Distribution of bovine and rabbit lens alpha-crystallin products by MALDI imaging mass spectrometry.
OBJECTIVE
To develop a general tissue preparation protocol for MALDI (Matrix-Assisted Laser Desorption Ionization) imaging mass spectrometry of ocular lens tissue, and to compare the spatial distributions of alpha-crystallin and its modified forms in bovine and rabbit lenses.


METHODS
Frozen bovine and rabbit lenses were cryosectioned equatorially at -20 degrees C into 12 microm-thick tissue sections. Lens sections were mounted onto conductive glass slides by ethanol soft-landing to maintain tissue integrity. An ethanol/xylene washing procedure was applied to each section before matrix application to facilitate uniform matrix crystal formation across the entire tissue section. Molecular images of both alpha-crystallin subunits and their modified forms were obtained from mass spectral data acquired at 100 microm steps across both whole rabbit and half bovine lens sections.


RESULTS
Distinct spatial patterns for the two subunits of alpha-crystallin and their modified forms were observed in the rabbit and bovine lens sections. While alphaA-crystallin was extensively degraded in the lens core of both species, rabbit lenses exhibited a greater degree of larger molecular weight truncation products. In contrast, alphaB-crystallin degradation was limited in both species. Interestingly, phosphorylation of alphaA- and alphaB-crystallin was most abundant in the middle cortex of both species.


CONCLUSIONS
An improved method for investigating the spatial distribution of alpha-crystallin in the ocular lens by MALDI imaging mass spectrometry has been developed. The localization of multiple degradation products and specific regions of alpha-crystallin phosphorylation in bovine and rabbit lenses gives new insight into the program of lens fiber cell differentiation and normal lens function.
Grey
Angus C
AC
Department of Cell and Molecular Pharmacology, Medical University of South Carolina, Charleston, SC 29425-5050, USA.
Schey
Kevin L
KL
D000818:Animals; D002417:Cattle; D007908:Lens, Crystalline / Q000166:cytology / Q000378:metabolism*; D010446:Peptide Fragments / Q000737:chemistry / Q000378:metabolism; D010766:Phosphorylation; D011499:Protein Processing, Post-Translational; D011817:Rabbits; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D038201:alpha-Crystallins / Q000737:chemistry / Q000378:metabolism*
D010446:Peptide Fragments; D038201:alpha-Crystallins