Gissendanner CR et al. (2014) Genesis "C. elegans nuclear receptor NHR-6 functionally interacts with the jun-1 transcription factor during spermatheca development."

Praslicka B, El Sayed M, Cardin D, Harmson JS, Rowan BG, Gissendanner CR, Dubose CJ

[Genesis, 2014]

The NR4A nuclear receptor NHR-6 is an essential regulator of spermatheca organogenesis in C. elegans. In this study, we perform a focused, RNAi-based screen to identify modifiers of partial nhr-6 loss of function. Ninety-eight genes that encode signaling proteins expressed in the spermatheca were screened for enhancement of the nhr-6 RNAi phenotype. We identify the C. elegans gene jun-1, which encodes the homolog of the Jun transcription factor, as a strong enhancer of nhr-6 partial loss of function. We show that nhr-6 and jun-1 function together to regulate development of the spermatheca and are necessary for generating an organ with the normal number of cells. jun-1 is expressed in all cells of the developing spermatheca. We also provide evidence that NHR-6 and JUN-1 can physically interact in a yeast two-hybrid assay. Our results provide in vivo evidence that NR4A nuclear receptor and Jun transcription factor interactions are essential in regulating developmental processes in metazoans.

Hiatt SM et al. (2009) Mol Biol Cell "Caenorhabditis elegans FOS-1 and JUN-1 regulate plc-1 expression in the spermatheca to control ovulation."

Matsumoto K, Kerppola TK, Ellis RE, Shyu YJ, Hiatt SM, Hisamoto N, Duren HM, Hu CD, Kariya KI

[Mol Biol Cell, 2009]

Fos and Jun are components of activator protein-1 (AP-1) and play crucial roles in the regulation of many cellular, developmental, and physiological processes. Caenorhabditis elegans fos-1 has been shown to act in uterine and vulval development. Here, we provide evidence that C. elegans fos-1 and jun-1 control ovulation, a tightly regulated rhythmic program in animals. Knockdown of fos-1 or jun-1 blocks dilation of the distal spermathecal valve, a critical step for the entry of mature oocytes into the spermatheca for fertilization. Furthermore, fos-1 and jun-1 regulate the spermathecal-specific expression of plc-1, a gene that encodes a phospholipase C (PLC) isozyme that is rate-limiting for inositol triphosphate production and ovulation, and overexpression of PLC-1 rescues the ovulation defect in fos-1(RNAi) worms. Unlike fos-1, regulation of ovulation by jun-1 requires genetic interactions with eri-1 and lin-15B, which are involved in the RNA interference pathway and chromatin remodeling, respectively. At least two isoforms of jun-1 are coexpressed with fos-1b in the spermatheca, and different AP-1 dimers formed between these isoforms have distinct effects on the activation of a reporter gene. These findings uncover a novel role for FOS-1 and JUN-1 in the reproductive system and establish C. elegans as a model for studying AP-1 dimerization.

Kavita S Oommen et al. (2005) International Worm Meeting "Crosstalk between Notch and Fos/Jun during specification of a uterine cell fate"

Kavita S Oommen, Anna P Newman

[International Worm Meeting, 2005]

