Biology subjects

Wu, C.-W.

Publications and source records attributed to Wu, C.-W..

7 recordsLinked to original sources

bioRxiv2026

Discovery of a small peptide that activates both APCCdc20 and APCCdh1 in yeast, driving increased lifespan

Aging is accompanied by molecular hallmarks conserved from yeast to humans. One such hallmark, senescence, is an irreversible nondividing state that promotes aging. We previously reported that mutants of the yeast Anaphase Promoting Complex (APC) shorten the lifespan of both dividing and nondividing cells. Here, we propose that activation of the APC will promote the maintenance of quiescent yeast cells, thereby delaying senescence and aging. We observed that APC activity in aging quiescent cells becomes increasingly impaired, as the APC substrates Clb1 and Mps1 accumulated within aging cells reintroduced back into the cell cycle. To identify peptide activators of the APC, we used a yeast 2-hybrid screen to recover peptides that interacted with the APC subunit Apc10. Recovered peptides were tested for their effects on the replicative lifespan (RLS) of dividing cells, and the chronological lifespan (CLS) of nondividing cells. Several peptides increased the RLS of wild type cells, but only one peptide (C43-4) increased CLS, which requires the APC Cdh1 co-activator, but not the Cdc20 co-activator. In cells expressing C43-4, Clb1 and Mps1 levels remained low when aging quiescent cells were reintroduced back into the cell cycle. The addition of a synthetic C43-4 to aging quiescent cells increased CLS even when added at late stages of aging. Mutations to APC subunits and co-activators blocked the ability of C43-4 to extend CLS. Htz1, which shares sequence homology with C43-4, was found to suppress APC mutant phenotypes and stabilize Apc10. We individually mutated the 6 amino acids in C43-4 that were shared with Htz1; one of the mutants disrupted the C43-4-Apc10 2-hybrid interaction and abolished the C43-4-dependent increase in CLS. C43-4 action is evolutionarily conserved, as C43-4 increased C. elegans lifespan in a daf-16- and aak-2-dependent manner. Our results describe the discovery of an evolutionarily conserved translational peptide that is the first of its kind to specifically activate the anti-aging APCCdh1 complex.

genetics↗

bioRxiv2024

Chemical genetics in C. elegans identifies anticancer mycotoxins chaetocin and chetomin as potent inducers of a nuclear metal homeostasis response

C. elegans numr-1/2 (nuclear-localized metal-responsive) is an identical gene pair encoding a nuclear protein previously shown to be activated by cadmium and disruption of the integrator RNA metabolism complex. We took a chemical genetic approach to further characterize regulation of this novel metal response by screening 41,716 compounds and extracts for numr-1p::GFP activation. The most potent activator was chaetocin, a fungal 3,6-epidithiodiketopiperazine (ETP) with promising anticancer activity. Chaetocin activates numr-1/2 strongly in the alimentary canal but is distinct from metal exposure because it represses canonical cadmium-responsive metallothionine genes. Chaetocin has diverse targets in cancer cells including thioredoxin reductase, histone lysine methyltransferase, and acetyltransferase p300/CBP; further work is needed to identify the mechanism in C. elegans as genetic disruption and RNAi screening of homologs did not induce numr-1/2 in the alimentary canal and chaetocin did not affect markers of integrator dysfunction. We demonstrate that disulfides in chaetocin and chetomin, a dimeric ETP analog, are required to induce numr-1/2. ETP monomer gliotoxin, despite possessing a disulfide linkage, had almost no effect on numr-1/2, suggesting a dimer requirement. Chetomin inhibits C. elegans growth at low micromolar levels and loss of numr-1/2 increases sensitivity; C. elegans and Chaetomiaceae fungi inhabit similar environments raising the possibility that numr-1/2 functions as a defense mechanism. There is no direct ortholog of numr-1/2 in humans, but RNAseq suggests that chaetocin affects expression of cellular processes linked to stress response and metal homeostasis in colorectal cancer cells. Our results reveal interactions between metal response gene regulation and ETPs and identify a potential mechanism of resistance to this versatile class of preclinical compounds.

pharmacology and toxicology↗

bioRxiv2023

A role for the C. elegans Argonaute protein CSR-1b isoform in small nuclear RNA 3' processing

The Integrator is a multi-subunit protein complex that catalyzes the maturation of snRNA transcripts via 3 cleavage, a step required for snRNA incorporation with snRNP for spliceosome biogenesis. Here we developed a GFP based in vivo snRNA misprocessing reporter as a readout of Integrator function and performed a genome-wide RNAi screen for Integrator regulators. We found that loss of the Argonaute encoding csr-1 gene resulted in widespread 3 misprocessing of snRNA transcripts that is accompanied by a significant increase in alternative splicing. Loss of csr-1 down-regulates the germline expression of Integrator subunit-4 and is accompanied by a reduced protein translation efficiency of multiple Integrator catalytic and non-catalytic subunits. Through isoform analysis, we identify that csr-1b is specifically required for snRNA processing and this is dependent on its catalytic slicer activity. Moreover, mRNA-sequencing revealed high similarity in the transcriptome profile between csr-1 and Integrator subunit knockdown via RNAi. Together, our findings reveal csr-1b as a new regulator of the Integrator complex and implicate a novel role of this Argonaute protein in snRNA 3 processing.

genetics↗

bioRxiv2022

CCR4-NOT subunit CCF-1/CNOT7 interacts with the PAL-1/CDX-1 transcription factor to regulate multiple stress responses in Caenorhabditis elegans

