June 2017
Volume 58, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2017
Characteristics of zebrafish with morpholino knockdown of SYDE2, a candidate gene for optic atrophy
Author Affiliations & Notes
  • John Borchert
    Massachusetts Eye and Ear Infirmary, Boston, Massachusetts, United States
  • Angela Gauthier
    Massachusetts Eye and Ear Infirmary, Boston, Massachusetts, United States
  • Anna Larson
    Massachusetts Eye and Ear Infirmary, Boston, Massachusetts, United States
  • Janey L. Wiggs
    Massachusetts Eye and Ear Infirmary, Boston, Massachusetts, United States
  • Footnotes
    Commercial Relationships   John Borchert, None; Angela Gauthier, None; Anna Larson, None; Janey Wiggs, None
  • Footnotes
    Support  Funded in part by an award from the Alcon Research Institute
Investigative Ophthalmology & Visual Science June 2017, Vol.58, 2138. doi:
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      John Borchert, Angela Gauthier, Anna Larson, Janey L. Wiggs; Characteristics of zebrafish with morpholino knockdown of SYDE2, a candidate gene for optic atrophy. Invest. Ophthalmol. Vis. Sci. 2017;58(8):2138.

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      © ARVO (1962-2015); The Authors (2016-present)

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Abstract

Purpose : Inherited disorders of the optic nerve cause significant visual impairment in children and adults. While approximately 50% of patients with inherited forms of optic neuropathy have mutations in OPA1, mitochondrial DNA (LHON) or other known genes, the genetic etiology is unknown for many patients. We have completed whole exome sequencing (WES) that has identified mutations in Synapse Defective 1, Rho GTPase, Homolog 2 (SYDE2) in two families affected by recessively inherited optic atrophy. In this study we used morpholino-mediated knockdown of SYDE2 in zebrafish embryos to investigate possible roles for SYDE2 mutations in ocular development and optic atrophy pathogenesis.

Methods : Translation blocking (TB) and splice site blocking (SS) morpholinos were designed to knockdown SYDE2 using Genetools. 2.3 nl of 0.25mM morpholino solution was injected into the yolk of zebrafish embryos prior to reaching the eight cell stage to ensure homogenous uptake into all cells. An anti-p53 morpholino of the same concentration as the SYDE2 morpholinos (TB and SS) was co-injected to prevent off-target effects. Negative and uninjected controls were included for each injection. Embryos were raised in E3 media at 28 Celcius. Light microscopy photos were taken daily to track development with detailed measurements performed postmortem to maintain consistent orientation of the fry. Eye length (anterior to posterior), eye area, head area and body length were measured using ImageJ.

Results : Both the TB and SS blocking morpholinos resulted in fry that had statistically significant (P<0.05) smaller eye length, eye area and head area when compared to negative and uninjected controls. Head/eye length ratios were also statistically different from the controls, indicating that the eyes are disproportionately affected.

Conclusions : Morpholino-mediated knockdown of SYDE2 influences eye size in developing zebrafish. Further evaluation of the optic nerve in the morphants and rescue experiments using wild-type and mutant human cDNA are necessary to fully define a role for SYDE2 in optic nerve disease.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.

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