May 2008
Volume 49, Issue 13
Free
ARVO Annual Meeting Abstract  |   May 2008
Identification and Characterization of Tubby-Like Protein 1 (Tulp1) as a Phagocytic Ligand in RPE Cells
Author Affiliations & Notes
  • N. B. Caberoy
    Bascom Palmer Eye Institute, University of Miami Miller School of Medicine, Miami, Florida
  • D. Maiguel
    Bascom Palmer Eye Institute, University of Miami Miller School of Medicine, Miami, Florida
  • Y. Kim
    Bascom Palmer Eye Institute, University of Miami Miller School of Medicine, Miami, Florida
  • W. Li
    Bascom Palmer Eye Institute, University of Miami Miller School of Medicine, Miami, Florida
  • Footnotes
    Commercial Relationships  N.B. Caberoy, None; D. Maiguel, None; Y. Kim, None; W. Li, None.
  • Footnotes
    Support  NIHR01EY016211, P30EY014801
Investigative Ophthalmology & Visual Science May 2008, Vol.49, 4442. doi:https://doi.org/
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    • Get Citation

      N. B. Caberoy, D. Maiguel, Y. Kim, W. Li; Identification and Characterization of Tubby-Like Protein 1 (Tulp1) as a Phagocytic Ligand in RPE Cells. Invest. Ophthalmol. Vis. Sci. 2008;49(13):4442. doi: https://doi.org/.

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

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Abstract

Purpose: : : Retinal pigment epithelium (RPE) cells play a critical role in the regeneration of photoreceptor outer segment (POS) by removing shed POS through phagocytosis. The molecular mechanism of RPE phagocytosis is poorly defined. Despite the identification of a few phagocytic receptors, an intriguing question is whether there is any phagocytic protein ligand for RPE phagocytosis. The purpose of this study is to identify and characterize RPE phagocytic protein ligands and to define their roles in retinal degeneration.

Methods: : A novel functional cloning strategy of phage display has been developed to screen mouse eye cDNA library in human ARPE-19 cell line. After four rounds of selection, individual phage clones with higher phagocytic activity than control phage were isolated and identified by DNA sequencing. The minimal phagocytic domains were defined functionally by a unique phage-based phagocytic assay (PPA). Their phagocytic activities were further validated and characterized by non-phage-based phagocytic assays.

Results: : Among several identified phagocytic ligands was Tulp179-199. Mutations in human Tulp1 have been previously reported to associate with retinitis pigmentosa 14. Deletion and mutation analyses by PPA revealed that full-length Tulp1 consists of five minimal phagocytic domains in its N-terminal region. No phagocytic activity was detected in its C-terminus. Tulp1 phagocytic activity was demonstrated in all RPE cells tested, including RPE primary culture cells, APRE-19, D407 and RPE-J cell lines, but not in non-RPE epithelial cell lines, such as HeLa and MCF-7 cells. Physiological and pathological relevance of Tulp1 phagocytic activity was further established.

Conclusions: : Tulp1 has been identified as the first phagocytic protein ligand in RPE cells by the innovative functional cloning strategy of phage display, and the role of its phagocytic activity in retinal degeneration has been demonstrated. The novel functional cloning strategy of phage display described in this study is a powerful tool to investigate molecular mechanisms of phagocytosis in RPE and other professional phagocytic cells.

Keywords: retinal pigment epithelium • retinal degenerations: cell biology • phagocytosis and killing 
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