Our cells possess an intrinsic ability to fight infection. This provides a crucial last line of defence when the innate and adaptive immune systems have failed to control a pathogen. In 2010 it was discovered that antibodies brought into the cell by invading viruses and bacteria are detected by a protein called TRIM21 (Mallery et al. 2010 PNAS). This activates a rapid and specific degradation response by the cell’s waste disposal system, the proteasome, that prevents infection. Though TRIM21 acts in a cell-autonomous manner, it can also alerts the wider immune system to the site of infection through the activation of inflammatory transcriptional networks (McEwan et al. 2013 Nat Immunol). Cytoplasmic antibodies therefore act as a potent danger signal to control viral infection (review McEwan 2016 Antibodies).
The specificity of TRIM21 is dictated solely by the antibody to which it has bound. Antibodies are widely used in molecular biology and an enormous diversity of antibodies targeting the majority of human proteins has been amassed by the research community. In 2017, we showed that by introducing antibodies into cell by electroporation or microinjection, we could promote the rapid and selective degradation of cellular proteins (Clift et al. 2017 Cell). Trim-Away represents the first broadly-applicable protein-level depletion technique that does not rely on prior genetic modification of the target protein. It therefore complements techniques that act on DNA (CRISPR/Cas9) and RNA (RNA interference) to enable functional studies of genes at the three fundamental levels of biological information.
Advantages of protein-level depletion
Gene editing and RNAi are enormously powerful as research tools in their ability to determine gene function. However, acute depletion at the protein-level can provide specific advantages in certain situations including:
- Long lived proteins Certain proteins are extremely long-lived, with a half-life of months or years. Depletion using genetic means is therefore challenging, especially when such proteins are essential for gene function.
- Genetic compensation Compensation is a common confounding issue when genes are depleted over long timescales. It is difficult to determine the extent of compensation and it may obscure the true function of a gene.
- Post-translational modifications RNAi and CRISPR are usually unable to specify depletion of post-translationally modified protein variants. Where antibodies against PTMs exist, Trim-Away may enable their selective depletion.
Using TRIM21 to target pathological proteins
The use of monoclonal antibodies has been revolutionary in modern medicine. For example adalimumab (Humira), which targets TNFα, has transformed the treatment of numerous auto-immune and inflammatory diseases. However, current approaches can only target proteins accessible to the extracellular space, leaving a sizeable proportion of the proteome out of reach. The enduring problem is an inability to introduce large biomolecules to the intracellular environment. We are taking two approaches to overcome these limitations:
- Antibodies may be naturally trafficked into cells in some specialised situations. We have recently shown that misfolded assemblies of tau, a protein that aggregates in Alzheimer’s disease, can import antibodies and activate the destructive capacity of TRIM21 (McEwan et al. 2017 PNAS).
- Cytoplasmically expressed antibody-like molecules are capable of eliciting specific protein degradation (Clift et al. 2017 PNAS).