Rohrer SP et al. (1994) Biochem J "Immunoaffinity purification of avermectin-binding proteins from the free-living nematode Caenorhabditis elegans and the fruitfly Drosophila melanogaster."

Rohrer SP, Jacobson EB, Hayes EC, Birzin ET, Schaeffer JM

[Biochem J, 1994]

Avermectin-binding proteins from the free-living nematode worm Caenorhabditis elegans and from the fruitfly Drosophila melanogaster were purified to homogeneity via a three-step procedure. The binding proteins were covalently labelled using- a radioactive photoaffinity probe and then partially purified on a Sephacryl S-300 gel-filtration column. The radiolabelled binding proteins were then purified by immunoaffinity chromatography using a monoclonal antibody to avermectin covalently attached to Protein A-Sepharose beads. Three affinity-labelled Drosophila proteins with molecular masses between 45 and 50 kDa were isolated in this way and then separated from each other by electroelution. This three-step protocol provides a rapid technique for receptor purification which may be of use in the purification of other binding proteins.

Schmidt R et al. (2017) J Cell Biol "Two populations of cytoplasmic dynein contribute to spindle positioning in C. elegans embryos."

Schmidt R, Fielmich LE, Akhmanova A, Katrukha EA, van den Heuvel S, Grigoriev I

[J Cell Biol, 2017]

The position of the mitotic spindle is tightly controlled in animal cells as it determines the plane and orientation of cell division. Contacts between cytoplasmic dynein and astral microtubules (MTs) at the cell cortex generate pulling forces that position the spindle. An evolutionarily conserved G-GPR-1/2(Pins/LGN)-LIN-5(Mud/NuMA) cortical complex interacts with dynein and is required for pulling force generation, but the dynamics of this process remain unclear. In this study, by fluorescently labeling endogenous proteins in Caenorhabditis elegans embryos, we show that dynein exists in two distinct cortical populations. One population directly depends on LIN-5, whereas the other is concentrated at MT plus ends and depends on end-binding (EB) proteins. Knockout mutants lacking all EBs are viable and fertile and display normal pulling forces and spindle positioning. However, EB protein-dependent dynein plus end tracking was found to contribute to force generation in embryos with a partially perturbed dynein function, indicating the existence of two mechanisms that together create a highly robust force-generating system.

Yang S et al. (2022) Autophagy "Regulation of TFEB nuclear localization by HSP90AA1 promotes autophagy and longevity."

Hu Y, Qi J, She H, He D, Huang L, Mao Z, Lu F, Feng D, Kukar T, Yang Q, Ma L, Nie T, Yang S, Tao K, Zhu L

[Autophagy, 2022]

TFEB (transcription factor EB) regulates multiple genes involved in the process of macroautophagy/autophagy and plays a critical role in lifespan determination. However, the detailed mechanisms that regulate TFEB activity are not fully clear. In this study, we identified a role for HSP90AA1 in modulating TFEB. HSP90AA1 was phosphorylated by CDK5 at Ser 595 under basal condition. This phosphorylation inhibited HSP90AA1, disrupted its binding to TFEB, and impeded TFEB's nuclear localization and subsequent autophagy induction. Pro-autophagy signaling attenuated CDK5 activity and enhanced TFEB function in an HSP90AA1-dependent manner. Inhibition of HSP90AA1 function or decrease in its expression significantly attenuated TFEB's nuclear localization and transcriptional function following autophagy induction. HSP90AA1-mediated regulation of a TFEB ortholog was involved in the extended lifespan of Caenorhabditis elegans in the absence of its food source bacteria. Collectively, these findings reveal that this regulatory process plays an important role in modulation of TFEB, autophagy, and longevity.Abbreviations : AL: autolysosome; AP: autophagosome; ATG: autophagy related; BafA1: bafilomycin A1; CDK5: cyclin-dependent kinase 5; CDK5R1: cyclin dependent kinase 5 regulatory subunit 1; CR: calorie restriction; FUDR: 5-fluorodeoxyuridine; HSP90AA1: heat shock protein 90 alpha family class A member 1; MAP1LC3: microtubule associated protein 1 light chain 3; NB: novobiocin sodium; SQSTM1: sequestosome 1; TFEB: transcription factor EB; WT: wild type.

(1990) Worm Breeder's Gazette "CGC Bibliography in FileMaker and HyperCard (Mac) and Memory Mate ( IBM)"

[Worm Breeder's Gazette, 1990]

The CGC Bibliography has been translated into a couple of programs other than dBase by various worm people. David Barker and Andy Fire both have sent HyperCard versions to the CGC and Mark Blaxter has sent us a version in FileMaker 2.0. None of these is perfectly up-to-date, so you'll have to be somewhat familiar with the programs to add new references. The data files are available free from the CGC; to get yours, just send a blank 3.5' diskette to Mark Edgley at the CGC with a request letter. In addition, Lew Jacobson has translated the bibliography into a DOS program called Memory Mate and he is willing to distribute the data file to anyone who sends him a blank 5.25' 360 Kb diskette. Memory Mate can be operated as a TSR and called up with a hotkey from the middle of a word processor or other program. Addresses for Mark and Lew can be found in the Subscriber Directory Update in this issue.

