Ebbing A et al. (2019) Development "Partially overlapping guidance pathways focus the activity of UNC-40/DCC along the anteroposterior axis of polarizing neuroblasts."

Middelkoop TC, Bodewes E, Betist M, Ebbing A, Korswagen HC

[Development, 2019]

Directional migration of neurons and neuronal precursor cells is a central process in nervous system development. In the nematode <i>C. elegans</i>, the two Q neuroblasts polarize and migrate in opposite directions along the anteroposterior body axis. Several key regulators of Q cell polarization have been identified, including MIG-21, DPY-19/DPY19L1, the netrin receptor UNC-40/DCC, the Fat-like cadherin CDH-4, and CDH-3/Fat, which we describe in this study. How these different transmembrane proteins act together to direct Q neuroblast polarization and migration is still largely unknown. Here, we demonstrate that MIG-21 and DPY-19, CDH-3 and CDH-4, and UNC-40 define three distinct pathways that have partially redundant roles in protrusion formation, but also separate functions in regulating protrusion direction. Moreover, we show that the MIG-21 - DPY-19 and Fat-like cadherin pathways control the localization and clustering of UNC-40 at the leading edge of the polarizing Q neuroblast, and that this is independent of the UNC-40 ligands UNC-6/netrin and MADD-4. Our results provide insight into a novel mechanism for ligand-independent localization of UNC-40 that directs the activity of UNC-40 along the anteroposterior axis.

Ebbing A et al. (2018) Dev Cell "Spatial Transcriptomics of C.elegans Males and Hermaphrodites Identifies Sex-Specific Differences in Gene Expression Patterns."

Junker JP, Ebbing A, Korswagen HC, Spanjaard B, Berezikov E, Betist MC, van Oudenaarden A, Vertesy A

[Dev Cell, 2018]

To advance our understanding of the genetic programs that drive cell and tissue specialization, it is necessary to obtain a comprehensive overview of gene expression patterns. Here, we have used spatial transcriptomics to generate high-resolution, anteroposterior gene expression maps of C.elegans males and hermaphrodites. To explore these maps, we have developed computational methods for discovering region- and tissue-specific genes. We have found extensive sex-specific gene expression differences in the germline and sperm and discovered genes that are specifically expressed in the male reproductive tract. These include a group of uncharacterized genes that encode small secreted proteins that are required for male fertility. We conclude that spatial gene expression maps provide a powerful resource for identifying tissue-specific gene functions in C.elegans. Importantly, we found that expression maps from different animals can be precisely aligned, enabling transcriptome-wide comparisons of gene expression patterns.

Goodwin EB et al. (1997) Development "A genetic pathway for regulation of tra-2 translation."

Mango SE, Goodwin EB, Hurney CA, Hofstra K, Kimble JE

[Development, 1997]

In Caenorhabditis elegans, the tra-2 sex-determining gene is regulated at the translational level by two 28 nt direct repeat elements (DREs) located in its 3' untranslated region (3'UTR). DRF is a factor that binds the DREs and may be a trans-acting translational regulator of tra-2. Here we identify two genes that are required for the normal pattern of translational control. A newly identified gene, called laf-1, is required for translational repression by the tra-2 3'UTR. In addition, the sex-determining gene, tra-3, appears to promote female development by freeing tra-2 from laf-1 repression. Finally, we show that DRF activity correlates with translational repression of tra-2 during development and that tra-3 regulates DRF activity. We suggest that tra-3 may promote female development by releasing tra-2 from translation repression by laf-1 and that translational control is important for proper sex determination--both in the early embryo and during postembryonic development.

Goodwin EB et al. (1997) Semin Cell Dev Biol "Translational control of development in C. elegans."

Goodwin EB, Evans TC

[Semin Cell Dev Biol, 1997]

Translational control by the 3'untranslated regions (3'UTRs) of mRNAs contributes to important events throughout the development of C. elegans. In oocytes and early embryos, maternal mRNAs are controlled by 3'UTR elements to restrict translation of their protein products to specific blastomeres. Localized translation is probably critical for specifying blastomere identity. In both germline and somatic cells, mRNAs from sex determining genes are translationally repressed by 3'UTR controls. These controls balance the activities that specify male and female cell fates. During larval development, the temporal sequence of cell lineages requires 3'UTR-mediated regulation of heterochronic genes by a small non-protein coding RNA. We review what is known about these translational control mechanisms in C. elegans. This overview illustrates that translational control by 3'UTR elements is a powerful mechanism for regulating the expression of multiple gene products in diverse cell types during development of a multi-cellular animal.Copyright 1997 Academic Press Limited.

Kayser EB et al. (2001) J Biol Chem "Mitochondrial expression and function of GAS-1 in Caenorhabditis elegans."

