The proteasome is a large protein complex responsible for degradation of intracellular proteins and is crucial for maintaining homeostasis of protein levels within a cell. Studying the proteasome’s activity, and how this changes rates of protein degradation, can reveal information that is lost when simply looking at the proteome.
At the Festival of Genomics and Biodata in January 2022, Yifat Merbl (Group Leader, Immunology Lab at the Weizmann Institute of Science) detailed how her team have been rummaging through the “garbage can” of the cell to provide ground-breaking insights into cancer therapy. You can watch Yifat’s presentation in full here.
Yifat highlighted a lack of investigative ability and precision when considering the EpiProteome, which describes the set of all post-translational modifications (PTMs) made to proteins. She also made sure to emphasise the critical role that proteasomes play in anti-tumour immunity through influencing the degradation landscape of a cell (degradome). The degradome can be changed by the presence of proteasome regulatory caps, and different catalytic cores. Changes to these regulatory elements can have a big impact on the levels of protein degradation within a cell.
Current proteomic analysis focuses on bulk analysis of the protein content of a cell at any given time. However, what it does not do is convey how proteins are being processed by the cell.
To answer this, Yifat and her team developed MAPP – Mass Spectrometry Analysis of Proteolytic Peptides – which essentially sequences any peptides that have been sent to be degraded by the cell. Additionally, MAPP can better detect peptides with a higher turnover rate than traditional proteomics.
This is highly applicable for research as MAPP can identify patterns of degradation in particular diseases, as well as specific cleavage sites. Knowing these sites allows researchers to identify which proteasome (e.g., the constitutive proteasome or the immuno-proteasome) is associated with the degradation of particular peptides.
Implications in lung cancer
The disease area which Yifat’s team focused on was lung cancer. Lung cancer has the highest incidence and mortality worldwide, with very limited therapy options. Proteasome footprinting with MAPP in tumour tissue and adjacent tissue revealed tumour specific degradation targets. To identify in greater detail the mechanism by which degradation is altered in tumour tissue, specific subunits of the proteasome were analysed. It was found that the regulatory cap PSME4 was significantly upregulated in the tumour cells when compared to adjacent tissue.
In collaboration with Silke Meiner’s lab in Helmotz, it was found that patient survival in individuals with high PSME4 levels was significantly lower than those with low levels of PSME4. Nonetheless, there was very little in the literature to support a causal link, and the role of PSME4 in cancer was poorly described.
The next phase of research involved figuring out exactly how the PSME4 regulatory subunit was influencing the proteasome. Yifat’s team found that PSME4 associated with both the constitutive and the immuno-proteasome but affected them in different ways. Increased PSME4 upregulated the constitutive proteasome which induced caspase-like cleavage and downregulated the immuno-proteasome, which normally induces tryptic-like cleavage that is more favourable for MHC1 presentation. These changes meant that antigen presentation for the immune response was negatively affected.
Improving immune therapy response
In vivo mouse models of PSME4 knock-down, wild-type, and overexpression showed that a lower level of PSME4 resulted in a decreased tumour burden. This was shown through both tumour size and abundance, as well as through monitoring levels of T-cells and cytotoxic T-cells as a representation of immune response. The increase in immune response was verified separately using trained splenocytes that were targeted to the lung tumour cells. PSME4-knockdown mice experienced much higher levels of targeted tumour cell death than those with overexpressed PSME4.
Until this point, the research on the effects of altered peptide degradation and PSME4 expression had been conducted with lung cancer in mind. Using a meta-analysis published in Cell in 2021, Yifat and her team expanded the scope of the study to examine the effects of PSME4 in bladder cancer, renal cancer, and melanoma. When looking at the percentage response to immunotherapy, they found that there was a significantly lower response in all cancer types in patients with high levels of PSME4.
Yifat highlighted that this work demonstrates how MAPP can be used as a novel drug discovery platform by utilising the understudied area of the degradome in cancer. It could also provide another tool for precision oncologists to deliver immunotherapy to the patients that will benefit from it most.
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