Myocilin Mutations Causing Glaucoma Inhibit the Intracellular Endoproteolytic Cleavage of Myocilin between Amino Acids Arg226 and Ile227

Aroca-Aguilar, José-Daniel; Sánchez-Sánchez, Francisco; Ghosh, Sikha; Coca‐Prados, Miguel; Escribano, Julio

doi:10.1074/jbc.m501340200

Cited by 67 publications

(118 citation statements)

References 36 publications

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“…1A). Published data suggest that the C-terminal fragment of myocilin is secreted (3,23,66), while data concerning secretion of the N-terminal fragment are controversial (23,66). To test whether N-and C-terminal fragments of myocilin are (52) and anti-FLAG antibodies, respectively (Fig.…”

Section: Resultsmentioning

confidence: 99%

Myocilin Is a Modulator of Wnt Signaling

Kwon

Lee

et al. 2009

Molecular and Cellular Biology

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It is well documented that mutations in the MYOCILINGlaucoma is a leading cause of blindness in the world. Primary open-angle glaucoma is the most common form of glaucoma. It will affect more than 60 million and blind about 4.5 million people worldwide by the year 2010 (62). It is now well established that a genetic component may contribute to glaucoma, and several glaucoma-associated genes have been identified. The first-identified and the most-studied gene is MYOCILIN (MYOC), which is highly expressed in and secreted by the trabecular meshwork (1,19,72,75,77,78), one of the key components of the eye aqueous humor outflow system. Dominant mutations in MYOC may lead to juvenile openangle glaucoma and are found in 3 to 4% of patients with primary open-angle glaucoma (18,19). The encoded protein, myocilin, belongs to a family of glycosylated proteins containing a C-terminal olfactomedin domain (57,90,91). Olfactomedin domain was originally identified in a glycoprotein isolated from the olfactory epithelium of frogs (71) and subsequently was found in a group of proteins forming a family of olfactomedin domaincontaining proteins. The family of olfactomedin domain-containing proteins includes both secreted and membrane-bound proteins that each exhibit a characteristic distribution in different tissues (4,10,27,30,44,51,58,78,79). Most of the glaucomacausing mutations in myocilin are located in the olfactomedin domain (1,2,18,19,24,72).Besides the trabecular meshwork and sclera (1, 77), high levels of MYOC were observed in the ciliary body (1, 13, 39), iris (89), retinal pigment epithelium/choroid (78, 88), and optic nerve (74). Low levels of MYOC expression were detected in several nonocular tissues (75). Myocilin, being a secreted protein, was also found in the aqueous humor, an intraocular fluid responsible for the supply of nutrients and for removal of metabolites from the avascular tissues of the eye anterior segment (63, 65). Some mutations in the MYOC gene lead to the inhibition of mutated myocilin secretion. Secretion of wildtype myocilin also can be reduced or blocked in the presence of mutated myocilin (22,36,52). It has been suggested that the intracellular accumulation of myocilin aggregates is deleterious to the trabecular meshwork cells, resulting in the deterioration of their function and subsequent elevation of intraocular pressure (IOP) (38,49).Although the MYOC gene has been studied for more than 10 years, the functions of myocilin protein are still not well understood (19,75). Biochemical data indicate that myocilin may interact with several intracellular and extracellular matrix proteins (16,37,48,61,73,78), although the biological significance of such interactions is not clear. The absence of openangle glaucoma in an elderly woman homozygous for the Arg46Stop mutation (45), as well as the absence of glaucoma in people hemizygous for MYOC (87), suggests that the loss of functional myocilin is not critical for the development of glaucoma or for normal eye functioning. These observations are supporte...

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Section: Resultsmentioning

confidence: 99%

Myocilin Is a Modulator of Wnt Signaling

Kwon

Lee

et al. 2009

Molecular and Cellular Biology

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“…heart and skeletal muscle) and in ocular tissues, such as iris, ciliary body (CB), and trabecular meshwork (TM) (3)(4)(5)(6)(7)(8). The protein is also present in aqueous humor (9,10). Interestingly, most MYOC mutations reported to date in sporadic cases of primary open angle glaucoma are heterozygous and are confined to exon 3 (2,5,6,(11)(12)(13).…”

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confidence: 99%

“…Interestingly, most MYOC mutations reported to date in sporadic cases of primary open angle glaucoma are heterozygous and are confined to exon 3 (2,5,6,(11)(12)(13). Recently, we showed that myocilin undergoes an intracellular endoproteolytic cleavage in the endoplasmic reticulum (ER) between amino acids Arg 226 and Ile 227 located in the putative linker domain (10). This processing predicts the production of two fragments corresponding to the N-and C-terminal domains.…”

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confidence: 99%

“…Several extracellular proteins have been identified to interact with myocilin, including fibronectin (through the HepII domain) (14), the C-terminal regions of fibrilin-1 (15), and hevin, an extracellular matrix protein member of the BM-40/SPARC/osteonectin family (16). Although the functional meaning of the proteolytic processing of myocilin is currently unknown, it has been suggested that controlled changes in the proportion of processed myocilin might contribute to regulate the normal TM structure by differential intermolecular interactions with other extracellular matrix molecules, therefore contributing to modulate aqueous humor outflow and IOP (10).…”

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Characterization of the Intracellular Proteolytic Cleavage of Myocilin and Identification of Calpain II as a Myocilin-processing Protease

