July 2019
Volume 60, Issue 9
Open Access
ARVO Annual Meeting Abstract  |   July 2019
DCTN5 mutant mice reveal a role for dynactin in lens biogenesis
Author Affiliations & Notes
  • Ting-Yu Yeh
    Biology, Johns Hopkins University, Baltimore, Maryland, United States
  • Emily Zion
    Biology, Johns Hopkins University, Baltimore, Maryland, United States
  • Ann Caverts
    Biology, Johns Hopkins University, Baltimore, Maryland, United States
  • Manish Ayushman
    Biology, Johns Hopkins University, Baltimore, Maryland, United States
    Chemical Engineering, Indian Institute of Technology Kharagpur, Kharagpur, West Bengal, India
  • Frederick Dong
    Biology, Johns Hopkins University, Baltimore, Maryland, United States
  • Haiqing Zhao
    Biology, Johns Hopkins University, Baltimore, Maryland, United States
  • Trina Schroer
    Biology, Johns Hopkins University, Baltimore, Maryland, United States
  • Footnotes
    Commercial Relationships   Ting-Yu Yeh, None; Emily Zion, None; Ann Caverts, None; Manish Ayushman, None; Frederick Dong, None; Haiqing Zhao, None; Trina Schroer, None
  • Footnotes
    Support  NIH Grant GM 44589; HL137076
Investigative Ophthalmology & Visual Science July 2019, Vol.60, 5686. doi:
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      Ting-Yu Yeh, Emily Zion, Ann Caverts, Manish Ayushman, Frederick Dong, Haiqing Zhao, Trina Schroer; DCTN5 mutant mice reveal a role for dynactin in lens biogenesis. Invest. Ophthalmol. Vis. Sci. 2019;60(9):5686.

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

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Abstract

Purpose : Lens clarity depends on proper interactions of lens fiber cells with each other and the overlying epithelium. A great is known about the role of the actin cytoskeleton in lens organization, but how microtubules and their associated proteins contribute to lens biology remains poorly defined. We explored this question using a novel mutation in the mouse DCTN5 gene, b2b315Clo (henceforth called DCTN5-315).

Methods : Refractive defects of lenses from 57 wild type and 47 DCTN5-315 C57/BL6 and DBA mice (P18-P56, 59 male, 45 female) were inspected. Whole lens F-actin was stained with Alexa568-phalloidin. Live cell imaging of RFP-F-tractin and Lum-mCherry was performed in cells expressing DCTN5-315 and wild type p25.

Results : DCTN5-315 is a point mutation in an intron of DCTN5, the gene that encodes the p25 component of dynactin, an essential component of the cytoplasmic dynein motor. DCTN5-315 homozygotes die at ≈ e16, but heterozygotes are viable and fertile. DCTN5-315 transcripts are differentially spliced to yield a truncated p25 isoform that differs at the C-terminus, a part of p25 implicated in cargo interactions. The truncated p25 protein incorporates normally into dynactin and does not affect dynactin abundance or stability. Heterozygous DCTN5 knockout mice exhibit congenital cataracts with high penetrance but this is rare in DCTN5-315 heterozygotes. However, DCTN5-315 heterozygous lenses are more susceptible to temperature-dependent cataract formation and show refractive index discontinuities suggesting abnormal cell organization. Fluorescent phalloidin staining reveals defects in epithelial cell organization and aberrant cortical actin deposition. Cultured cells expressing the truncated form in place of WT p25 exhibit aberrant actin dynamics and cortical ruffling, as well as promiscuous primary cilia formation. By contrast, post-Golgi carrier movement is normal, indicating that the truncated form of p25 retains partial function.

Conclusions : DCTN5-315 heterozygous mice may represent a sensitized background for cataracts formation. Our data suggest that the p25 C-terminus is required for proper regulation of actin dynamics and primary ciliogenesis. Our work provides the first view of the role played by cytoplasmic dynein in lens cell biology, and support a model wherein this essential motor contributes in a novel way to maintenance of the cortical structures required for normal cell-cell interactions.

This abstract was presented at the 2019 ARVO Annual Meeting, held in Vancouver, Canada, April 28 - May 2, 2019.

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