One signaling pathway alone can specify numerous yet distinct cell fates during development. Additional mechanisms must be in place to ensure that reiteratively utilized pathways can achieve specificity. In C. elegans, evolutionarily conserved LIN-12/Notch signaling in the developing ventral uterus establishes the cell fate. The cells undergo a well-characterized lineage and differentiate into cell types that form a proper uterine-vulval connection critical for egg laying. The SOX domain transcription factor encoded by egl-13 is specifically expressed in the cells from the time of specification through differentiation and morphogenesis. Through deletional analysis of the upstream regulatory sequences (URS) of an egl-13 transcriptional gene fusion, we deduced a minimal regulatory region critical for driving -specific expression of egl-13. Furthermore, we deleted discrete, highly conserved binding sites within this defined region to pinpoint which cis factor(s) may be relevant for transcriptional regulation of egl-13. In this manner, we isolated a single binding site for LAG-1, a member of the CSL (CBF-1/Su(H)/LAG-1) family of DNA binding proteins known to mediate Notch signaling, that is required for egl-13 expression at the time of cell specification. To our surprise, we discovered that a conserved binding site for the dimeric Fos/Jun proteins is also required for egl-13 expression at specification as well. Fos and Jun proteins constitute a subclass of basic domain leucine zipper transcription factors characterized as oncogenes in mammals. In addition, both sites are necessary for cDNA-mediated rescue of egl-13 mutants. Observations including RNAi of Fos/Jun homologs and genetic mutant characterization of the closest C. elegans Fos homolog support the involvement of Fos/Jun factors in normal uterine development and egl-13 expression. We have conducted in vitro assays to identify the corresponding Jun factor(s) and confirmed direct binding of FOS/JUN proteins to their respective binding site within the egl-13 URS. This study is the first to elucidate a cooperative regulatory mechanism between LIN-12/Notch and Fos/Jun pathways and may explain how Notch signaling in C. elegans is able to establish a distinct cell fate in the cell paradigm.

Susan M Fox et al. (2004) Mid-west Worm Meeting "Identification and characterization of C. elegans Fos and Jun homologs"

Chang-Deng Hu, Elizabeth J Taparowsky, Tom K Kerppola, Alfred Zullo, Ronald E Ellis, Susan M Fox, Soochin Cho

[Mid-west Worm Meeting, 2004]

The basic region leucine zipper (bZIP) proteins constitute of one of the largest families of transcription factors in mammals, Drosophila , C. elegans and plants. They bind to specific DNA sequences as heterodimers or homodimers , and regulate a variety of cellular processes. For example, the activator protein 1 (AP-1) group of bZIP proteins contains the Fos , Jun, ATF2 and Maf subfamilies, which bind to the AP-1 consensus sequences. AP-1 proteins activate a variety of target genes that regulate cellular proliferation, differentiation and death, as well as stress responses. Furthermore, genetic analyses using mouse and Drosophila have revealed that AP-1 proteins play pivotal roles in regulating specific developmental events. P> P> How do bZIP proteins form the correct heterodimers and homodimers at appropriate times during development, out of many other possible combinations? We are developing C. elegans as a system to answer this question. First, we searched the C. elegans genome sequence database and identified 21 unique bZIP-containing proteins. Sequence alignment and phylogenetic analysis of the bZIP domains along with those of Drosophila and human bZIP proteins showed that the proteins encoded by F29G9.4 and T24H10.2 are the closest homologs of Drosophila and human Fos and Jun, respectively. We tentatively name these genes Ce-fos-1 and Ce-jun-1 . P> P> To determine whether these genes regulate specific developmental events in C. elegans , we used RNA interference. Initial studies suggest that Ce-fos-1 is required for normal vulva development, and Ce-jun-1 is required for embryonic viability. To characterize their DNA-binding ability and dimerization selectivity, we carried out gel mobility shift assays, and found that they bind AP-1 consensus sequences as a heterodimer . As is the case with mammalian Fos and Jun proteins, Ce-JUN-1 can also form homodimers , but Ce-FOS-1 cannot. Furthermore, both Ce-FOS-1 and Ce-JUN-1 could form heterodimers with rat c-Fos to bind DNA, suggesting that they are functionally interchangeable in terms of DNA binding. Finally, these interactions have been confirmed in living cells and in living animals using a bimolecular fluorescence complementation ( BiFC ) assay. Taken together, our results strongly suggest that Ce-FOS-1 and Ce-JUN-1 are the C. elegans Fos and Jun proteins, and that these genes play important roles in C. elegans development. Our ability to assay dimerization in vivo should allow us to elucidate how their pattern of dimerization changes during development.

