There are at least two main mechanisms for change in copy number: NAHR and microhomology-mediated events. NAHR can be formed either by classical HR-mediated DSB repair via a double Holliday junction, or from BIR, which restarts broken replication forks by HR. However, the LCRs that mediate NAHR were presumably formed predominantly by the same mechanism as non-recurrent copy-number changes that are being formed now. Thus the mechanisms of microhomology-mediated copy number change underlie most copy-number change. Based on the evidence favouring replicative mechanisms, the known enzymology of DNA transactions in model organisms, and the evidence presented above concerning the potential involvement of stress responses in altering the availability of DNA repair proteins, we suggest that a mechanism like MMBIR presently constitutes our best working hypothesis for most events of copy number change. It is also likely that end-joining mechanisms including NHEJ, MMEJ and SSA will play a role, especially in cells of the immune system. The breakage-fusion-bridge cycle has been shown to operate in experimental systems and appears to be important in amplification in some cancers. However, we need not assume that only one mechanism acts in any given event. Microhomology-mediated events might trigger the breakage-fusion-bridge cycle by forming a dicentric chromosome, which must eventually be resolved to a more stable genotype. This could also happen when NAHR causes formation of a dicentric chromosome. Similarly, end-joining mechanisms might have a role to play in cleaning up loose ends that result from other events. Notably, the fusion step of the breakage-fusion-bridge cycle might be mediated by any of these end-joining mechanisms.
The molecular events proposed in MMBIR have not been demonstrated experimentally. Because of its molecular detail, several aspects of the model are testable. The hypothesized involvement of stress responses will also be important to test. The potential for extensive LOH downstream from the initiating event has already been seen in some systems, and further testing of this correlation should be available from the study of genome-wide single nucleotide polymorphism data. This LOH might extend as far as the next replication fork travelling in the opposite direction, or it might process to the telomere.
If it can be substantiated that CNV stems from stress response, this has interesting implications for physiology, evolution and disease. First, stress-inducible chromosomal structural variation suggests that cells have an inducible ability to evolve (“evolvability”). If the mechanism is indeed stress-inducible, then cells and organisms will be predisposed to genome rearrangement when they are stressed and activate stress responses. This is specifically when they are maladapted to their environments. If stress fuels CNV formation, then the generation of genetic diversity upon which natural selection acts will be maximal specifically when a population will benefit most from such diversity, potentially fueling evolution at that time. This idea was developed to explain mutagenesis affecting evolution of bacterial populations (reviewed by129, 130
) including generation of antibiotic resistance upon antibiotic exposure131, 132
. If stress inducibility applies to CNV formation, then CNVs may not only be important promoters of evolutionary divergence12
but, in addition, their formation may be an “evolvability” enhancing mechanism. Although human cells show stress-inducible genetic change109, 133, 134
, it is not yet known in any organism with a differentiated germline whether such stress-inducible genome instability mechanisms can contribute to the germline and so to evolution. This is an important topic for future study.
Second, similarly, the predicted occurrence of LOH with CNV formation is likely to be important in human cancer in which both mutations and LOH drive tumor progression and resistance to therapies. The same mechanism is proposed to produce translocations when the microhomology used in repair resides in a different chromosome24
, further exacerbating the problem. The problem of stress-inducible cancer progression and resistance mechanisms are discussed elsewhere108, 129
Third, given the large and ever increasing number of developmental, neurological, and psychiatric syndromes linked to CNV, we wish to propose an important analogy or corollary from cancer biology to these other CNV-based diseases. It is well known that mutations that increase mutation rate, “mutator mutations”, promote cancer predisposition because cancer is a genetic disease fueled by mutagenesis and genome instability135-137
. Mutator mutations are common in microbes and many are known in a wide variety of cancer-predisposition and aging syndromes25
. We propose that in addition to CNVs' directly promoting various syndromes, that there will be human variants with increased rates of CNV change, because of mutations in any of many possible genes affecting DNA-damage processing and repair (among other processes). We expect that such alleles will predispose families and individuals to the mental, developmental, neurological and other syndromes caused by CNVs. Future work should seek to identify such modifier-locus mutations that affect human developmental and other disorders, some of which, we predict, will do so by elevating CNV-formation rates. Individuals with such mutations might also be cancer prone. Moreover, in cases where the disease can be caused by a CNV present as a mosaic, therapies to reduce CNV genesis might reduce penetrance of such disease. Drugs that do this are neither known nor on the drawing board. Identification of the many genes, proteins and pathways affecting CNV formation rates and understanding their mechanisms of action are prerequisites to considering any potential therapeutic approaches.