Inhibiting Mitochondrial RNA Degradosome Complex SUV3 and PNPT1 Increases dsRNA in the Cytoplasm, Triggers Viral Mimicry Response and Sensitizes AML Cells to Immune Mediated Killing
Bibliographic record
Abstract
Eukaryotic cells have two separate genomes; nuclear DNA organized in chromosomes, and circular mitochondrial DNA located within mitochondria. Mitochondrial DNA is comprised of a double-stranded circular genome that is 16.6 kB in length, lacks introns, and encodes two rRNAs, 22 t-RNAs and 13 of the 90 proteins in the mitochondrial respiratory chain. To maintain homeostasis, mitochondria possess RNA degradation machinery to regulate mitochondrial RNA turnover. The ATP-dependent helicase, SUV3 (encoded by the gene SUPV3L1) and the exonuclease PNPase (encoded by the gene PNPT1) function in a complex to degrade mitochondrial dsRNA. We identified PNPT1 and SUPV3L1 through an unbiased in-silico screen using bioinformatics platform to identify novel targets in AML. Our analysis revealed a strong correlation between these genes and ontologies of exogenous dsRNA, response to viruses, and RNA catabolic processes. PNPase and SUV3 protein were increased in 7/7 AML patient samples and 10/10 of AML cell lines compared to the normal hematopoietic cells by immunoblotting. Analysis of the TARGET AML dataset revealed that AML patients with increased expression of SUPV3L1 (p = 0.051, p= 0.045) and PNPT1 (p = 0.0013, p = 0.018) had decreased overall survival and event-free survival respectively. We knocked down or knocked out PNPT1 and SUPV3L1 with shRNA or sgRNA in AML cells, to study the importance of these genes in AML. Genetic knockdown or knockout of PNPT1 or SUPV3L1 decreased the growth and viability of OCI-AML2, TEX, K562, U937, NB4 and 8227 leukemia cells. Moreover, SUPV3L1 & PNPT1 ranked top 5.2% and 7.4% of essential genes in 26 leukemia cell lines in CRISPR screens and 2.7% and 4.9% in RNAi screens (https://depmap.org/portal). Knockdown of PNPT1 & SUPV3L1 also reduced the clonogenic growth of AML cells. Demonstrating the functional importance of PNPT1 & SUPV3L1 on leukemia-initiating cells in vivo, genetic knockdown of PNPT1 and SUPV3L1 significantly reduced engraftment of TEX cells into the marrow of immune-deficient mice. Finally, primary AML cells with SUPV3L1 knockdown had reduced engraftment into the marrow of immune-deficient mice. Whereas, the knockdown of SUPV3L did not alter the primary engraftment of CD34+ enriched cord blood cells. Mechanistically, knockdown of PNPT1 and SUPV3L1 in OCI-AML2 cells increased levels of cytoplasmic dsRNA 3-4 fold compared to control. Knockdown of PNPT1 and SUPV3L1 also increased cytoplasmic dsRNA in 143B cells, but not Rho(0) 143B cells that lack mitochondrial DNA, demonstrating a mitochondrial source for the increased dsRNA. Increased cytoplasmic dsRNA can mimic viral infection and trigger a type 1 Interferon response. Knockdown of PNPT1 or SUPV3L1 increased expression of genes (INFgR1, ICAM, IRF7 & JAK/STAT) associated with a type 1 interferon response compared to control. We also observed that PNPT1 and SUPV3L1 knockdown in AML cells sensitized the leukemic cells to T-cell mediated killing upon co-culturing with double negative T cells. In addition, using an immunocompetent Balbc mouse model, we demonstrated endogenous T cells were required to observe the greatest reduction in tumor burden after SUPV3L1 knockdown. In summary, the mitochondrial RNA degradosome complex SUPV3L1 and PNPT1 are overexpressed in AML and are essential for AML cells and stem/progenitors. These enzymes regulate the levels of mitochondrial dsRNA and their inhibition leads to a viral mimicry response and heightened sensitivity to immune-mediated killing.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.000 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.001 |
| Insufficient payload (model declined to judge) | 0.001 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".