610 Medizin und Gesundheit
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The transcription factor MYC is a onco-protein, found to be deregulated in many human cancers. High MYC levels correlate with an aggressive tumor outcome and poor survival rates. Despite MYC being discovered as an oncogene already in the 1970s, how MYC regulates transcription of its target genes, which are involved in cellular growth and proliferation, is not fully understood yet.
In this study, the question how MYC influences factors interacting with the RNA polymerase II ensuring productive transcription of its target genes was addressed using quantitative mass spectrometry. By comparing the interactome of RNA polymerase II under varying MYC levels, several potential factors involved in transcriptional elongation were identified. Furthermore, the question which of those factors interact with MYC was answered by employing quantitative mass spectrometry of MYC itself. Thereby, the direct interaction of MYC with the transcription elongation factor SPT5, a subunit of the DRB-sensitivity inducing factor, was discovered and analyzed in greater detail. SPT5 was shown to be recruited to chromatin by MYC. In addition, the interaction site of MYC on SPT5 was narrowed down to its evolutionary conserved NGN-domain, which is the known binding site for SPT4, the earlier characterized second subunit of the DRB-sensitivity inducing factor. This finding suggests a model in which MYC and SPT4 compete for binding the NGN-domain of SPT5.
Investigations of the SPT5-interacting region on MYC showed binding of SPT5 to MYC’s N-terminus including MYC-boxes 0, I and II.
In order to analyze proteins interacting specifically with the N-terminal region of MYC, a truncated MYC-mutant was used for quantitative mass spectrometric analysis uncovering reduced binding for several proteins including the well-known interactor TRRAP and TRRAP-associated complexes.
Summarized, ...
The transcription factor SPT5 physically interacts with MYC oncoproteins and is essential for efficient transcriptional activation of MYC targets in cultured cells. Here, we use Drosophila to address the relevance of this interaction in a living organism. Spt5 displays moderate synergy with Myc in fast proliferating young imaginal disc cells. During later development, Spt5-knockdown has no detectable consequences on its own, but strongly enhances eye defects caused by Myc overexpression. Similarly, Spt5-knockdown in larval type 2 neuroblasts has only mild effects on brain development and survival of control flies, but dramatically shrinks the volumes of experimentally induced neuroblast tumors and significantly extends the lifespan of tumor-bearing animals. This beneficial effect is still observed when Spt5 is knocked down systemically and after tumor initiation, highlighting SPT5 as a potential drug target in human oncology.
Pancreatic ductal adenocarcinoma (PDAC) is predominantly driven by mutations in KRAS and TP53. However, PDAC tumors display deregulated levels of MYC and are a paradigm example for MYC-driven and -addicted tumors. For many years MYC was described as a transcription factor that regulates a pleiotropic number of genes to drive proliferation. Recent work sheds a different light on MYC biology. First, changes in gene expression that come along with the activation of MYC are mild and MYC seems to act more as a factor that reduces stress and increases resilience towards challenges during transcription. Second, MYC is a strong driver of immune evasion in different entities. In this study we depleted MYC in murine PDAC cells and revealed the immune dependent regression of tumors in an orthotope transplant model, as well as the activation of the innate immune system using global expression analysis, immunoblotting and fCLIP.
These experiments revealed that endogenous double-stranded RNA is binding as a viral mimicry to Toll-like receptor 3, causing activation of TBK1 and downstream activation of a proimmunogenic transcription program. The regression of tumors upon depletion of MYC is dependent on this pathway since the knockout of TBK1 prevents regression of tumors after depletion of MYC.
We can summarize this study in three main findings: First, the dominant and most important function of MYC in tumors is not to drive proliferation but to promote immune evasion and prevent immune-dependent regression of tumors. Second, cells monitor defects or delay in splicing and RNA processing and activate the immune system to clear cells that face problems with co-transcriptional processing. Third, MYC suppresses the activation of the cell-intrinsic innate immune system and shields highly proliferating cells from the recognition by the immune system.
To translate this into a therapeutically approach, we replaced the shRNA mediated depletion of MYC by treatment with cardiac glycosides. Upon treatment with cardiac glycosides tumor cells reduce uptake of nutrients, causing a downregulation of MYC translation, inhibition of proliferation, glycolysis and lactate secretion. Lactate is a major reason for immune evasion in solid tumors since it dampens, amongst others, cytotoxic T cells and promotes regulatory T cells.
Treatment of mice with cardiac glycosides causes a complete and immune-dependent remission of PDAC tumors in vivo, pointing out that cardiac glycosides have strong proimmunogenic, anti-cancer effects. More detailed analyses will be needed to dissect the full mechanism how cardiac glycosides act on MYC translation and immune evasion in PDAC tumors.