FOXC1 modulates MYOC secretion through regulation of the exocytic proteins RAB3GAP1, RAB3GAP2 and SNAP25

Abstract

The neurodegenerative disease glaucoma is one of the leading causes of blindness in the world. Glaucoma is characterized by progressive visual field loss caused by retinal ganglion cell (RGC) death. Both surgical glaucoma treatments and medications are available, however, they only halt glaucoma progression and are unable to reverse damage. Furthermore, many patients do not respond well to treatments. It is therefore important to better understand the mechanisms involved in glaucoma pathogenesis. Patients with Axenfeld-Rieger syndrome (ARS) offer important insight into glaucoma progression. ARS patients are at 50% risk of developing early onset glaucoma and respond poorly to treatments, even when surgical treatments are combined with medications. Mutations in the transcription factor FOXC1 cause ARS. Alterations in FOXC1 levels cause ocular malformations and disrupt stress response in ocular tissues, thereby contributing to glaucoma progression. In this study, using biochemical and molecular techniques, we show that FOXC1 regulates the expression of RAB3GAP1, RAB3GAP2 and SNAP25, three genes with central roles in both exocytosis and endocytosis, responsible for extracellular trafficking. FOXC1 positively regulates RAB3GAP1 and RAB3GAP2, while either increase or decrease in FOXC1 levels beyond its normal range results in decreased SNAP25. In addition, we found that FOXC1 regulation of RAB3GAP1, RAB3GAP2 and SNAP25 affects secretion of Myocilin (MYOC), a protein associated with juvenile onset glaucoma and steroid-induced glaucoma. The present work reveals that FOXC1 is an important regulator of exocytosis and establishes a new link between FOXC1 and MYOC-associated glaucoma.

In vitro gentamicin exposure alters caveolae protein profile in cochlear spiral ligament pericytes

“…Numerous findings propose that caveolae transcytose low density lipoporteins (LDLs), albumin and insulin [106]. Rab8a has been associated with basolateral-to-apical transcytosis in polarized Madin-Darby Canine Kidney Cells (MDCK) cells [107], while Rab3gap2 had been shown to form with the protein Rab3gap1 a complex acting in exocytosis and endocytosis, and extracellular trafficking [108].…”

FOXC1 Regulates Expression of Prostaglandin Receptors Leading to an Attenuated Response to Latanoprost

“…12 Briefly, confluent HeLa cells were fixed and their nuclei were lysed. Chromatin was incubated with 5 lg of either rabbit anti-IgG (CALTAG Laboratories, Buckingham, UK), a negative control; rabbit anti-H3AcK9 (Enzo Life Sciences, Farmingdale, NY, USA), a positive control for transcriptionally active chromatin; or goat anti-FOXC1 (OriGene, Rockville, MD, USA).…”

“…A luciferase-containing vector (pGL3) was modified with a thymidine kinase (TK) basal promoter sequence as previously described 12 (herein referred to as pGL3-TK) to test for the presence of an enhancer element. The fragment from PTGER3 identified previously 10 was cloned downstream of the TK promoter using BglII and SacI restriction sites for unidirectional insertion to create pGL3-TL-PTGER3.…”

“…12 In brief, TM1 and HeLa cells were transfected with an siRNA for FOXC1 or a control scrambled siRNA. Cell lysates were harvested and electrophoretically separated on a polyacrylamide gel and then transferred to a nitrocellulose membrane.…”

“…The DD Ct method was used to calculate the relative quantities of RNA in comparison to the siRNA scrambled control as previously described. 12 An unpaired, two-tailed Student's t-test was used to calculate statistical significance.…”