Conradt B (2000) Ernst Schering Res Found Workshop "Programmed cell death and its regulation and initiation in C. elegans."

Conradt B

[Ernst Schering Res Found Workshop, 2000]

The proliferation of cells is an integral part of development and tissue homeostatsis in multicellular animals(reviewed by Raff 1996; Folletee and O'farrel 1997). Two opposing processes, the division of cells on one hand and the programmed death of cells on the other hand, determine the overall rate of cell proliferation, The proper regulation of these two physiological processes is therefore a crucial aspect of development and of tissue homeostatsis(reviewed by Edgar and Lehner 1996; Shrr 1997;Jacobson et al. 1997). While the importance of the process of cell division has long been recognized, the role and extent of programmed cell death, or apoptosis, has only been realized within the last decades (Glucksmann 1950; Kerr et al. 19972). Massive programmed cell death occurs, for instance, during the development of the nervous system and in the immune system: more than 50% on all neurons and oligodendrocytes formed in the periperal and central vertebrate nervous system undergo programmed cell death neurogenesis...

Olgun A et al. (2007) Ann N Y Acad Sci "Mitochondrial DNA-deficient models and aging."

Akman S, Olgun A

[Ann N Y Acad Sci, 2007]

Human mitochondrial DNA (mtDNA) encodes 13 subunits of oxidative phosphorylation (OXPHOS) enzyme complexes I, III, IV, and V except complex II. MtDNA is more sensitive to oxidative damage than nuclear DNA. MtDNA defects are involved in many pathologies including aging. Several mtDNA-deficient cell culture, yeast, and animal models were generated to study the role of mtDNA in many physiological processes. Ethidium bromide (EB), an agent that is known to inhibit mtDNA replication with a negligible effect on nuclear DNA, is generally used to generate mtDNA-deficient models. The antibiotics chloramphenicol and doxycycline, which were known to inhibit mitochondrial translation, were also used to generate the same phenotype. Cultured mtDNA-deficient cells need uridine and pyruvate to survive. At the organismal level, uridine can be supplemented, but pyruvate supplementation can cause a worser phenotype because of lactic acidosis. In C. elegans, EB, when used during larval development, increases life span, but decreases, when used after the beginning of adult stage. This should be kept in mind since mitochondria-related genes are generally detected in genome-wide screening studies for longevity. We believe that conditional knockout studies need to be carried out for these genes after reaching adulthood. MtDNA mutator mouse did not show an increase of free radical production. Therefore, the downstream phenomena to mtDNA defects are likely ineffective pyrimidine synthesis (dihydroorotate dehydrogenase, DHODH, needs a functional respiratory chain) and excess NADH (decreased NAD pool) in addition to free radicals.

Karen Curto et al. (2007) International Worm Meeting "Regulation of protein degradation in muscle: CaMKII and calcineurin as potential regulators of MAP kinase signaling."

Karen Curto, Nate Szewczyk, Neil Umbreit, Lew Jacobson

[International Worm Meeting, 2007]

Multiple receptor signals modulate protein degradation in C. elegans muscle through multiple mechanisms. Activation of EGL-15 FGF receptor promotes activation of MPK-1 MAP kinase, which in turn triggers non-proteasome dependent protein degradation (Szewczyk & Jacobson, 2003). This is opposed by signal from the DAF-2 insulin/IGF receptor (Szewczyk & Jacobson, 2007). In a seemingly independent mechanism, influx of calcium upon activation of nicotinic acetylcholine receptor inhibits proteasome-mediated muscle protein degradation (Szewczyk et al., 2000). We have now found that UNC-43 calcium/calmodulin-dependent protein kinase II (CaMKII) also has a role in controlling muscle protein degradation. Gain-of-function mutation unc-43(n498) activates CaMKII (without increasing its phosphorylation) and promotes progressive protein degradation. The CaMKII-gf mutation causes an increase in phospho-MAPK, whereas transgene-activated MAPK does not increase phospho-CaMKII. A mutant (egl-19gf) with elevated intramuscular calcium also contains increased levels of both phospho-CaMKII and phospho-MAPK, suggesting that elevated calcium activates CaMKII, which in turn activates MAPK by an unknown mechanism. Conversely, we find that reduction-of-function mutant unc-43(e408) is resistant to protein degradation that occurs in wild-type after either hyperactivation of the FGFR pathway or inhibition of the IGFR pathway. The amount of total MPK-1 protein is increased in unc-43rf mutants, possibly implying that the worm attempts to compensate for low CaMKII signal by up-regulating MAPK. Our current hypothesis is that UNC-43 CaMKII acts as a calcium-responsive positive modulator (sensitivity control?) of the FGFR-Ras-Raf-MEK-MAPK cascade. Reduction-of-function mutations in either tax-6 or cnb-1, which encode the two subunits of the calcium/calmodulin-dependent protein phosphatase calcineurin, result in decreased protein levels in muscle, suggesting that calcineurin may act to oppose muscle protein degradation. Neither reduction- nor gain-of-function mutations in tax-6 or cnb-1 alter the levels of total or phospho-CaMKII; thus, calcineurin may act downstream of or in parallel to CaMKII to provide yet another mechanism by which calcium regulates muscle protein degradation. Supported by NSF grant MCB-0542355.

