Damage to the DNA in our cells can have detrimental consequences. For example, DNA lesions can impede all DNA-transacting processes, including transcription and replication. Luckily most lesions are dealt with by the cellular DNA repair pathways. However, DNA repair pathways are not always accurate and, even worse, might alter the code of the DNA if they are not properly regulated. These activities may result in mutations (changes in the DNA code). As DNA encodes for proteins, these mutations might lead to differences in proteins. Both the order of amino acids as the the expression level of proteins (when for example the regulatory elements surrounding a gene are affected) might change. Currently it is unclear to what extend DNA damage impacts our proteomes nor how the cellular pathways that deal what protein stress (the protein quality control pathways) respond to DNA damage. My lab investigates this link between genome instability and protein quality control.
Genome instability influencing protein quality control
Our proteomes are maintained by the cellular protein quality control pathways that include both molecular chaperones and the cellular degradation machineries. This maintenance is necessary for nascent chains (the newly born polypeptides) but also to deal with other forms of proteotoxic stress that arise due to internal or external stimuli. As mentioned above the alteration of the genetic code will impact the proteome and may in fact cause proteotoxic stress as well. However, how somatic mutations affect our proteomes and which cellular pathways are dealing with this presumed link is currently unclear. In collaboration with the Kampinga group we are currently investigating this link in more detail.
Protein quality control influencing DNA repair
DNA can be damaged in many different ways as a result of exogenous and endogenous forms of cellular stress. DNA damage usually induces the activation of DNA repair pathways and often a cell cycle arrest (a check point response). This reaction, referred to as the DNA-damage response (DDR), is typically initiated by proteins that recognize DNA lesions, and is followed by the recruitment and activation of proteins that trigger checkpoint signaling or directly perform the necessary repair steps. When repair is completed the machinery needs to be disassembled and the DDR turned off. Therefore, the DDR is a tightly controlled process, which is regulated at multiple levels.
Over the years, it became clear that DNA repair is fine-tuned by an enormous amount of DNA damage-induced postranslational modifications. The main idea is that posttranslational modifications, including those of the ubiquitin family, lead to alterations in interactions between proteins that perform the necessary repair steps. DNA damage-induced posttranslational modifications are therefore a cellular way to assemble the DNA repair protein complexes required to counteract the genomic insults.
However, other regulatory mechanisms are also active. DNA repair complexes are highly dynamic and often abort during the reaction. The disassembly of DNA damage induced repair complexes must thus be highly orchestrated as well. In fact, there are indications that this is indeed the case. For example, upon binding to DNA lesions Rad23b dissociates from XPC (two proteins involved in nucleotide excision repair) in vivo3. This finding is surprising, as the formation of this complex is essential to initiate repair. Another example is how the chaperone-like segregase Cdc48/p97 seperates the DNA-bound Rad51 and Rad52 complex –two essential recombination/DNA repair enzymes. These findings were especially important since they pointed out that dissociation of DNA repair complexes is not necessarily spontaneous but in fact regulated. Moreover, this regulation occurs at the DNA damaged site.
The disassembly/remodelling when the reaction is done or stalled is under-investigated and therefore barely understood. The direct need for more fundamental insight becomes urgent with for example the newest insights in the extreme genome instability in tumors. This instability is partly caused by the uncontrolled reactions of the various DNA repair pathways (e.g. homologous recombination and non-homologous end joining). Our aim is to investigate how DNA repair complexes are disassembled at the DNA.