, when the two halves of each centromere separate they go in opposite directions. We have a “bridge” spanning the gap between the two centromeres.

The bit of chromosome between them gets stretched and can break.

The bridge is stretched and can break.

The broken chromosomes in the new cell join together - the top daughter cell gets an extra copy of the yellow gene. The bottom cell loses this copy of yellow gene.

The broken chromosomes in the new cell join together – the top daughter cell gets an extra copy of the yellow gene. The bottom cell loses this copy of the yellow gene.

The new chromosome copies itself to make two equal halves.

The new chromosome copies itself to make two equal halves.

If this process repeats..

If this process repeats..

Fusion of the broken bits of chromosome in the top cell.

Fusion of the broken pieces creates a chromosome with four copies of the yellow gene.

After replication - the new chromosome with four copies of the yellow gene courtesy of the breakage-fusion-bridge cycle.

After replication.

If the yellow gene in the pictures is a cancer gene (“oncogene”) the cell with extra copies might grow and multiply faster than its neighbours. We call this natural selection – the cells that can grow faster than their neighbours become more common which means the genetic change causing that is undergoing “positive selection”. Yes, the cells in our body can evolve and we know this best as cancer.

All this change happens between the two centromeres where the bridge forms. So if we find a chromosome with this type of change on one side of the centromere only it’s a clue that this might have been caused by the breakage-fusion-bridge cycle.

These are modelling clay images from my breakage-fusion-bridge claymation. They’re a bit rough but I hope it helps you understand what happens. Many, perhaps most, images demonstrating the BFB cycle show a different version – where the abnormal chromosome is created by two chromatids of one chromosome breaking and joining together. Most examples don’t show the version I’ve presented – where two different chromosomes have joined together. Check this out on Google Images (search for breakage-fusion-bridge).

Here’s the answer to the quiz from the telomere post. The arrows point to the ring chromosomes. Being rings they have no ends, so no telomeres.

answer to ring chr

Further Reading:

B. McClintock 1939. The Behavior in Successive Nuclear Divisions of a Chromosome Broken at Meiosis. Proc Natl Acad Sci U S A. 1939 August; 25(8): 405–416.

M. Kinsella and V. Bafna 2012. Combinatorics of the Breakage-Fusion-Bridge Mechanism. J Comput Biol. 2012 June; 19(6): 662–678.

R. MacKinnon and L. Campbell 2011. The Role of Dicentric Chromosome Formation and Secondary Centromere Deletion in the Evolution of Myeloid Malignancy. Genetics Research InternationalVolume 2011 (2011), Article ID 643628.

What makes a cell become cancerous?

This can happen if the chromosomes are incorrectly distributed to a new cell. The cell could get too many copies of a cancer-promoting gene, or too few copies of a cancer-protecting gene.

The breakage-fusion-bridge cycle is one way that this can happen. This  week I was asked to provide a cartoon showing the breakage-fusion-bridge cycle and how it relates to a chromosome abnormality I was describing.

By lucky coincidence my colleague Lan Ta just this week published a paper with a neat breakage-fusion-bridge cartoon that was put together by Bruce Mercer. As well as being a scientist, Bruce is a graphic artist – a very handy combination. So I was trying to draw the modified cartoon for Bruce in two dimensions, without much success. Then modelling clay came to the rescue and I was able to show him what I meant.

So I would like to share the 3D modelling clay version, but before we look at the breakage-fusion-bridge cycle, which is an abnormal pattern of chromosome division, we had better look at normal chromosome division (or mitosis).

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These are chromosomes in modelling clay. There are 23 pairs of chromosomes in a human cell but we will follow 3 chromosomes for simplicity.  The chromosomes only take on a recognisable shape when the cell is ready to divide. Each chromosome is made of two halves called chromatids, which are identical. These are held together at the centromere.

mitosis 1

In a cell the chromosomes are usually not recognisable – the DNA is unravelled in the nucleus. In a growing cell each unravelled  chromosome is producing a copy of itself so that there will be a chromosome for each of the two new daughter cells when the cell divides, or reproduces itself.

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After the chromosomes take shape they line up together (at the metaphase plate) between two ends, or poles, of the cell. The centromere has another very important function in cell division. It attaches to fine fibres (microtubules) which stretch between the poles.

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The two halves of the centromere separate and each draws its chromatid (which is now a new daughter chromosome) along these microtubules.

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So when the cell divides in two to make two new cells, each chromatid becomes a chromosome in one of the new cells.