CCR4-NOT is a versatile eukaryotic protein complex that controls multiple steps in gene expression regulation from synthesis to decay. In yeast, CCR4-NOT has been implicated in stress response regulation, though this function in other organisms remains unclear. In a genome-wide RNAi screen, we identified a subunit of the CCR4-NOT complex, ccf-1, as a requirement for the C. elegans transcriptional response to cadmium and acrylamide stress. Using whole-transcriptome RNA sequencing, we show that knockdown of ccf-1 attenuates the activation of a broad range of stress protective genes in response to cadmium and acrylamide, including those encoding heat shock proteins and glutathione s-transferases. Consistently, survival assays show that knockdown of ccf-1 decreases C. elegans stress resistance. A yeast-2-hybrid screen using a CCF-1 bait identified the homeobox transcription factor PAL-1 as a physical interactor. Knockdown of pal-1 inhibits the activation of ccf-1 dependent stress genes and reduces C. elegans stress resistance. Gene expression analysis reveals that knockdown of pal-1 down-regulates the mRNA levels of elt-2 and elt-3, which serves as the master transcriptional co-regulators of stress response in the C. elegans intestinal and epidermal tissues respectively. These results reveal a new role for CCR4-NOT in stress response regulation with PAL-1 through the transcriptional control of elt-2 and elt-3 in C. elegans.

genetics↗

bioRxiv2021

Translational suppression via IFG-1/eIF4G confers resistance to stress-induced RNA alternative splicing in Caenorhabditis elegans

Splicing of pre-mRNA is an essential process for dividing cells and splicing defects have been linked to aging and various chronic diseases. Environmental stress has recently been shown to alter splicing fidelity and molecular mechanisms that protect against splicing disruption remains unclear. Using an in vivo RNA splicing reporter, we performed a genome-wide RNAi screen in Caenorhabditis elegans and found that protein translation suppression via silencing of the conserved initiation factor 4G (IFG-1/eIF4G) protects against cadmium-induced splicing disruption. Transcriptome analysis of an ifg-1 deficient mutant revealed an overall increase in splicing fidelity and resistance towards cadmium-induced alternative splicing compared to the wild-type. We found that the ifg-1 mutant up-regulates >80 RNA splicing regulatory genes that are controlled by the TGF-{beta} transcription factor SMA-2. The extended lifespan of the ifg-1 mutant is partially reduced upon sma-2 depletion and completely nullified when core spliceosome genes including snr-1, snr-2, and uaf-2 are knocked down. Together, these data describe a molecular mechanism that provides resistance towards stress-induced alternative splicing and demonstrate an essential role for RNA homeostasis in promoting longevity in a translation-compromised mutant.

genetics↗

bioRxiv2021

Tumor Suppressor p53 Restrains Cancer Cell Dissemination by Modulating Mitochondrial Dynamics

Tumor suppressor p53 plays a central role in preventing tumorigenesis. Here, we unravel how p53 modulates mitochondrial dynamics to restrain the metastatic properties of cancer cells. p53 inhibits the mammalian target of rapamycin complex 1 (mTORC1) signaling to attenuate the protein level of mitochondrial fission process 1 (MTFP1), which fosters the pro-fission dynamin-related protein 1 (Drp1) phosphorylation. This regulatory mechanism allows p53 to restrict cell migration and invasion governed by Drp1-mediated mitochondrial fission. Downregulating p53 expression or elevating the molecular signature of mitochondrial fission correlates with aggressive tumor phenotypes and poor prognosis in cancer patients. Upon p53 loss, exaggerated mitochondrial fragmentation stimulates the activation of the extracellular signal-regulated kinase 1/2 (ERK1/2) signaling resulting in epithelial-to-mesenchymal transition (EMT)-like changes in cell morphology, accompanied by accelerated matrix metalloproteinase-9 (MMP9) expression and invasive cell migration. Notably, blocking the activation of mTORC1/MTFP1/Drp1/ERK1/2 axis completely abolishes the p53 deficiency-driven cellular morphological switch, MMP9 expression, and cancer cell dissemination. Our findings unveil a hitherto unrecognized mitochondria-dependent molecular mechanism underlying the metastatic phenotypes of p53-compromised cancers.

molecular biology↗

bioRxiv2018

Targeting BMI1 and MCL1 for Lung Adenocarcinoma Treatment

Lung cancer is the leading cause of cancer-associated death worldwide. Early metastasis and the recurrence remain major challenges for lung cancer treatment in clinic. Targeting the cancer stemness could be a potential strategy to restrain tumor progression. In the current study, we found that in lung adenocarcinoma (LAC), BMI1 and MCL1 play crucial roles in invasion, chemo-resistance, and tumor initiation. JNK signaling is a link between oncogenic pathway or environment stress to cancer stemness. The activation of JNK, either by EGFR or chemotherapy agent, stabilized BMI1 and MCL1 proteins through suppressing the expression of E3-ubiquitin ligase HUWE1. In lung cancer patient samples, high level of BMI1 is correlated with poor survival, and the expression of BMI1 is positively correlated with MCL1. A novel small-molecule BI-44 was synthesized, which effectively suppressed BMI1/MCL1 expression and inhibited tumor formation and progression in preclinical models. Targeting BMI1/MCL1 provides the basis for a new therapeutic approach in the treatment of LAC.

cancer biology↗