Morgan PG, Sedensky MM, Kayser EB, Hoppel CL

[J Biol Chem, 2001]

A mutation in the gene gas-1 alters sensitivity to volatile anesthetics, fecundity, and life span in the nematode Caenorhabditis elegans. gas-1 encodes a close homologue of the 49-kDa iron protein subunit of Complex I of the mitochondrial electron transport chain from bovine heart. gas-1 is widely expressed in the nematode neuromuscular system and in a subcellular pattern consistent with that of a mitochondrial protein. Pharmacological studies indicate that gas-1 functions partially via presynaptic effects. In addition, a mutation in the gas-1 gene profoundly decreases Complex I-dependent metabolism in mitochondria as measured by rates of both oxidative phosphorylation and electron transport. An increase in Complex II-dependent metabolism also is seen in mitochondria from gas-1 animals. There is no apparent alteration in physical structure in mitochondria from gas-1 nematodes compared with those from wild type. These data indicate that gas-1 is the major 49-kDa protein of complex I and that the GAS-1 protein is critical to mitochondrial function in C. elegans. They also reveal the importance of mitochondrial function in determining not only aging and life span, but also anesthetic sensitivity, in this model organism.

Maryon EB et al. (1998) J Cell Sci "Muscle-specific functions of ryanodine receptor channels in Caenorhabditis elegans."

Maryon EB, Anderson P, Saari B

[J Cell Sci, 1998]

Ryanodine receptor channels regulate contraction of striated muscle by gating the release of calcium ions from the sarcoplasmic reticulum. Ryanodine receptors are expressed in excitable and non-excitable cells of numerous species, including the nematode C. elegans. Unlike vertebrates, which have at least three ryanodine receptor genes, C. elegans has a single gene encoded by the unc-68 locus. We show that unc-68 is expressed in most muscle cells, and that the phenotypic defects exhibited by unc-68 null mutants result from the loss of unc-68 function in pharyngeal and body-wall muscle cells. The loss of unc-68 function in the isthmus and terminal bulb muscles of the pharynx causes a reduction in growth rate and brood size. unc-68 null mutants exhibit defective pharyngeal pumping (feeding) and have abnormal vacuoles in the terminal bulb of the pharynx. unc-68 is required in body-wall muscle cells for normal motility. We show that UNC-68 is localized in body-wall muscle cells to flattened vesicular sacs positioned between the apical plasma membrane and the myofilament lattice. In unc-68 mutants, the vesicles are enlarged and densely stained. The flattened vesicles in body-wall muscle cells thus represent the C. elegans sarcoplasmic reticulum. Morphological and behavioral phenotypes of unc-68 mutants suggest that intracellular calcium release is not essential for excitation-contraction coupling in C. elegans.

Kayser EB et al. (2004) Anesthesiology "Mitochondrial complex I function affects halothane sensitivity in Caenorhabditis elegans."

Morgan PG, Kayser EB, Sedensky MM

[Anesthesiology, 2004]

BACKGROUND:: The gene gas-1 encodes a subunit of complex I of the mitochondrial electron transport chain in Caenorhabditis elegans. A mutation in gas-1 profoundly increases sensitivity of C. elegans to volatile anesthetics. It is unclear which aspects of mitochondrial function account for the hypersensitivity of the mutant. METHODS:: Oxidative phosphorylation was determined by measuring mitochondrial oxygen consumption using electron donors specific for either complex I or complex II. Adenosine triphosphate concentrations were determined by measuring luciferase activity. Oxidative damage to mitochondrial proteins was identified using specific antibodies. RESULTS:: Halothane inhibited oxidative phosphorylation in isolated wild-type mitochondria within a concentration range that immobilizes intact worms. At equal halothane concentrations, complex I activity but not complex II activity was lower in mitochondria from mutant (gas-1) animals than from wild-type (N2) animals. The halothane concentrations needed to immobilize 50% of N2 or gas-1 animals, respectively, did not reduce oxidative phosphorylation to identical rates in the two strains. In air, adenosine triphosphate concentrations were similar for N2 and gas-1 but were decreased in the presence of halothane only in gas-1 animals. Oxygen tension changed the sensitivity of both strains to halothane. When nematodes were raised in room air, oxidative damage to mitochondrial proteins was increased in the mutant animal compared with the wild type. CONCLUSIONS:: Rates of oxidative phosphorylation and changes in adenosine triphosphate concentrations by themselves do not control anesthetic-induced immobility of wild-type C. elegans. However, they may contribute to the increased sensitivity to volatile anesthetics of the gas-1 mutant. Oxidative damage to proteins may be an important contributor to sensitivity to volatile anesthetics in C. elegans.

Parra-Millan R et al. (2018) mSphere "Intracellular Trafficking and Persistence of Acinetobacter baumannii Requires Transcription Factor EB."

