BBS Faculty Member - Gary Ruvkun

Gary Ruvkun

Department of Genetics

Massachusetts General Hospital
Simches Research Building, CPZN 7250
185 Cambridge Street
Boston, MA 02114
Tel: 617-726-5959
Email: ruvkun@molbio.mgh.harvard.edu
Lab Members: 8 postdoctoral fellows, 2 project scientists
Visit my lab page here.



The Ruvkun lab uses C. elegans molecular genetics and genomics to study miRNA and RNAi pathways as well as mechanisms of aging and microbial surveillance. We have identified genes that positively or negatively regulate RNAi and microRNA pathways. For example, our screens for C. elegans mutants with enhanced RNA interference identified a specialized Argonaute pathway that recognizes recently acquired genes, for example integrated viruses. Mutants defective in these specialized RNA silencing pathways express these integrated viruses, inducing the Unfolded Protein Response. We are currently exploring how RNA editing allows the antiviral systems to detect viruses divergent from the previous viral exposures that are recorded in the genome.

Our genetic analysis of C. elegans lifespan revealed a surprising surveillance of conserved core components of cells, such as the ribosome, the mitochondrion, and a coupling to the induction of detoxification and innate immune responses. We are now dissecting this surveillance system by isolating mutations in C. elegans that fail to recognize deficits in ribosomal, mitochondrial or proteasomal function, or constitutively induce such responses. For example, we found that germline-specific mutations in translation components are detected by this system to induce detoxification and immune responses in distinct somatic cells. An RNA interference screen revealed gene inactivations that act at multiple steps in a bile acid biosynthetic and kinase pathways upstream of MAP kinase to mediate the systemic communication of translation defects to induce detoxification genes. Mammalian bile acids can rescue the defect in detoxification gene induction caused by C. elegans bile acid biosynthetic gene inactivations. These eukaryotic antibacterial countermeasures are not ignored by bacteria: for example Kocuria rhizophila, suppress normal C. elegans detoxification responses to mutations in translation factors. By genetic analysis of the Kocuria, we identified mutations in carotenoid biosynthesis that are defective in this inhibition of the C. elegans translational toxin defense response. Extracts of this C50 carotenoid from wild type K. rhizophila could restore this bacterial anti-immunity activity. Corynebacterium glutamicum, also inhibits the C. elegans translation detoxification response by producing the C50 carotenoid decaprenoxanthin. These bacterial carotenoids sensitize C. elegans to a lower translation toxin dose and inhibit food aversion behaviors normally induced by protein translation toxins or mutations. Since the surveillance and response to toxins is mediated by signaling pathways conserved across animal phylogeny, bacterial carotenoids may also suppress aberrant human xenobiotic responses, such as in chemotherapy-induced nausea, anorexia nervosa, or autoimmune disorders.

The proteasome mediates selective protein degradation and is dynamically regulated in response to proteotoxic challenges. C. elegans genes encoding proteasome subunits and detoxification factors are strongly upregulated in response to disruption of proteasome function. A large-scale EMS mutagenesis screen on wild type animals carrying a proteasome-GFP fusion gene identified dozens of mutants that constitutively activate the GFP reporter, including weak alleles of proteasome subunit genes. We used one of the weak proteasome subunit alleles that activates a proteasome GFP fusion gene to identify multiple suppressor mutants that fail to up-regulate the proteasome fusion genes in the proteasome mutant background, that is a screen for mutants "blind" to their defective proteasome. Full genome sequencing of these mutant strains identified multiple alleles of png-1/N-glycanase, ddi-1/protease. These proteins mediate the deglycosylation and cleavage activation of the SKN-1/mammalian NRF1 bZip transcription factor as it exits from the ER and is normally degraded by the proteasome. Thus, SKN-1A/Nrf1, an endoplasmic reticulum (ER)-associated transcription factor that undergoes N-linked glycosylation, serves as a sensor of proteasome dysfunction and triggers compensatory upregulation of proteasome subunit genes. The PNG-1/NGLY1 peptide:N-glycanase edits the sequence of SKN-1A protein by converting particular N-glycosylated asparagine residues to aspartic acid. Genetically introducing aspartates at these N-glycosylation sites bypasses the requirement for PNG-1/NGLY1, showing that protein sequence editing rather than deglycosylation is key to SKN-1A function. Misfolded endogenous proteins and the human amyloid beta peptide also trigger activation of proteasome subunit expression by SKN-1A/Nrf1. SKN-1A activation is protective against age-dependent defects caused by accumulation of misfolded and aggregation-prone proteins. In a C. elegans Alzheimer's disease model, SKN-1A/Nrf1 slows accumulation of the amyloid beta peptide and delays adult-onset cellular dysfunction. Thus, SKN-1A surveys cellular protein folding and adjusts proteasome capacity to meet the demands of protein quality control pathways.

