Molecular characterization of pannexins in the lens.
OBJECTIVE
Cell communication in the lens is critical for the life-long homeostasis of this tissue. Abundant gap junctions and cell-cell fusions are reported to be indispensable to the metabolic requirements and optical properties of the highly interconnected syncytial lens tissue. The expression of the recently characterized Panx1 and Panx2 gap junction proteins in the lens is, therefore, rather intriguing. Co-expression of pannexins and abundant connexins in the lens suggests that the two gap junction protein families have distinct roles in cell communication.


METHODS
Panx1 and Panx2 expression was studied by in situ hybridization and quantitative RT-PCR. We examined properties and tissue distribution of Panx1 isoforms by Western blot analysis. Immunohistochemistry was used to visualize lens regions that accumulate Panx1 to study intercellular localization and spatial relationship with lens connexin gap junctions.


RESULTS
Panx1 and Panx2 expression peaked in lens epithelial cells prior to differentiation. We detected one ubiquitously expressed Panx1 isoform and two additional isoforms that were only detected in the lens and the retina. Our results indicated that the ubiquitous 58 kDa and the oligomeric 120 kDa isoforms were plasma membrane-bound, resistant to Triton X-100 treatment, and was likely associated with cholesterol-enriched membrane microdomains. Immunohistochemistry revealed Panx1-specific punctuate labeling in the plasma membrane, and intensive labeling of the organelles in the epithelial and immature fiber cells. In addition, we detected Panx1 immunoreactivity in blood endothelial cells of the tunica vasculosa lentis capillaries and in blood erythrocytes.


CONCLUSIONS
Despite similarity in detergent solubility of pannexins and connexins, the lack of spatial co-localization in the lens membranes suggested a distinct, non-redundant to connexin function for these proteins in the membrane.
Dvoriantchikova
Galina
G
Bascom Palmer Eye Institute, Department of Ophthalmology, University of Miami Miller School of Medicine, Miami, FL 33136, USA. gdvoriantchikova@med.miami.edu <gdvoriantchikova@med.miami.edu>
Ivanov
Dmitry
D
Pestova
Anna
A
Shestopalov
Valery
V
D000818:Animals; D017630:Connexins / Q000235:genetics / Q000378:metabolism*; D004912:Erythrocytes / Q000378:metabolism; D006367:HeLa Cells; D006801:Humans; D007150:Immunohistochemistry / Q000379:methods; D007908:Lens, Crystalline / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D009419:Nerve Tissue Proteins / Q000097:blood / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013194:Staining and Labeling
D017630:Connexins; D009419:Nerve Tissue Proteins; C503631:Panx1 protein, mouse; C516139:Panx2 protein, mouse; D012333:RNA, Messenger