The answer lies in a self-seeding paradigm, whereby once CENP-A is deposited it continues to direct the maintenance of its own position, and to ensure that this is passed on to all daughter cells.{97} This is independent of DNA sequence. Instead it seems to depend on small chemical modifications to the histone octamers.
Histone proteins in the octamers can be modified in a huge number of different ways. Proteins are made up of combinations of 20 different amino acids, many of which can be modified. And there are lots of different modifications that can be made to a protein. This is just as true of histones as of any other proteins.
In human centromeres, the octamers that contain CENP-A don’t have a complete monopoly. Instead, blocks of these octamers alternate with ones containing the standard histone protein, as shown in Figure 6.5. The standard octamers carry a very characteristic combination of chemical modifications. These in turn attract other proteins that bind to these modifications, part of whose function is to make sure these modifications are maintained.{98} This all acts to keep the octamers that contain CENP-A localised to the same region of the genome, and means that they only form at one position on the chromosome. This is probably why the junk DNA sequence at centromeres is so variable between species, even though it provides the geographical scaffold for one of the most fundamental processes in any cell.
Figure 6.5 The alternating pattern of standard and CENP-A histone octamers at the centromeres. For clarity, only small numbers of octamers are shown, whereas there are thousands present in the cell. Each circle represents an entire octamer.
The chemical modifications at the centromere also have the effect of keeping that region of the genome silent. Although there are recent data suggesting that there may be low-level expression of RNA from some centromeric regions, it’s very unclear if this has any functional significance. Essentially, the DNA at the centromeres has no real function except to be junk. It just acts as the regions where CENP-A and all its associated proteins can bind. That’s the only thing the cell needs from it. It’s better that it doesn’t have any other purpose, because that might be disrupted when the octamers containing CENP-A bind. That’s why this region of DNA has been able to change so much during evolution, because the sequence really doesn’t matter.
It might seem that there is still a missing stage in this. How does the CENP-A ‘know’ to bind to the right region of junk DNA in the first place? Because that tends to be how we all think, wanting to know what starts something off. But if we examine that assumption, we realise it leads us into a dead-end. Once again in this chapter we can invoke the lyricist Oscar Hammerstein, although this time in Austria rather than Siam/Thailand.
In The Sound of Music, Captain von Trapp and Maria sing that ‘Nothing comes from nothing. Nothing ever could’.{99}
How right they were.
Naked human DNA is a completely non-functional molecule. It does nothing at all, and certainly can’t direct the production of a new human being. It needs all the accessory information, such as the histones and their modifications, and it needs to be in a functioning cell. When the replicated chromosomes are separated and pulled to opposite ends of the cell, they each carry off some histone octamers in the correct positions, and with appropriate modifications. There are enough of these that they can act as the seed region to recreate the full picture of histones and modifications in the daughter cells. This is true not just of standard histone octamers, but also of the ones that contain CENP-A and thus show where the centromeres are formed. For these non-standard octamers, the regions of the CENP-A protein that contain different amino acids from the standard histones are important for attracting the appropriate proteins.{100}
This information — the chemical modifications — is even retained when eggs and sperm are produced.{101} The octamers that contain CENP-A stay in place when the egg and sperm fuse to form the one cell that will ultimately give rise to all the trillions of others in the human body. Our centromeres have been passed down through all of human evolution, and long before that in our distant ancestors, based on the position of the proteins, and not the DNA sequence to which they bind.
There are drugs that interfere with the way in which the spindle apparatus pulls the replicated chromosomes to opposite ends of the cells. The spindle apparatus is formed by the coming together of a large number of proteins, and these only combine at the time when a cell is ready to pull the chromosomes apart. A drug called paclitaxel works by making the spindle apparatus too stable, so that the complex of proteins can’t disaggregate.{102}
We can visualise why this is a bad thing for a cell by comparing the scenario with one of those fire engines that carries an extending ladder. It’s great that the ladder can be extended to rescue people from upper storeys of a burning building. But if the fire crew can’t get the ladder folded back down again after the emergency and have to drive around with it fully extended, it won’t be long before they have a pretty serious accident. The same happens in the cells treated with paclitaxel. Systems in the cell recognise that the spindle apparatus hasn’t been deactivated properly, and this triggers destruction of the cell. In the UK, paclitaxel is licensed for use in a number of cancers including non-small cell lung cancer, breast cancer and ovarian cancer.{103}
Paclitaxel is probably effective because cancer cells divide rapidly. By using a drug that targets cell division, it’s possible to kill the cancer cells at a higher rate than the normal body cells, which are not proliferating so quickly. But we also know that abnormal separation of chromosomes is itself a hallmark of many cancers.
If the separation of chromosomes goes wrong, one daughter cell may inherit both the ‘original’ chromosome and its replicate. The other daughter cell won’t inherit either. The first daughter cell will have one chromosome too many, the other daughter cell will have one too few. This situation, where the number of chromosomes is wrong, is known as aneuploidy. The word is derived from Greek. In this case, an means ‘not’, eu means ‘good’ and ploos means ‘-fold’ (as in ‘twofold’, ‘threefold’, etc.). In other words, it represents an unbalanced genomic state.
Astonishingly, about 90 per cent of solid tumours contain cells that are aneuploid, i.e. contain the wrong number of chromosomes.{104} The pattern of aneuploidy can be really complicated, as there is probably a strong degree of randomness to how the chromosomes are mis-segregated if the process is going wrong. In a single cancer cell there may be four copies of one chromosome, two copies of another and one copy of a third, or some other combination. Because of this variability, it’s very difficult to determine if the aneuploidy itself drives the cancer process, or if it’s just an innocent marker of the cancer status of the cells. The likelihood, because of the essentially random patterns of abnormal chromosome numbers, is that there’s probably a spectrum. Some cancer cells may develop combinations of chromosomes that drive cell proliferation faster. Other cells may have combinations with the opposite effect, and which may even trigger the cancer cell’s suicide system. And in some cells the combination may be ultimately neutral.{105}