In Caenorhabditis elegans, mitochondrial dysfunction caused by mutation or toxins activates programs of detoxification and immune response. A genetic screen for mutations that constitutively induce C. elegans mitochondrial defense revealed reduction-of-function mutations in the mitochondrial chaperone hsp-6/mtHSP70 and gain-of-function mutations in the Mediator component mdt-15/MED15. The activation of detoxification and immune responses is transcriptionally mediated by mdt-15/MED15 and nuclear hormone receptor nhr-45. Mitochondrial dysfunction triggers redistribution of intestinal mitochondria, which requires the mitochondrial Rho GTPase miro-1 and its adaptor trak-1/TRAK1, but not nhr-45-regulated responses. Disabling the mdt-15/nhr-45 pathway renders animals more susceptible to a mitochondrial toxin or pathogenic Pseudomonas aeruginosa but paradoxically improves health and extends lifespan in animals with mitochondrial dysfunction caused by a mutation. Thus, some of the health deficits in mitochondrial disorders may be caused by the ineffective activation of detoxification and immune responses, which may be inhibited to improve health.

Interactions between C. elegans and its microbiome.
C. elegans consumes bacteria which can supply essential vitamins and cofactors especially for mitochondrial functions ancestrally related to bacteria. We screened the Keio E. coli knockout library for mutations that induce a C. elegans mitochondrial damage response gene to identify 45 E. coli mutations that induce a C. elegans mitochondrial UPR gene. Four of these E. coli mutations that disrupt the import or removal of iron from the bacterial siderophore enterobactin were synthetic lethal in combination with C. elegans mutations that disrupt particular iron-sulfur proteins of the electron transport chain. Antioxidants suppress this inviability, suggesting that reactive oxygen species (ROS) are produced by the mutant mitochondria in combination with the bacterial enterobactin-iron complex. The siderophores produced by bacteria are often hijacked by other bacteria utilizing diverse electron transport terminal oxidases. It is possible that the Fe-charged siderophore is a weapon against competitor bacteria that cannot remove the iron from the siderphore.

The kingdoms of life share many small molecule cofactors and coenzymes, for example many vitamins. Molybdenum cofactor (Moco) is synthesized by many archaea, bacteria, and eukaryotes in a multistep pathway from GTP. The genome of Caenorhabditis elegans contains all of the Moco biosynthesis genes, and surprisingly these genes are not essential if the animals are fed a bacterial diet that synthesizes Moco. C. elegans lacking both endogenous Moco synthesis and dietary Moco from bacteria arrest development, demonstrating interkingdom Moco transfer. Our screen of Escherichia coli mutants identified genes necessary for synthesis of bacterial Moco or transfer to C. elegans. A comprehensive genetic screen for C. elegans mutations that can survive the developmental arrest of Moco-deficient C. elegans, showed that arrest is suppressed by mutations in either C. elegans cystathionine gamma-lyase or cysteine dioxygenase, blocking toxic sulfite production from cystathionine. It is the lack of loss of sulfite oxidase, a Moco-requiring enzyme, in Moco deficient animals that causes an increase in toxic sulfite. That sulfite is generated by the cystathionine gamma-lyase or cysteine dioxygenase pathway to sulfite.



Last Update: 8/8/2019



Publications

For a complete listing of publications click here.

 


 

Lehrbach NJ and G Ruvkun. 2019. ER-associated SKN-1A/Nrf1 mediates a cytoplasmic unfolded protein response and promotes longevity. Elife. 2019 Apr 11;8. pii: e44425. doi: 10.7554/eLife.44425. PMID: 30973820

Lehrbach NJ, PC Breen, and G Ruvkun. 2019. Protein sequence editing of SKN-1A/Nrf1 by peptide:N-glycanase controls proteasome gene expression. Cell. 2019 Apr 18;177(3):737-750.e15. doi: 10.1016/j.cell.2019.03.035. PMID: 31002798

Warnhoff K and G Ruvkun. 2019. Molybdenum cofactor transfer from bacteria to nematode mediates sulfite detoxification. Nat Chem Biol. 2019 May;15(5):480-488. doi: 10.1038/s41589-019-0249-y. Epub 2019 Mar 25. PMID: 30911177

Mao K, F Ji, PC Breen, A Sewell, M Han, R Sadreyev, and G Ruvkun. 2019. Mitochondrial dysfunction in C. elegans activates mitochondrial relocalization and nuclear hormone receptor-dependent detoxification genes. Cell Metab. 2019 Feb 14. pii: S1550-4131(19)30022-1. doi: 10.1016/j.cmet.2019.01.022. [Epub ahead of print] PMID: 30799287

Lehrbach NJ and G Ruvkun. 2016. Proteasome dysfunction triggers activation of SKN-1A/Nrf1 by the aspartic protease DDI-1. bioRxiv preprint posted online May. 12, 2016; doi: http://dx.doi.org/10.1101/052936. Elife. 2016 Aug 16;5. pii: e17721. doi: 10.7554/eLife.17721. PMID:27528192

Samuel BS, H Rowedder, C Braendle, MA Félix, G Ruvkun. 2016. Caenorhabditis elegans responses to bacteria from its natural habitats. Proc Natl Acad Sci U S A., 2016.. pii: 201607183. PMID:27317746

Govindan JA, E Jayamani, X Zhang, P Breen, J Larkins-Ford, E Mylonakis and G Ruvkun. 2015. Lipid signalling couples translational surveillance to systemic detoxification in Caenorhabditis elegans. Nat Cell Biol. 2015 Oct;17(10):1294-303. doi: 10.1038/ncb3229. Epub 2015 Aug 31.PMID: 26322678



© 2016 President and Fellows
of Harvard College