Sohn J et al. (2024) Autophagy "HLH-30/TFEB mediates sexual dimorphism in immunity in caenorhabditis elegans."

Jung Y, Ha SG, Sohn J, Lee SV, Lee J, Kwon S, Ham S, Lee Y, Lee GY, Lee D, Kim SS, Park HH, Park S

[Autophagy, 2024]

Sexual dimorphism affects various biological functions, including immune responses. However, the mechanisms by which sex alters immunity remain largely unknown. Using <i>Caenorhabditis elegans</i> as a model species, we showed that males exhibit enhanced immunity against various pathogenic bacteria through the upregulation of HLH-30 (Helix Loop Helix 30/TFEB (transcription factor EB), a transcription factor crucial for macroautophagy/autophagy. Compared with hermaphroditic <i>C. elegans</i>, males displayed increased activity of HLH-30/TFEB, which contributed to enhanced antibacterial immunity. <i>atg-2</i> (AuTophaGy (yeast Atg homolog) 2) upregulated by HLH-30/TFEB mediated increased immunity in male <i>C. elegans</i>. Thus, the males appear to be equipped with enhanced HLH-30/TFEB-mediated autophagy, which increases pathogen resistance, and this may functionally prolong mate-searching ability with reduced risk of infection.

Settembre C et al. (2013) Nat Cell Biol "TFEB controls cellular lipid metabolism through a starvation-induced autoregulatory loop."

Irazoqui JE, Ballabio A, Chan L, Saha PK, Settembre C, Vetrini F, Di Bernardo D, Huynh T, Klisch TJ, Palmer D, Wollenberg AC, Mansueto G, Carissimo A, De Cegli R, Visvikis O

[Nat Cell Biol, 2013]

The lysosomal-autophagic pathway is activated by starvation and plays an important role in both cellular clearance and lipid catabolism. However, the transcriptional regulation of this pathway in response to metabolic cues is uncharacterized. Here we show that the transcription factor EB (TFEB), a master regulator of lysosomal biogenesis and autophagy, is induced by starvation through an autoregulatory feedback loop and exerts a global transcriptional control on lipid catabolism via Ppargc1 and Ppar1. Thus, during starvation a transcriptional mechanism links the autophagic pathway to cellular energy metabolism. The conservation of this mechanism in Caenorhabditis elegans suggests a fundamental role for TFEB in the evolution of the adaptive response to food deprivation. Viral delivery of TFEB to the liver prevented weight gain and metabolic syndrome in both diet-induced and genetic mouse models of obesity, suggesting a new therapeutic strategy for disorders of lipid metabolism.

Lapierre LR et al. (2013) Nat Commun "The TFEB orthologue HLH-30 regulates autophagy and modulates longevity in Caenorhabditis elegans."

Irazoqui JE, Hansen M, McQuary PR, Dillin A, Lapierre LR, Gelino S, Chu CC, Ong B, De Magalhaes Filho CD, Chang JT, Davis AE, Visvikis O

[Nat Commun, 2013]

Autophagy is a cellular recycling process that has an important anti-aging role, but the underlying molecular mechanism is not well understood. The mammalian transcription factor EB (TFEB) was recently shown to regulate multiple genes in the autophagy process. Here we show that the predicted TFEB orthologue HLH-30 regulates autophagy in Caenorhabditis elegans and, in addition, has a key role in lifespan determination. We demonstrate that hlh-30 is essential for the extended lifespan of Caenorhabditis elegans in six mechanistically distinct longevity models, and overexpression of HLH-30 extends lifespan. Nuclear localization of HLH-30 is increased in all six Caenorhabditis elegans models and, notably, nuclear TFEB levels are augmented in the livers of mice subjected to dietary restriction, a known longevity-extending regimen. Collectively, our results demonstrate a conserved role for HLH-30 and TFEB in autophagy, and possibly longevity, and identify HLH-30 as a uniquely important transcription factor for lifespan modulation in Caenorhabditis elegans.