Nishikori, K. et al. (2017) International Worm Meeting "RNAi screening of genetic factors required for the biogenesis of intestinal granular organelles in C. elegans - Involvement of intestinal proton transporters."

Hosoda, A., Tanji, T., Kawano, K., Aoyama, R., Shiraishi, H., Ohashi-Kobayashi, A., Nishikori, K.

[International Worm Meeting, 2017]

The cytosol of the intestinal cells of C. elegans was apparently occupied by superficially indistinguishable granular organelles (intestinal granules), which consist of at least three types of organelles: acidified lysosome-related gut granules, lipid droplets, and HEBE (HAF-4/HAF-9-enriched body evanescent with age) granules. Although the former two types of organelles were studied well, HEBE granules have been largely overlooked. HAF-4 and HAF-9 are homologous to mammalian lysosomal peptide transporter ABCB9 and localize to the membrane of HEBE granules. Population of HEBE granules changes dynamically by aging and food conditions. Although characters we found suggest roles of HEBE granules in nutritional resource allocation in response to physiologic changes during the young adult stage, regulatory mechanisms of the organelle dynamics have yet to be determined. To reveal the genetic mechanisms underlying the biogenesis of HEBE granules, we performed RNAi screening according to the following scheme: synchronized L1 larvae were fed with RNAi feeding library clones (Open Biosystems) on 12-well plates. Worms at adult day 1, when HEBE granules are most prominent in the control condition, were directly observed using digital microscopes to efficiently evaluate the presence of intestinal granules. Positive clones were then subjected to the second screening by differential interference contrast microscopy. We previously reported the progress of the screening with preliminary analyses of genes for pentose phosphate pathway (PPP) and vacuolar proton transporter (V-ATPase) subunits (Shiraishi et al., and Nishikori et al., 19th and 20th International C. elegans Meeting). In this meeting, we will present results of the whole screening, in which worms fed with 11,263 out of 11,479 RNAi clones reached adulthood and 142 positive clones were obtained. The gene ontology analysis revealed enrichment of genes associated with metabolic processes including PPP and vacuolar proton transport. In view of proton transport, we found a Na+/H+ exchanger NHX-2 and its regulator PBO-1 among the positive clones. Since V-ATPase and NHX-2 are assumed to play a role in nutritional uptake via acidification of the intestinal lumen in C. elegans, we examined involvement of H+/oligopeptide symporter PEPT-1 in the biogenesis of HEBE granules using pept-1 mutant. These results suggest that oligopeptide uptake is responsible for the biogenesis of HEBE granules.

Thomas JH et al. (1994) Worm Breeder's Gazette "lin-36, a Class B Synthetic Multivulva Gene, Encodes a Novel Protein"

Thomas JH, Horvitz HR

[Worm Breeder's Gazette, 1994]

lin-36, a Class B Synthetic Multivulva Gene, Encodes a Novel Protein Jeffrey H. Thomas and H. Robert Horvitz, HHMI, Dept. Biology, MIT, Cambridge, MA 02139, USA

Sun J et al. (2013) Exp Parasitol "Parasitism of secondary-stage juvenile of Heterodera glycines and four larva stages of Caenorhabditis elegans by Hirsutella spp."

Sun J, Xie H, Xiang M, Liu X, Qiu J

[Exp Parasitol, 2013]

The fungi Hirsutella rhossiliensis and Hirsutella minnesotensis generally parasitize only plant-parasitic nematodes in nature but parasitize the bacterivorous nematode Caenorhabditis elegans on agar plates. To establish a model system for studying the interaction between fungi and nematodes, we compared the parasitism of the first- to fourth-stage larvae (L1-L4) of C. elegans and second-stage juvenile (J2) of Heterodera glycines by twenty isolates of Hirsutella spp. Although parasitism differed substantially among isolates, both H. minnesotensis and H. rhossiliensis parasitized a higher percentage of H. glycines J2s than of C. elegans larvae. Parasitism of C. elegans L1s was correlated with parasitism of H. glycines J2s. Parasitism of C. elegans by H. rhossiliensis and H. minnesotensis was negatively correlated with larva size and motility, i.e., parasitism was higher for the younger stages. The C. elegans L1 is recommended for studying parasitism of nematodes by H. rhossiliensis and H. minnesotensis.

