ncRNAs have recently been implicated in Lamarckian transmission of inherited characteristics. In one example, fertilised mouse eggs were injected with a miRNA which targeted a key gene involved in growth of heart tissue. The mice which developed from these eggs had enlarged hearts (cardiac hypertrophy) suggesting that the early injection of the miRNA disturbed the normal developmental processes. Remarkably, the offspring of these mice also had a high frequency of cardiac hypertrophy. This was apparently because the abnormal expression of the miRNA was recreated during generation of sperm in these mice. There was no change in the DNA code of the mice, so this was a clear case of a miRNA driving epigenetic inheritance[157].
But if ncRNAs are so important for cellular function, surely we would expect to find that sometimes diseases are caused by problems with them. Shouldn’t there be lots of examples where defects in production or expression of ncRNAs lead to clinical disorders, aside from the imprinting or X inactivation conditions? Well, yes and no. Because these ncRNAs are predominantly regulatory molecules, acting in networks that are rich in compensatory mechanisms, defects may only have relatively subtle impacts. The problem this creates experimentally is that most genetic screens are good at detecting the major phenotypes caused by mutations in proteins, but may not be so useful for more subtle effects.
There is a small ncRNA called BC1 which is expressed in specific neurons in mice. When researchers at the University of Munster in Germany deleted this ncRNA, the mice seemed fine. But then the scientists moved the mutant animals from the very controlled laboratory setting into a more natural environment. Under these conditions, it became clear that the mutants were not the same as normal mice. They were reluctant to explore their surroundings and were anxious[158]. If they had simply been left in their cages, we would never have appreciated that loss of the BC1 ncRNA actually had a quite pronounced effect on behaviour. A clear case of what we see being dependent on how we look.
The impact of ncRNAs in clinical conditions is starting to come into focus, at least for a few examples. There is a breed of sheep called a Texel, and the kindest description would be that it’s chunky. The Texel is well known for having a lot of muscle, which is a good thing in an animal that’s being bred to be eaten. The muscularity of the breed has been shown to be at least partially due to a change in a miRNA binding site in the 3′ UTR of a specific gene. The protein coded for by this gene is called myostatin, and it normally slows down muscle growth[159]. The impact of the single base change is summarised in Figure 10.4. The final size of the Texel sheep has been exaggerated for clarity.
Figure 10.4 A single base change which is in a part of the myostatin gene that does not code for protein nevertheless has a dramatic impact on the phenotype in the Texel sheep breed. The presence of an A base instead of a G in the myostatin mRNA leads to binding of two specific miRNAs. This alters myostatin expression, resulting in sheep with very pronounced muscle growth.
Tourette’s syndrome is a neurodevelopmental disorder where the patient frequently suffers from involuntary convulsive movements (tics) which in some cases are associated with involuntary swearing. Two unrelated individuals with this disorder were shown to have the same single base change in the 3′ UTR of a gene called SLITRK1[160]. SLITRK1 appears to be required for neuronal development. The base change in the Tourette’s patients introduced a binding site for a short ncRNA called miR-189. This suggests that SLITRK1 expression may be abnormally down-regulated via such binding, at critical points in development. This alteration is only present in a few cases of Tourette’s but raises the tantalising suggestion that mis-regulation of miRNA binding sites in other neuronal genes may be involved in other patients.
Earlier in this chapter we encountered the theory that ncRNAs may have been vitally important for the development of increased brain complexity and sophistication in humans. If that is the case, we might predict that the brain would be particularly susceptible to defects in ncRNA activity and function. Indeed, the Tourette’s cases in the previous paragraph give an intriguing glimpse of such a scenario.
There is a condition in humans called DiGeorge syndrome in which a region of about 3,000,000 bases has been lost from one of the two copies of chromosome 22[161]. This region contains more than 25 genes. It’s probably not surprising that many different organ systems may be affected in patients with this condition, including genito-urinary, cardiovascular and skeletal. Forty per cent of DiGeorge patients suffer seizures and 25 per cent of adults with this condition develop schizophrenia. Mild to moderate mental retardation is also common. Different genes in the 3,000,000 base-pair region probably contribute to different aspects of the disorder. One of the genes is called DGCR8 and the DGCR8 protein is essential for the normal production of miRNAs. Genetically modified mice have been created with just one functional copy of Dgcr8. These mice develop cognitive problems, especially in learning and spatial processing[162]. This supports the idea that miRNA production may be important in neurological function.
We know that ncRNAs are important in the control of cellular pluripotency and cellular differentiation. It’s not much of a leap from that to hypothesise that miRNAs may be important in cancer. Cancer is classically a disease in which cells can keep proliferating. This has parallels with stem cells. Additionally, in cancer, the tumours often look relatively undifferentiated and disorganised under the microscope. This is in contrast to the fully differentiated and well-organised appearance of normal, healthy tissues. There is now a strong body of evidence that ncRNAs play a role in cancer. This role may involve either loss of selected miRNAs or over-expression of other miRNAs, as shown in Figure 10.5.
Figure 10.5 Decreased levels of certain types of microRNAs, or increased levels of others, may each ultimately have the same disruptive effect on gene expression. The end result may be increased expression of genes that drive cells into a highly proliferative state, increasing the likelihood of cancer development.
Chronic lymphocytic leukaemia is the commonest human leukaemia. Approximately 70 per cent of cases of this type of cancer[163] have lost the ncRNAs called miR-15a and miR-16-1. Cancer is a multi-step disease and a lot of things need to go wrong in an individual cell before it becomes cancerous. The fact that so many cases of this type of leukaemia, the most common human leukaemia, lacked these particular miRNAs suggested that loss of these sequences happened early in the development of the disease.
An example of the alternative mechanism – over-expression of miRNAs in cancer – is the case of the miR-17-92 cluster. This cluster is over-expressed in a range of cancers[164]. In fact, a considerable number of reports have now been published on abnormal expression of miRNAs in cancer[165]. In addition, a gene called TARBP2 is mutated in some inherited cancer conditions[166]. The TARBP2 protein is involved in normal processing of miRNAs. This strengthens the case for a role of miRNAs in the initiation and development of certain human cancers.
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For a useful review see Garzon et al. (2010),