Research Article

Translational Fusion of a G‐protein Coupled‐Receptor from the Hookworm Ancylostoma ceylanicum Expressed in Caenorhabditis elegans

Published: 2019-4

Journal: The FASEB Journal

DOI: 10.1096/fasebj.2019.33.1_supplement.649.8

Abstract

Hookworms are infectious parasites that affect more than 500 million people worldwide, causing extensive morbidity and economic burden in developing countries. Mass drug administration is a current control method, but resistance is of concern. Many hookworm species, including those in the genusAncylostoma, are specialists and require specific host signals to continue development upon infection. Little is known about the signaling mechanisms, although evolutionary conservation between parasitic nematodes and the model organismC. eleganssuggests that G‐protein coupled receptors (GPCRs) expressed in amphid neurons are chemosensory detection candidates. The amphid neurons ofC. elegansare exposed to the environment, even during the dauer stage, an alternate phase of larval development that is similar to the parasitic infective juvenile stage. Previous experiments involvingA. ceylanicumtranscriptional GFP fusions inC. elegansidentified putative GPCR genes expressed in neurons during dauer stage. To confirm that hookworm proteins are neuronally localized under dauer conditions, a translational GFP fusion of a putative hookworm GPCR was generated. The entire coding sequence, plus introns, was cloned using homologous recombination‐based, yeast assembly methods to create a C‐terminal GFP fusion. The putativeA. ceylanicumpromoter consisted of the upstream intergenic region, which is approximately 5000 base pairs, and the transcriptional terminator fromC. elegans unc‐54was used. To generate the expression plasmid, amplicons containing homology to the adjacent segment in the assembly were amplified using polymerase chain reaction (PCR) and transformed into yeast. The plasmid was verified by junction‐confirming PCR analysis, restriction enzyme mapping, and sequencing. After microinjection, GFP expression in amphid neurons of stably transformedC. elegansduring the dauer stage would support the hypothesis that this putative GPCR plays a role in reception of host signaling molecules and may provide a possible therapeutic target. Support or Funding Information This work was supported by the Salisbury University Faculty Mini‐Grant Program. This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published inThe FASEB Journal.

Faculty Members

  • Damien O'Halloran - Biological Sciences The George Washington University Washington DC
  • Ramesh Ratnappan - Microbiology, Immunology and Tropical Medicine The George Washington University Washington DC
  • Patti T. Erickson - Biological Sciences Salisbury University Salisbury MD
  • Myra Dickey - Biological Sciences Salisbury University Salisbury MD
  • Brandon Norman - Biological Sciences Salisbury University Salisbury MD
  • Mollie Jewell - Biological Sciences Salisbury University Salisbury MD
  • Lauren DeLong - Biological Sciences Salisbury University Salisbury MD
  • John M. Hawdon - Microbiology, Immunology and Tropical Medicine The George Washington University Washington DC

Themes

  • Hookworm Biology
  • G-Protein Coupled Receptors (GPCRs)
  • Global Health
  • Infectious Diseases
  • Therapeutic Targets
  • Host-Parasite Interactions
  • Developmental Biology

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