Fei YJ et al. (2000) J Biol Chem "A novel H(+)-coupled oligopeptide transporter (OPT3) from Caenorhabditis elegans with a predominant function as a H(+) channel and an exclusive expression in neurons."

Leibach FH, Romero MF, Krause MW, Huang W, Ganapathy V, Fei YJ, Liu JC

[J Biol Chem, 2000]

We have cloned and functionally characterized a novel, neuron-specific, H(+)-coupled oligopeptide transporter (OPT3) from Caenorhabditis elegans that functions predominantly as a H(+) channel. The opt3 gene is approximately 4.4 kilobases long and consists of 13 exons. The cDNA codes for a protein of 701 amino acids with 11 putative transmembrane domains. When expressed in mammalian cells and in Xenopus laevis oocytes, OPT3 cDNA induces H(+)-coupled transport of the dipeptide glycylsarcosine. Electrophysiological studies of the transport function of OPT3 in Xenopus oocytes show that this transporter, although capable of mediating H(+)-coupled peptide transport, functions predominantly as a H(+) channel. The H(+) channel activity of OPT3 is approximately 3-4-fold greater than the H(+)/peptide cotransport activity as determined by measurements of H(+) gradient-induced inward currents in the absence and presence of the dipeptide using the two-microelectrode voltage clamp technique. A downhill influx of H(+) was accompanied by a large intracellular acidification as evidenced from the changes in intracellular pH using an ion-selective microelectrode. The H(+) channel activity exhibits a K(0.5)(H) of 1.0 microM at a membrane potential of -50 mV. At the level of primary structure, OPT3 has moderate homology with OPT1 and OPT2, two other H(+)-coupled oligopeptide transporters previously cloned from C. elegans. Expression studies using the opt3::gfp fusion constructs in transgenic C. elegans demonstrate that opt3 gene is exclusively expressed in neurons. OPT3 may play an important physiological role as a pH balancer in the maintenance of H(+) homeostasis in C. elegans.

Imadzu H et al. (1994) Worm Breeder's Gazette "Two Tropomyosins Expressed in Body Wall and the Third Did In Pharynx of Caenorhabditis elegans"

Kagawa H, Sakube Y, Imadzu H

[Worm Breeder's Gazette, 1994]

TWO TROPOMYOSINS EXPRBSSBD IN BODY WALL AND THE THIRD DID IN PHARYNX OF CAENORHABDDITIS ELEGANS. H. Imadzu, Y. Sakube and H. gagawa. Department of Biology, Faculty of Science, Okayama University, Okayama, 700 Japan.

Wilson PA et al. (1982) Parasitology "Strongyloides ratti (homogonic): the time-course of early migration in the generalized host deduced from experiments in lactating rats."

Steven GE, Simpson NE, Wilson PA

[Parasitology, 1982]

Unsuckled mother rats given a 1 h suckling stimulus 3 h after subcutaneous injection of an exact dose of homogonic Strongyloides ratti allow fewer worms to develop in their intestines by day 9 than nulliparous rats (Wilson & Simpson, 1981). This effect is studied in more detail in terms of the length of time between weaning and stimulus (W leads to S) and injection and stimulus (I leads to S). It was observable with a W leads to S of 30 h but this and a period of 5 h were less effective than 24 h. With W leads to S constant at 24 h, significantly more worms developed in mothers when I leads to S was 24 h compared to 3 h and 10 h (P less than 0.005). The data, combined with those from nulliparous controls, are presented as a measure of the change with time of numbers of larvae in that compartment of the system which gives access to the stimulated mammary gland. It is argued that the particular compartment is the local lymph node draining the injection site and that the kinetics deduced are applicable to migration in the rat in general.

Miller DL et al. (2007) Proc Natl Acad Sci U S A "Hydrogen sulfide increases thermotolerance and lifespan in Caenorhabditis elegans."

Miller DL, Roth MB

[Proc Natl Acad Sci U S A, 2007]

Hydrogen sulfide (H(2)S) is naturally produced in animal cells. Exogenous H(2)S has been shown to effect physiological changes that improve the capacity of mammals to survive in otherwise lethal conditions. However, the mechanisms required for such alterations are unknown. We investigated the physiological response of Caenorhabditis elegans to H(2)S to elucidate the molecular mechanisms of H(2)S action. Here we show that nematodes exposed to H(2)S are apparently healthy and do not exhibit phenotypes consistent with metabolic inhibition. Instead, animals exposed to H(2)S are thermotolerant and long-lived. These phenotypes require SIR-2.1 activity but are genetically independent of the insulin signaling pathway, mitochondrial dysfunction, and caloric restriction. These studies suggest that SIR-2.1 activity may translate environmental change into physiological alterations that improve survival. It is interesting to consider the possibility that the mechanisms by which H(2)S increases thermotolerance and lifespan in nematodes are conserved and that studies using C. elegans may help explain the beneficial effects observed in mammals exposed to H(2)S.