Pachon J, Parra-Millan R, Pachon-Ibanez ME, Ayerbe-Algaba R, Guerrero-Gomez D, Miranda-Vizuete A, Smani Y

[mSphere, 2018]

Acinetobacter baumannii is a significant human pathogen associated with hospital-acquired infections. While adhesion, an initial and important step in A.baumannii infection, is well characterized, the intracellular trafficking of this pathogen inside host cells remains poorly studied. Here, we demonstrate that transcription factor EB (TFEB) is activated after A.baumannii infection of human lung epithelial cells (A549). We also show that TFEB is required for the invasion and persistence inside A549 cells. Consequently, lysosomal biogenesis and autophagy activation were observed after TFEB activation which could increase the death of A549 cells. In addition, using the Caenorhabditis elegans infection model by A.baumannii, the TFEB orthologue HLH-30 was required for survival of the nematode to infection, although nuclear translocation of HLH-30 was not required. These results identify TFEB as a conserved key factor in the pathogenesis of A.baumannii.<b>IMPORTANCE</b>Adhesion is an initial and important step in Acinetobacter baumannii infections. However, the mechanism of entrance and persistence inside host cells is unclear and remains to be understood. In this study, we report that, in addition to its known role in host defense against Gram-positive bacterial infection, TFEB also plays an important role in the intracellular trafficking of A.baumannii in host cells. TFEB was activated shortly after A.baumannii infection and is required for its persistence within host cells. Additionally, using the C.elegans infection model by A.baumannii, the TFEB orthologue HLH-30 was required for survival of the nematode to infection, although nuclear translocation of HLH-30 was not required.

Kayser EB et al. (1999) Anesthesiology "GAS-1: a mitochondrial protein controls sensitivity to volatile anesthetics in the nematode Caenorhabditis elegans."

Sedensky MM, Morgan PG, Kayser EB

[Anesthesiology, 1999]

BACKGROUND: Mutations in several genes of Caenorhabditis elegans confer altered sensitivities to volatile anesthetics. A mutation in one gene, gas-1(fc21), causes animals to be immobilized at lower concentrations of all volatile anesthetics than in the wild-type, and it does not depend on mutations in other genes to control anesthetic sensitivity. gas-1 confers different sensitivities to stereoisomers of isoflurane, and thus may be a direct target for volatile anesthetics. The authors have cloned and characterized the gas-1 gene and the mutant allele fc21. METHODS: Genetic techniques for nematodes were as previously described. Polymerase chain reaction, sequencing, and other molecular biology techniques were performed by standard methods. Mutant rescue was done by injecting DNA fragments into the gonad of mutant animals and scoring the offspring for loss of the mutant phenotype. RESULTS: The gas-1 gene was cloned and identified. The protein GAS-1 is a homologue of the 49-kDa (IP) subunit of the mitochondrial NADH:ubiquinone-oxidoreductase (complex I of the respiratory chain). gas-1(fc21) is a missense mutation replacing a strictly conserved arginine with lysine. CONCLUSIONS: The function of the 49-kDa (IP) subunit of complex I is unknown. The finding that mutations in complex I increase sensitivity of C elegans to volatile anesthetics may implicate this physiologic process in the determination of anesthetic sensitivity. The hypersensitivity of animals with a mutation in the gas-1 gene may be caused by a direct anesthetic effect on a mitochondrial protein or secondary effects at other sites caused by mitochondrial dysfunction.

Espiritu EB et al. (2012) Dev Biol "CLASPs function redundantly to regulate astral microtubules in the C. elegans embryo."

Espiritu EB, Rose LS, Krueger LE, Ye A

[Dev Biol, 2012]

Microtubule dynamics are thought to play an important role in regulating microtubule interactions with cortical force generating motor proteins that position the spindle during asymmetric cell division. CLASPs are microtubule-associated proteins that have a conserved role in regulating microtubule dynamics in diverse cell types. Caenorhabditis elegans has three CLASP homologs in its genome. CLS-2 is known to localize to kinetochores and is needed for chromosome segregation at meiosis and mitosis; however CLS-1 and CLS-3 have not been reported to have any role in embryonic development. Here, we show that depletion of CLS-2 in combination with either CLS-1 or CLS-3 results in defects in nuclear rotation, maintenance of spindle length, and spindle displacement in the one-cell embryo. Polarity is normal in these embryos, but reduced numbers of astral microtubules reach all regions of the cortex at the time of spindle positioning. Analysis of the microtubule plus-end tracker EB1 also revealed a reduced number of growing microtubules reaching the cortex in CLASP depleted embryos, but the polymerization rate of astral microtubules was not slower than in wild type. These results indicate that C. elegans CLASPs act partially redundantly to regulate astral microtubules and position the spindle during asymmetric cell division. Further, we show that these spindle pole-positioning roles are independent of the CLS-2 binding proteins HCP-1 and HCP-2.