Daigle, S. | bioRxiv Science

bioRxiv2026

Validation of tissue-specific RNAi systems in C. elegans reveals a converging role for polyubiquitin UBQ-1/UBC in vitellogenin metabolism and lifespan

Numerous studies in C. elegans have taken advantage of gene silencing RNAi libraries and tissue-specific RNAi systems to temporally and spatially understand gene function. Tissue-specific RNAi strains were created via tissue-specific functional expression of a key gene in the RNAi system in a presumably RNAi insensitive correspondingly mutated background. Here, we tested the level of RNAi insensitivity across different commonly used RNAi-defective mutants (rde-1 and sid-1) using lifespan analyses. Remarkably, while we found similarly important lifespan shortening in wild-type animals by silencing the polyubiquitin gene ubq-1 (UBC) at both 20{degrees}C and 25{degrees}C, we found wide and temperature-sensitive variations in RNAi sensitivity across different rde-1 and sid-1 strains from substantial sensitivity to complete insensitivity. This riveting finding warrants a re-evaluation of the tissue-specific interpretations in numerous tissue-specific RNAi studies in C. elegans. Using validated tissue-specific RNAi systems, we determined that proteasomal burden largely differs between the germline and the soma. Notably, gonadally accumulating vitellogenin proteins substantially contributed to the total polyubiquitinated proteins observed during aging. Interestingly, we found that UBQ-1 was essential to maintain the transcription of highly expressed genes, including vitellogenins, supporting a broad and key role for polyubiquitin beyond proteostasis. Altogether, we authenticated commonly used tissue-specific RNAi systems and uncovered a new approach to improve RNAi insensitivity in RNAi-defective strains. With validated tissue-specific RNAi strains, we spatially analyzed proteasomal function and age-related polyubiquitinated protein accumulation and revealed a converging regulatory link between UBQ-1, vitellogenin metabolism and lifespan in C. elegans.

genetics↗

bioRxiv2022

Inhibition of mutant IDH1 promotes cycling of acute myeloid leukemia stem cells

Acute myeloid leukemias (AML) are comprised of multiple cell types with distinct capabilities to propagate the disease and resist therapy. Approximately 20% of AML patients carry gain- of-function mutations in IDH1 or IDH2 that result in over-production of the onco-metabolite 2-HG. Although IDH inhibitors can induce complete morphological remission, almost all patients eventually relapse. Analysis of clinical samples suggests that a population of IDH mutant cells is able to persist during treatment eventually acquiring 2-HG independence and drug resistance. Herein we characterized the molecular and cellular responses to the clinical IDH1 inhibitor AG-120 at high resolution using a novel multi-allelic mouse model of IDH1 mutant AML. We demonstrate that AG-120 exerts cell type-dependent effects on leukemic cells promoting delayed disease regression. Although IDH1 inhibition alone was not able to fully eradicate the disease, we uncovered that it increases cycling of rare leukemic stem cells and triggers transcriptional upregulation of the pyrimidine salvage pathway. Accordingly, AG-120 sensitized IDH1 mutant AML to azacitidine with the combination of AG-120 and azacitidine showing vastly improved efficacy in vivo. Our data highlight the impact of non- genetic heterogeneity on treatment response and provide mechanistic rationale for a drug combination that is being tested in clinical trials. STATEMENT OF SIGNIFICANCEInhibition of mutant IDH1 in AML is insufficient to eliminate the disease but promotes proliferation of quiescent leukemic stem cells. Our data provide a mechanistic explanation for the observed synergy between IDH inhibitors and azacitidine and suggest that IDH inhibitors may also synergize with other drugs that preferentially target actively dividing cells.

cancer biology↗