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Cell division complete, the chromosomes unravel and copy themselves again ready for the next cell division.

This process happens successfully millions of times every day to create new cells in our bodies – amazing.

In the year 2000 the draft human genome sequence was announced by Tony Blair and Bill Clinton. It was said to be complete in 2003, in time for the 50th anniversary of the discovery of the structure of DNA. Well actually it wasn’t quite finished. Actually it’s still not finished. Besides the tweaking that still goes on here and there, there are still big gaps. And what’s in these gaps sometimes has a significant role in cancer.

The biggest gaps are centromeres. Centromeres and telomeres are made of what is known as repetitive DNA, and this is hard to sequence. There’s more detail below.

Cancer chromosomes often have centromere or telomere abnormalities. In fact these abnormalities can cause cancer or make it progress faster.

In cancer research there’s a big push to sequence the genomes of different types of cancer  to try and understand the many different DNA changes that can cause cancer. Some researchers try to understand telomeres and centromeres and their role in cancer, but in cancer sequencing projects, and also in diagnostics, centromeres and telomeres are pretty much ignored. Although they’re difficult to sequence, the repetitive DNA does make them easy to study by some other techniques.

One of the goals of personalised medicine is to be able to read a person’s complete genome. For cancer this would include the abnormal cancer genome. But at the moment these gaps mean that we can’t describe the abnormal cancer chromosomes from end to end by sequencing them. The approach I use allows me to work out what’s in each chromosome and discover telomere and centromere abnormalities.

HOW DOES MY RESEARCH FIT IN?

By looking at things that most people don’t worry about I’ve overturned a few assumptions and made some unexpected discoveries, particularly about centromeres in leukaemia.

A normal chromosome has one centromere. I found that chromosomes with two centromeres are more common in  acute myeloid leukaemia (AML) and myelodysplastic syndromes (MDS) than was thought. I found that most of these chromosomes with two centromeres were probably made by two chromosomes joining together.

Telomeres are at the ends of chromosomes and stop them from sticking to each other, so when the telomeres are eroded,  chromosomes can join together. They can be eroded by exposure to chemical toxins, cancer drugs and radiation. So it’s interesting that the leukaemias that are caused by these exposures have more of these two-centromered chromosomes than other leukaemias.

centromeres - dicentric

Fluorescent DNA probes can label up centromeres (blue) and genes (red). In this image there is also a chromosome 20 paint – the green regions are from chromosome 20.

MORE DETAIL ON THE SCIENCE

Telomeres and centromeres are made of highly repetitive DNA and make up some of the gaps in the human genome sequence.

Sequencing a genome is like reading a story. But first we cut the book up into tiny fragments. We read them piece by piece, then try to join the pieces together to make the story, by matching the overlapping parts. Where this approach falls apart is that some sections are repetitive. Some pages are made up of a single word or phrase repeated over and over and over and over and over and over (I won’t repeat that hundreds of thousands of times, but you should get the picture). So if a lot of fragments just say the same thing “over and over and over”, it’s very hard to put them together meaningfully.

The centromere guides the chromosome to the two daughter cells during cell division. A normal chromosome has one centromere. When a chromosome has two centromeres (we call this a dicentric chromosome), the chromosome can be pulled in opposite directions, breaking the chromosome and causing more chromosome disorganisation.Telomeres cap the ends of normal healthy chromosomes. One of their functions is to stop the chromosomes from sticking to each other. So when the telomeres are lost or eroded the chromosomes can join together. That’s one way of creating a chromosome with two centromeres.

Telomere loss is a natural part of ageing. There are also many environmental and lifestyle factors that are thought to affect telomere length. Short telomeres are thought to be a cancer risk because dicentric chromosomes are more likely to arise.

Telomeres and centromeres are very important parts of a normal chromosome. You could say they hold the chromosome together. They have a lot of influence on whether chromosomes are normal and stay normal.

FURTHER READING

Murnane JP 2012. Telomere dysfunction and chromosome instability. Mutat Res. 2012 Feb 1;730(1-2):28-36. (Open access)

MacKinnon RN and Campbell, LJ. 2011. The role of dicentric chromosome formation and secondary centromere deletion in the evolution of myeloid malignancy. Genetics Research International. Article ID 643628. (Open access)

MacKinnon RN, Duivenvoorden HM and Campbell LJ. 2011. Unbalanced translocation of 20q in AML and MDS often involves interstitial rather than terminal deletion of 20q. Cancer Genet. 204(3):153-61.