Vinci CR et al. (2007) J Biol Chem "Recognition of age-damaged (R,S)-adenosyl-L-methionine by two methyltransferases in the yeast Saccharomyces cerevisiae."

Vinci CR, Clarke SG

[J Biol Chem, 2007]

The biological methyl donor, S adenosylmethionine (AdoMet), can exist in two diastereoisomeric states with respect to its sulfonium ion. The "S" configuration, (S,S)AdoMet, is the only form that is produced enzymatically as well as the only form used in almost all biological methylation reactions. Under physiological conditions, however, the sulfonium ion can spontaneously racemize to the "R" form, producing (R,S)AdoMet. As of yet, (R,S)AdoMet has no known physiological function and may inhibit cellular reactions. In this study, two enzymes have been found in Saccharomyces cerevisiae that are capable of recognizing (R,S)AdoMet and using it to methylate homocysteine to form methionine. These enzymes are the products of the SAM4 and MHT1 genes, previously identified as homocysteine methyltransferases dependent upon AdoMet and S-methylmethionine respectively. We find here that Sam4 recognizes both (S,S) and (R,S)AdoMet, but its activity is much higher with the R,S form. Mht1 reacts with only the R,S form of AdoMet while no activity is seen with the S,S form. R,S-specific homocysteine methyltransferase activity is also shown here to occur in extracts of Arabidopsis thaliana, Drosophila melanogaster, and Caenorhabditis elegans, but has not been detected in several tissue extracts of Mus musculus. Such activity may function to prevent the accumulation of (R,S)AdoMet in these organisms.

Susan M Fox et al. (2005) International Worm Meeting "Identification and Characterization of C. elegans Fos and Jun Homologs"

John Shyu, Elizabeth J Taparowsky, Ronald E Ellis, Tom K Kerppola, Susan M Fox, Soochin Cho, Susan Strome, Alfred Zullo, Chang-Deng Hu

[International Worm Meeting, 2005]

The basic region leucine zipper (bZIP) proteins constitute one of the largest families of transcription factors in mammals, Drosophila, and C. elegans. These proteins function as dimers to bind specific DNA sequences and regulate a variety of cellular processes. For example, the activator protein 1 (AP-1) group of bZIP proteins, containing Fos, Jun, ATF2 and Maf subfamilies bind AP-1 consensus sequences and activate a variety of target genes that regulate cellular proliferation, differentiation and death, as well as stress responses. Furthermore, genetic analyses using mice and Drosophila have revealed that AP-1 proteins play pivotal roles in regulating specific developmental events. How do bZIP proteins chose the correct partner for a specific heterodimer or homodimer at appropriate times during development? We are developing C. elegans as a system to answer this. Following identification of 24 unique bZIP proteins, sequence alignment and phylogenetic analysis of the C. elegans bZIP domains along with those of Drosophila and human bZIP proteins showed that the proteins encoded by F29G9.4 and T24H10.7 are the closest homologs of Drosophila and human Fos and Jun, respectively. We tentatively named these genes Ce-fos-1 and Ce-jun-1. Using RNA interference, we have found that Ce-fos-1 is required for fertility, as 95% of offspring were sterile. Observation of native expression patterns is underway, using GFP fusion constructs, antibody staining and in situ hybridization. We have found that the bZIP regions of Ce-FOS-1 and Ce-JUN-1 bind AP-1 consensus sequences as a heterodimer, and Ce-JUN-1 can bind as a homodimer. These interactions have been confirmed in living cells and in living worms using the bimolecular fluorescence complementation (BiFC) assay, a novel approach used to visualize protein-protein interactions. The identification of native promoters of Ce-fos-1 and Ce-jun-1 and the visualization of the temporal and spatial interactions of Ce-FOS-1 and Ce-JUN-1 are underway.

