June 2021
Volume 62, Issue 8
Open Access
ARVO Annual Meeting Abstract  |   June 2021
Generation and characterization of novel Dhdds knock-in mutation mouse models of RP59
Author Affiliations & Notes
  • Mai N. Nguyen
    Optometry and Vision Science, Vision Science Research Center, The University of Alabama at Birmingham, Birmingham, Alabama, United States
  • Dibyendu Chakraborty
    Optometry and Vision Science, Vision Science Research Center, The University of Alabama at Birmingham, Birmingham, Alabama, United States
  • Phillip Isaac Cobb
    Optometry and Vision Science, Vision Science Research Center, The University of Alabama at Birmingham, Birmingham, Alabama, United States
  • Steven J. Fliesler
    Ophthalmology and Biochemistry, SUNY The State University of New York, Buffalo, New York, United States
    Research Service, VA Western New York Healthcare System Buffalo VA Medical Center, Buffalo, New York, United States
  • Steven J Pittler
    Optometry and Vision Science, Vision Science Research Center, The University of Alabama at Birmingham, Birmingham, Alabama, United States
  • Footnotes
    Commercial Relationships   Mai Nguyen, None; Dibyendu Chakraborty, None; Phillip Cobb, None; Steven Fliesler, None; Steven Pittler, None
  • Footnotes
    Support  NIH/NEI (1 R01 EY029341; SJP, SJF) and P30 EY003039 (SJP), NIH/NCATS (1UL1 TR001412; SJF), VAWNYHS facilities and resources (SJF), VA Research Career Scientist Award/BLR&D (SJF)
Investigative Ophthalmology & Visual Science June 2021, Vol.62, 1464. doi:
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    • Get Citation

      Mai N. Nguyen, Dibyendu Chakraborty, Phillip Isaac Cobb, Steven J. Fliesler, Steven J Pittler; Generation and characterization of novel Dhdds knock-in mutation mouse models of RP59. Invest. Ophthalmol. Vis. Sci. 2021;62(8):1464.

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

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Abstract

Purpose : Dehydrodolichyl diphosphate synthase (DHDDS) is ubiquitously expressed and serves an essential function throughout the body, yet certain defects in the gene cause non-syndromic retinitis pigmentosa (RP59). To understand the basis for this retina selectivity, we generated and characterized two novel Dhdds knock-in mouse models: DhddsT206A/T206A and DhddsT206A/K42E.

Methods : Heterozygous (DhddsT206A/+) knock-in (KI) mice were generated on a C57Bl/6J background using CRISPR/Cas9 technology (UAB Transgenic & Genetically Engineered Models Core). DhddsT206A/+ littermates were crossed to generate Dhdds+/+, DhddsT206A/+, and DhddsT206A/T206A mice, and with DhddsK42E/K42E to generate DhddsT206A/K42E mice (to at least 3rd (F3) generation). All mutations were verified by PCR and sequence analysis. Phenotypes were assessed by SD-OCT and electroretinography (ERG). Retinal structure was assessed for DhddsT206A/T206A, DhddsT206A/+, DhddsT206A/K42E, and wild type (WT) mice (N=3-5) at PN 1 and 3 mo., while ERG was performed at PN 3 mo. Statistical analysis: Student t-test, with p<0.05 as significance threshold.

Results : SD-OCT showed no significant differences in retinal morphology, comparing mutant vs. WT mice. Outer nuclear layer (ONL) thickness (µm; mean±S.D.): DhddsT206A/T206A (55±5); DhddsT206A/+ (54±6); DhddsT206A/K42E (56±5); vs. WT (55±7). Total retinal thickness (µm): DhddsT206A/T206A (222±8); DhddsT206A/+ (220±8); DhddsT206A/K42E (220±9); vs. WT (222±6). DhddsT206A/+ ERG maximum response amplitudes at saturation were not significantly different from WT values under dark-adapted (scotopic) or light-adapted (photopic) conditions. However, photopic ERG amplitude of DhddsT206A/K42E mice were significantly different from WT values, and trending lower for DhddsT206A/T206A mice. Additionally, scotopic b-/a-wave ratios (values in parentheses) of DhddsT206A/T206A (1.2:1) and DhddsT206A/K42E (1.3:1), vs. WT (2:1) mice were significantly diminished.

Conclusions : Homozygous (T206A/T206A) and compound heterozygous (K42E/T206A) Dhdds knock-in mouse models of RP59 exhibit negative ERGs under light- and dark-adapted conditions, indicative of defective photoreceptor-to-bipolar cell synaptic transmission. There may be common retina-specific target(s) that could explain the lack of systemic effects associated with DHDDS mutations in RP59.

This is a 2021 ARVO Annual Meeting abstract.

 

ERG maximum saturating response averages. *p≤0.05

ERG maximum saturating response averages. *p≤0.05

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