Pharmacological Inhibition of Protein Geranylgeranyltransferase Type I Increases Aqueous Humor Outflow through the Trabecular Meshwork
P. Vasantha Rao 1*,
Yuri K Peterson 2,
Toshihiro Inoue 3,
and
Patrick J Casey 2
1 Department of Ophthalmology, Duke University Medical Center, Durham, North Carolina, United States
2 Pharmacology and Cancer Biology, Duke University Medical Center, Durham, North Carolina, United States
3 Ophthalmology, Duke University Medical Center, Durham, North Carolina, United States
* To whom correspondence should be addressed. E-mail: rao00011{at}mc.duke.edu.
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Abstract |
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Purpose: To determine the effects of inhibition of protein geranylgeranyltransferase type 1 (GGTase-I), which isoprenylates so-called CaaX proteins, which include GTP-binding proteins such as Rho GTPases and the 
subunits of heterotrimeric G-proteins, on aqueous humor outflow and trabecular meshwork cytoskeletal integrity.
Methods: A selective small molecular inhibitor of GGTase-I, GGTI-DU40 was tested in this study to investigate its effects on actin cytosketal integrity, cell adhesions, cell-cell junctions, myosin II phosphosphorylation, and membrane localization of GTP-binding proteins in trabecular meshwork (TM) cells, using immunofluorescence detection and immunobloting analysis. The effects of GGTI-DU40 on aqueous humor outflow were determined using organ cultured, perfused anterior segments of porcine eyes.
Results: In the TM cell lysates, GGTI-DU40 was confirmed to inhibit GGTase-I activity in a dose-dependent manner. TM cells treated with GGTI-DU40 displayed dose-dependent changes in cell morphology and reversible decreases in actin stress fibers, focal adhesions and adherens junctions. Myosin light chain phosphorylation was decreased significantly, and membrane localization of isoprenylated small GTPases and G was impaired in drug-treated TM cells. Aqueous outflow facility was increased significantly in eyes perfused with GGTI-DU40.
Conclusions: These data demonstrate that inhibition of geranylgeranyl isoprenylation of CaaX proteins in the aqueous outflow pathway increases aqueous humor outflow, possibly through altered cell adhesive interactions and actin cytoskeletal organization in cells of the outflow pathway. This study indicates that the GGTase-I enzyme a promising molecular target for lowering increased ocular pressure in glaucoma patients.
Key Words:
Isoprenylation, cytoskeleton, trabecular meshwork, Geranylgeranyltransferase, aqueous humor outflow, Rho GTPases