Fanning S et al. (2018) Mol Cell "Lipidomic Analysis of -Synuclein Neurotoxicity Identifies Stearoyl CoA Desaturase as a Target for Parkinson Treatment."

Lou Y, Haque A, Freyzon Y, Farese RV, Terry-Kantor E, Hofbauer HF, Termine D, Welte MA, Barrasa MI, Imberdis T, Noble T, Lindquist S, Clish CB, Jaenisch R, Pincus D, Nuber S, Sandoe J, Kohlwein SD, Kim TE, Ho GPH, Ramalingam N, Walther TC, Baru V, Selkoe D, Srinivasan S, Landgraf D, Soldner F, Dettmer U, Fanning S, Becuwe M, Newby G

[Mol Cell, 2018]

In Parkinson's disease (PD), -synuclein (S) pathologically impacts the brain, a highly lipid-rich organ. We investigated how alterations in S or lipid/fattyacid homeostasis affect each other. Lipidomic profiling of human S-expressing yeast revealed increases in oleic acid (OA, 18:1), diglycerides, and triglycerides. These findings were recapitulated in rodent and human neuronal models of S dyshomeostasis (overexpression; patient-derived triplication or E46K mutation; E46K mice). Preventing lipid droplet formation or augmenting OA increased S yeast toxicity; suppressing the OA-generating enzyme stearoyl-CoA-desaturase (SCD) was protective. Genetic or pharmacological SCD inhibition ameliorated toxicity in S-overexpressing rat neurons. In a C.elegans model, SCD knockout prevented S-induced dopaminergic degeneration. Conversely, we observed detrimental effects of OA on S homeostasis: in human neural cells, excess OA caused S inclusion formation, which was reversed by SCD inhibition. Thus, monounsaturated fatty acid metabolism is pivotal for S-induced neurotoxicity, and inhibiting SCD represents a novel PD therapeutic approach.

Vandecandelaere I et al. (2017) PLoS One "Metabolic activity, urease production, antibiotic resistance and virulence in dual species biofilms of Staphylococcus epidermidis and Staphylococcus aureus."

Deforce D, Coenye T, Van Nieuwerburgh F, Vandecandelaere I

[PLoS One, 2017]

In this paper, the metabolic activity in single and dual species biofilms of Staphylococcus epidermidis and Staphylococcus aureus isolates was investigated. Our results demonstrated that there was less metabolic activity in dual species biofilms compared to S. aureus biofilms. However, this was not observed if S. aureus and S. epidermidis were obtained from the same sample. The largest effect on metabolic activity was observed in biofilms of S. aureus Mu50 and S. epidermidis ET-024. A transcriptomic analysis of these dual species biofilms showed that urease genes and genes encoding proteins involved in metabolism were downregulated in comparison to monospecies biofilms. These results were subsequently confirmed by phenotypic assays. As metabolic activity is related to acid production, the pH in dual species biofilms was slightly higher compared to S. aureus Mu50 biofilms. Our results showed that S. epidermidis ET-024 in dual species biofilms inhibits metabolic activity of S. aureus Mu50, leading to less acid production. As a consequence, less urease activity is required to compensate for low pH. Importantly, this effect was biofilm-specific. Also S. aureus Mu50 genes encoding virulence-associated proteins (Spa, SplF and Dps) were upregulated in dual species biofilms compared to monospecies biofilms and using Caenorhabditis elegans infection assays, we demonstrated that more nematodes survived when co-infected with S. epidermidis ET-024 and S. aureus mutants lacking functional spa, splF or dps genes, compared to nematodes infected with S. epidermidis ET-024 and wild- type S. aureus. Finally, S. epidermidis ET-024 genes encoding resistance to oxacillin, erythromycin and tobramycin were upregulated in dual species biofilms and increased resistance was subsequently confirmed. Our data indicate that both species in dual species biofilms of S. epidermidis and S. aureus influence each other's behavior, but additional studies are required necessary to elucidate the exact mechanism(s) involved.

