ZFIN ID: ZDB-PUB-081217-4
A modified acetylcholine receptor delta-subunit enables a null mutant to survive beyond sexual maturation
Epley, K.E., Urban, J.M., Ikenaga, T., and Ono, F.
Date: 2008
Source: The Journal of neuroscience : the official journal of the Society for Neuroscience   28(49): 13223-13231 (Journal)
Registered Authors: Ono, Fumihito, Urban, Jason
Keywords: zebrafish, neuromuscular junction, acetylcholine receptor, synapse, fetal akinesia deformation sequence, fluorescent protein
MeSH Terms:
  • Acetylcholine/metabolism
  • Animals
  • Animals, Genetically Modified
  • Feeding Behavior/physiology
  • Female
  • Gene Expression Regulation, Developmental/genetics
  • Longevity/genetics*
  • Luminescent Proteins/genetics
  • Male
  • Mutation/genetics*
  • Neuromuscular Junction/genetics
  • Neuromuscular Junction/metabolism
  • Neuromuscular Junction/physiopathology
  • Neuromuscular Junction Diseases/genetics*
  • Neuromuscular Junction Diseases/metabolism
  • Neuromuscular Junction Diseases/physiopathology
  • Protein Subunits/chemistry
  • Protein Subunits/genetics
  • Receptors, Cholinergic/genetics*
  • Recombinant Fusion Proteins/chemistry
  • Recombinant Fusion Proteins/genetics
  • Sexual Behavior, Animal/physiology
  • Sexual Maturation/genetics
  • Synaptic Membranes/genetics
  • Synaptic Membranes/metabolism
  • Synaptic Membranes/ultrastructure
  • Synaptic Transmission/genetics
  • Transgenes/genetics
  • Zebrafish/genetics*
  • Zebrafish/growth & development*
  • Zebrafish/metabolism
PubMed: 19052214 Full text @ J. Neurosci.
The contraction of skeletal muscle is dependent on synaptic transmission through acetylcholine receptors (AChRs) at the neuromuscular junction (NMJ). The lack of an AChR subunit causes a fetal akinesia in humans, leading to death in the first trimester and characteristic features of Fetal Akinesia Deformation Sequences (FADS). A corresponding null mutation of the delta-subunit in zebrafish (sofa potato; sop) leads to the death of embryos around 5 d postfertilization (dpf). In sop(-/-) mutants, we expressed modified delta-subunits, with one (delta1YFP) or two yellow fluorescent protein (delta2YFP) molecules fused at the intracellular loop, under the control of an alpha-actin promoter. AChRs containing these fusion proteins are fluorescent, assemble on the plasma membrane, make clusters under motor neuron endings, and generate synaptic current. We screened for germ-line transmission of the transgene and established a line of sop(-/-) fish stably expressing the delta2YFP. These delta2YFP/sop(-/-) embryos can mount escape behavior close to that of their wild-type siblings. Synaptic currents in these embryos had a smaller amplitude, slower rise time, and slower decay when compared with wild-type fish. Remarkably, these embryos grow to adulthood and display complex behaviors such as feeding and breeding. To the best of our knowledge, this is the first case of a mutant animal corresponding to first trimester lethality in human that has been rescued by a transgene and survived to adulthood. In the rescued fish, a foreign promoter drove the transgene expression and the NMJ had altered synaptic strength. The survival of the transgenic animal delineates requirements for gene therapies of NMJ.