RPGR isoforms in photoreceptor connecting cilia and the transitional zone of motile cilia.
OBJECTIVE
The retinitis pigmentosa guanosine triphosphatase (GTPase) regulator (RPGR) is essential for photoreceptor survival. There is as yet no consensus concerning the subcellular localization of RPGR. This study was undertaken as a comprehensive effort to resolve current controversies.


METHODS
RPGR in mice and other mammalian species was examined by immunofluorescence. RPGR variants were distinguished by using isoform-specific antibodies. Different tissue processing procedures were evaluated. Immunoblot analysis of serial cross-sections of photoreceptors was performed as a complementary approach to subcellular localization.


RESULTS
RPGR was found in the connecting cilia of rods and cones with no evidence for species-dependent variation. RPGR ORF15 was the predominant variant in photoreceptor connecting cilia whereas constitutive RPGR (default) was the sole variant in the transitional zone of motile cilia in airway epithelia. Removal of soluble materials in the interphotoreceptor matrix facilitated detection of RPGR in the connecting cilia in photoreceptors.


CONCLUSIONS
RPGR localizes in photoreceptor connecting cilia and in a homologous structure, the transitional zone of motile cilia. These data are important for understanding the multitude of clinical manifestations associated with mutations in RPGR. Interphotoreceptor matrix surrounding the connecting cilia is a key variable for in situ detection of a protein in the connecting cilia.
Hong
Dong-Hyun
DH
Berman-Gund Laboratory for the Study of Retinal Degenerations, Harvard Medical School, Boston, Massachusetts 02114, USA.
Pawlyk
Basil
B
Sokolov
Maxim
M
Strissel
Katherine J
KJ
Yang
Jun
J
Tulloch
Brian
B
Wright
Alan F
AF
Arshavsky
Vadim Y
VY
Li
Tiansen
T
D000818:Animals; D002352:Carrier Proteins / Q000378:metabolism*; D002417:Cattle; D002923:Cilia / Q000378:metabolism; D005136:Eye Proteins; D019084:Fluorescent Antibody Technique, Indirect; D015151:Immunoblotting; D051379:Mice; D008810:Mice, Inbred C57BL; D008856:Microscopy, Fluorescence; D020419:Photoreceptor Cells, Vertebrate / Q000378:metabolism*; D020033:Protein Isoforms; D013552:Swine
D002352:Carrier Proteins; D005136:Eye Proteins; D020033:Protein Isoforms; C099929:RPGR protein, human