April 2009
Volume 50, Issue 13
ARVO Annual Meeting Abstract  |   April 2009
Identification of Retinal Pro-Survival Wnt Regulator Protein Dickkopf3 (Dkk3) Interactors
Author Affiliations & Notes
  • R. E. Nakamura
    Ophthalmology, University of Miami, Miami, Florida
  • S. K. Bhattacharya
    Ophthalmology, University of Miami, Miami, Florida
  • A. S. Hackam
    Ophthalmology, University of Miami, Miami, Florida
  • Footnotes
    Commercial Relationships  R.E. Nakamura, None; S.K. Bhattacharya, None; A.S. Hackam, None.
  • Footnotes
    Support  Karl Kirchgessner Foundation, Lois Pope Life Fellowship, RPB Career Dev Award,RPB Unrestricted grant (BPEI),NIH Grant EY017837, NIH core grant P30EY014801
Investigative Ophthalmology & Visual Science April 2009, Vol.50, 6158. doi:
  • Views
  • Share
  • Tools
    • Alerts
      This feature is available to authenticated users only.
      Sign In or Create an Account ×
    • Get Citation

      R. E. Nakamura, S. K. Bhattacharya, A. S. Hackam; Identification of Retinal Pro-Survival Wnt Regulator Protein Dickkopf3 (Dkk3) Interactors. Invest. Ophthalmol. Vis. Sci. 2009;50(13):6158.

      Download citation file:

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

  • Supplements

Purpose: : The Wnt pathway is a critical cellular communication pathway involved in retinal development and retinal degeneration. Previously, we demonstrated that activation of the Wnt signaling pathway in Muller glia protects photoreceptors from oxidative stress (Yi, et al, 2007). Muller glia secrete Dkk3, a Wnt pathway protein that potentiates Wnt3a-mediated signaling and protects several cell lines from exogenous insults (Nakamura et al, 2007). The purpose of this study is to identify the interacting proteins that mediate these properties of Dkk3 to further understand the role of Dkk3 during retinal degeneration.

Methods: : GST-pull down, co-immunoprecipitation, and bis[sylfosuccinimidyl]suberate (BS3) cross linking were performed to test Dkk3 interaction with the Wnt regulators Krm1 and Krm2. FLAG-Dkk3 pull down or blot overlay followed by MS/MS analysis were carried out using human Muller glia cell line, mouse retina and brain lysates to identify novel interacting proteins. The interactions were verified via confocal analysis and TOP-FLASH luciferase Wnt reporter assays.

Results: : Our data showed that Dkk3 interacts with both Krm1 and Krm2 but not with low density related lipoprotein 6 receptor (LRP6) when these genes were co-expressed (n=5). Interestingly, co-expression of these genes did not modulate Wnt signaling activity measured via luciferase assay (n=5, p<NS). Furthermore, we demonstrated by blot overlay that Dkk3 interacts with cysteine and glycine rich protein 2 (CRP2) and co-expression of these genes enhanced Wnt3a-mediated Wnt signaling (n=3, p<0.01).

Conclusions: : Dkk3 interacts with the known Wnt regulators Krm1 and Krm2, and the non-Wnt pathway related protein CRP2. Since CRP2 is reported to interact with protein inhibitor of activated STAT1 (PIAS1), Dkk3 may mediate its pro-survival properties via a non-canonical pathway, most likely through the STAT pathway.

Keywords: Muller cells • protein purification and characterization • neuroprotection 

This PDF is available to Subscribers Only

Sign in or purchase a subscription to access this content. ×

You must be signed into an individual account to use this feature.