Vandecandelaere I et al. (2014) Pathog Dis "Protease production by Staphylococcus epidermidis and its effect on Staphylococcus aureus biofilms."

Coenye T, Vandecandelaere I, Depuydt P, Nelis HJ

[Pathog Dis, 2014]

Due to the resistance of Staphylococcus aureus to several antibiotics, treatment of S. aureus infections is often difficult. As an alternative to conventional antibiotics, the field of bacterial interference is investigated. Staphylococcus epidermidis produces a serine protease (Esp) which inhibits S. aureus biofilm formation and which degrades S. aureus biofilms. In this study, we investigated the protease production of 114 S. epidermidis isolates, obtained from biofilms on endotracheal tubes (ET). Most of the S. epidermidis isolates secreted a mixture of serine, cysteine and metalloproteases. We found a link between high protease production by S. epidermidis and the absence of S. aureus in ET biofilms obtained from the same patient. Treating S. aureus biofilms with the supernatant (SN) of the most active protease producing S. epidermidis isolates resulted in a significant biomass decrease compared to untreated controls, while the number of metabolically active cells was not affected. The effect on the biofilm biomass was mainly due to serine proteases. Staphylococcus aureus biofilms treated with the SN of protease producing S. epidermidis were thinner with almost no extracellular matrix. An increased survival of Caenorhabditis elegans, infected with S. aureus Mu50, was observed when the SN of protease positive S. epidermidis was added.

Duren, Holli Marie et al. (2009) International Worm Meeting "Role of C. elegans JUN-1 in UV-induced DNA damage."

Hu, Chang-Deng, Shyu, Y. John, Duren, Holli Marie, Hiatt, Susan Marie

[International Worm Meeting, 2009]

Genome integrity is critically important for organisms to survive and flourish. Cells have protective measures, such as cell cycle arrest and DNA repair, to combat the effects of DNA damage. If a particular signaling pathway such as cell cycle arrest, DNA repair, or apoptosis, is not functional properly, the possibility of mutations occurring and incorporating into the genome increases, thus leading to increased cancer risk. Skin cancer is the most common type of cancer in the United States with 90% of the non-melonoma skin cancers associated with UV exposure. Activator protein 1 (AP-1), a family of dimeric transcription factors comprised of Jun, Fos, and ATF subfamilies, has been shown to be an immediate early response gene upon UV exposure and overexpression of AP-1 has been shown to play a role in skin cancer development in vitro and in vivo suggesting a possible role of AP-1 in DNA damage-induced skin cancer. To determine the role of AP-1 genes in UV response in intact animals, we have identified the C. elegans Fos, Jun, and ATF2 homologous genes (fos-1, jun-1, and atf-1) and investigated how they are involved in UV response. Using RNA interference methods, we have shown that knockdown of these individual AP-1 genes sensitized the worms to UV-induced worm death. Because UV affects cell cycle arrest, cell death, and DNA repair; we proceeded to expose worms to UV and study the effect on cell cycle arrest. RNAi knockdown of the each AP-1 gene showed no significant effect; however, the jun-1 deletion mutant demonstrated an increase in arrest at the higher dose of 120 Jm-2, 12 hours after exposure. The atf-1 deletion mutant showed no change in arrest as compared to vector. Further, germline apoptosis assays showed that RNAi knock-down of jun-1, but not fos-1 or atf-1, sensitized the worms to UV-induced cell death. Similar results were obtained when a jun-1 deletion mutant was used. Further genetic analysis suggested that JUN-1 acts upstream of the canonical programmed cell death pathway in C. elegans and may regulate the transcription xpf-1, an XP gene involved in nucleotide excision repair (NER). The results presented here strongly suggest that C. elegans jun-1 is involved in the UV-mediated cell death pathway. Current effort is directed to determine what AP-1 dimers are involved in UV-induced cell death and DNA repair.