Lectin microarray profiling and relative quantification of glycome associated with proteins of neonatal wt and rd1 mice retinae

Invest Ophthalmol Vis Sci. 2013 May 7;54(5):3272-80. doi: 10.1167/iovs.12-11363.

Abstract

Purpose: Progressive dynamic, relative quantitative changes were compared in glycans associated with retinal proteins of wild type (wt) and retinal degeneration 1 (rd1) mice during neonatal development and degeneration of retinae.

Methods: Proteins extracted from retinae of postnatal days 2 (PN2), PN7, and PN14 wt and rd1 mice were labeled with Cy3-fluorescent dye. Glycome of these proteins was quantified relatively by lectin microarray technique. Net fluorescence emitted by individual complexes formed between 45 lectins and Cy3-labeled proteins was measured by evanescent-field fluorescence-assisted microarray reader.

Results: GlcNAcβ1-oligomer and high-mannose/Manα1-6Man were major glycans associated with the proteins of PN2, PN7, and PN14 wt and rd1 mice retinae. Gal/GalNAc/Man3-core-bi-/tri-antennary-complex, Sia2-3Galβ1-4GlcNAc, and high-mannose glycans were conjugated mainly to proteins from PN7 rd1 and PN14 wt retinae, respectively. With increasing neonatal age, mannosylated, GlcNAcβ, and sialylated (minor component) glycans were increased, and fucosylated GlcNAc/Galβ glycans were decreased significantly in wt retinal proteins. This trend was less evident in PN14 rd1 retinal proteins. Mouse retina was almost devoid of Siaα2-6 (except WGA bound Sia), Fucα1-2, and Gal/GalNAc-containing glycans. STL reacting GlcNAc oligomers were high in PN2 rd1 retinae.

Conclusions: Quantitative dynamic, relative variation in high-mannose and GlcNAc glycans, Siaα2-3Galβ1-4GlcNAc associated with proteins from PN2, PN7, and PN14 wt and rd1 mice retinae suggested that these glycans participate in retinal development and degeneration, and may be used as markers for retinal electrophysiologic integrity during transplantation/therapy studies; Siaα2-3Galβ1-4GlcNAc-specific Agrocybe cylindracea lectin and other lectins may be used to enrich/purify retinal ribbon synapse glycoproteins and other glycoproteins including rhodopsin. Further investigations are required.

Keywords: glycome quantification; lectin microarray; rd1 mouse; retinal degeneration; retinal development.

Publication types

  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Animals, Newborn
  • Carbocyanines / metabolism
  • Disease Models, Animal
  • Eye Proteins / metabolism*
  • Fluorescent Dyes / metabolism
  • Glycomics
  • Glycoproteins / metabolism*
  • Lectins / metabolism*
  • Mice
  • Polysaccharides / metabolism*
  • Protein Array Analysis
  • Retina / metabolism*
  • Retinitis Pigmentosa / metabolism*

Substances

  • Carbocyanines
  • Eye Proteins
  • Fluorescent Dyes
  • Glycoproteins
  • Lectins
  • Polysaccharides
  • cyanine dye 3