Sakube Y et al. (1994) Worm Breeder's Gazette "The Genome Structure and Expression of Promoter/lacZ Fusion Genes of C. elegans Ryanodine Receptor"

Imadzu H, Sakube Y, Kagawa H

[Worm Breeder's Gazette, 1994]

The Genome Structure and Expression of Promoter/lacZ Fusion Genes of the C. elegans Ryanodine Receptor Y. Sakube, H. Imadzu and H. Kagawa, Laboratory of Molecular Biology, Faculty of Science, Okayama University, Okayama, 700 Japan C51918@JPNKUDPC

Valles P et al. (2006) Semin Nephrol "Kidney vacuolar H+ -ATPase: physiology and regulation."

Wysocki J, Batlle D, Lapointe MS, Valles P

[Semin Nephrol, 2006]

The vacuolar H(+)-ATPase is a multisubunit protein consisting of a peripheral catalytic domain (V(1)) that binds and hydrolyzes adenosine triphosphate (ATP) and provides energy to pump H(+) through the transmembrane domain (V(0)) against a large gradient. This proton-translocating vacuolar H(+)-ATPase is present in both intracellular compartments and the plasma membrane of eukaryotic cells. Mutations in genes encoding kidney intercalated cell-specific V(0) a4 and V(1) B1 subunits of the vacuolar H(+)-ATPase cause the syndrome of distal tubular renal acidosis. This review focuses on the function, regulation, and the role of vacuolar H(+)-ATPases in renal physiology. The localization of vacuolar H(+)-ATPases in the kidney, and their role in intracellular pH (pHi) regulation, transepithelial proton transport, and acid-base homeostasis are discussed.

Rosas PC et al. (2016) PLoS One "Hsp72 (HSPA1A) Prevents Human Islet Amyloid Polypeptide Aggregation and Toxicity: A New Approach for Type 2 Diabetes Treatment."

Kaur P, Rosas PC, Wickman G, Panossian A, Al-Khamis FA, Nagaraja GM, Garcia LR, Asea AA

[PLoS One, 2016]

Type 2 diabetes is a growing public health concern and accounts for approximately 90% of all the cases of diabetes. Besides insulin resistance, type 2 diabetes is characterized by a deficit in -cell mass as a result of misfolded human islet amyloid polypeptide (h-IAPP) which forms toxic aggregates that destroy pancreatic -cells. Heat shock proteins (HSP) play an important role in combating the unwanted self-association of unfolded proteins. We hypothesized that Hsp72 (HSPA1A) prevents h-IAPP aggregation and toxicity. In this study, we demonstrated that thermal stress significantly up-regulates the intracellular expression of Hsp72, and prevents h-IAPP toxicity against pancreatic -cells. Moreover, Hsp72 (HSPA1A) overexpression in pancreatic -cells ameliorates h-IAPP toxicity. To test the hypothesis that Hsp72 (HSPA1A) prevents aggregation and fibril formation, we established a novel C. elegans model that expresses the highly amyloidogenic human pro-IAPP (h-proIAPP) that is implicated in amyloid formation and -cell toxicity. We demonstrated that h-proIAPP expression in body-wall muscles, pharynx and neurons adversely affects C. elegans development. In addition, we demonstrated that h-proIAPP forms insoluble aggregates and that the co-expression of h-Hsp72 in our h-proIAPP C. elegans model, increases h-proIAPP solubility. Furthermore, treatment of transgenic h-proIAPP C. elegans with ADAPT-232, known to induce the expression and release of Hsp72 (HSPA1A), significantly improved the growth retardation phenotype of transgenic worms. Taken together, this study identifies Hsp72 (HSPA1A) as a potential treatment to prevent -cell mass decline in type 2 diabetic patients and establishes for the first time a novel in vivo model that can be used to select compounds that attenuate h-proIAPP aggregation and toxicity.