APOBEC3 proteins restrict mobile and foreign DNA

Abstract

A cell's DNA provides its operating instructions. Consequently, cells rigorously maintain their DNA. At the same time, cells must prevent foreign DNA from persisting, or risk having their operations hijacked. This thesis describes how APOBEC3 proteins help fulfill this double imperative, by limiting the replication of mobile genetic elements, and by triggering the destabilization of foreign intracellular DNA.

The APOBEC3s are a family of DNA modifying enzymes that convert deoxycytidines to deoxyuridines. Introducing uridines into DNA can alter the sequence of genes or regulatory elements, in essence removing information content from the molecule. Such an alteration also often leads to the destabilization and degradation of the DNA. Previously, APOBEC3s had been shown to act on a relatively limited subset of viral and retrotransposon DNA replication intermediates. The major contribution of this work is to expand the range of biologically relevant APOBEC3 substrates to include LINE-1 retrotransposons and foreign intracellular DNA in general. LINE-1s are the only retrotransposons currently active in humans, and their mobilization can cause disease by a variety of mechanisms, the most straightforward of which is by inserting in or near a gene and disrupting its function. Expression of several APOBEC3 proteins reduces the rate of LINE-1 retrotransposition in human cells. Less retrotransposed LINE-1 DNA accumulates in these cells, suggesting that the block occurs prior to retrotransposon integration. The APOBEC3s therefore limit the mutagenic potential of LINE-1 elements. Foreign intracellular DNA is inherently dangerous and is often associated with microbial infection. In recognition of this danger, sensors within cells detect foreign DNA, but, previously, little was known about mechanisms that respond to DNA detection to mediate its clearance. This thesis demonstrates that APOBEC3 family members are induced following DNA detection and destabilize foreign DNA. This defense system is likely to have evolved to respond to intracellular microbial DNA, but it may also diminish the efficacy of processes such as genetic engineering and gene therapy. Finally, this thesis explores the molecular basis of the differential sub-cellular localization of the APOBEC3 proteins. Sequestering APOBEC3 proteins in different cellular compartments may be a way to regulate the DNA substrates to which they have access. This thesis therefore offers several insights into a family of proteins that modify potentially harmful DNA in order to protect the well-being and proper function of cells.