Sánchez-Sánchez

Martínez-Redondo

Aroca-Aguilar

et al. 2007

Journal of Biological Chemistry

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MYOC, a gene involved in different types of glaucoma, encodes myocilin, a secreted glycoprotein of unknown function, consisting of an N-terminal leucine-zipper-like domain, a central linker region, and a C-terminal olfactomedin-like domain. Recently, we have shown that myocilin undergoes an intracellular endoproteolytic processing. We show herein that the proteolytic cleavage in the linker region splits the two terminal domains. The C-terminal domain is secreted to the culture medium, whereas the N-terminal domain mainly remains intracellularly retained. In transiently transfected 293T cells, the cleavage was prevented by calpain inhibitors, such as calpeptin, calpain inhibitor IV, and calpastatin. Since calpains are calciumactivated proteases, we analyzed how changes in either intra-or extracellular calcium affected the cleavage of myocilin. Intracellular ionomycin-induced calcium uptake enhanced myocilin cleavage, whereas chelation of extracellular calcium by EGTA inhibited the proteolytic processing. Calpains I and II cleaved myocilin in vitro. However, in cells in culture, only RNA interference knockdown of calpain II reduced myocilin processing. Subcellular fractionation and digestion of the obtained fractions with proteinase K showed that full-length myocilin resides in the lumen of the endoplasmic reticulum together with a subpopulation of calpain II. These data revealed that calpain II is responsible for the intracellular processing of myocilin in the lumen of the endoplasmic reticulum. We propose that this cleavage might regulate extracellular interactions of myocilin, contributing to the control of intraocular pressure.

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“…There is evidence for physiological processing of other molecules that contain olfactomedin domains. For example, myocilin was recently shown to be cleaved endoproteolytically in the linker region between the N-terminal and olfactomedin domains (43). However, details concerning the processing of other olfactomedin domain-containing proteins and what differences or similarities that such processing may have to the processing of gliomedin described here remain to be characterized.…”

Section: Discussionmentioning

confidence: 89%

Cleavage and Oligomerization of Gliomedin, a Transmembrane Collagen Required for Node of Ranvier Formation

Maertens

Hopkins

Franzke

et al. 2007

Journal of Biological Chemistry

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Gliomedin, which has been implicated as a major player in genesis of the nodes of Ranvier, contains two collagenous domains and an olfactomedin-like domain and belongs to the group of type II transmembrane collagens that includes collagens XIII and XVII and ectodysplasin A. One characteristic of this protein family is that constituent proteins can exist in both transmembrane and soluble forms. Recently, gliomedin expressed at the tips of Schwann cell microvilli was found to bind axonal adhesion molecules neurofascin and NrCAM in interactions essential for Na ؉ -channel clustering at the nodes of Ranvier in myelinating peripheral nerves. Interestingly, exogenously added olfactomedin domain was found to have the same effect as intact gliomedin. Here we analyze the tissue form of gliomedin and demonstrate that the molecule not only exists as full-length gliomedin but also as a soluble form shed from the cell surface in a furin-dependent manner. In addition, gliomedin can be further proteolytically processed by bone morphogenetic protein 1/Tolloid-like enzymes, resulting in release of the olfactomedin domain from the collagen domains. Interestingly, the later cleavage induces formation of higher order, insoluble molecular aggregates that may play important roles in Na ؉ -channel clustering.Collagens and other proteins with collagenous domains constitute the most abundant components of the extracellular matrix. They also play essential roles in development and provision of structural integrity to most tissues and organs and can be divided into nine subgroups (1). Type II-oriented transmembrane collagens are one such subgroup. These play roles in cellmatrix interactions both as integral membrane proteins and as shed forms cleaved from cell surfaces by limited proteolysis (2). Transmembrane collagens XIII (3) and XXV (4) and transmembrane protein ectodysplasin A, which is not formally a collagen but which contains collagenous domains (5), are shed by cleavage at the juxtamembrane consensus sequence (K/R)X n (K/R) (n ϭ 0, 2, 4, 6). Such cleavage is by furin-like proprotein convertases of the S8 family of Kex/subtilisin-related serine endopeptidases (6 -8). In contrast, collagen XVII is not directly cleaved by furin-like convertases. Rather, furin-like convertases activate ADAM (a disintegrin and metalloproteinase) family proteinases that cleave collagen XVII (2, 9).The designation colmedin (collagen repeat plus olfactomedin domain) describes a new type of transmembrane collagen (10) containing an ϳ260-amino acid olfactomedin-like domain at the C terminus. The prototype, olfactomedin, was first identified in the extracellular mucus matrix of the olfactory neuroepithelium of Rana catesbeiana (11,12). With the exception of the sea urchin protein amassin (13), all subsequently characterized olfactomedin-like domain-containing proteins are expressed in neural tissues and seem involved in neural function. Noelin-1 participates in generation of neural crest cells (14, 15), photomedins are expressed in retina (16), tiarin is re...

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Myocilin Mutations Causing Glaucoma Inhibit the Intracellular Endoproteolytic Cleavage of Myocilin between Amino Acids Arg226 and Ile227

Cited by 67 publications

References 36 publications

Myocilin Is a Modulator of Wnt Signaling

Myocilin Is a Modulator of Wnt Signaling

Characterization of the Intracellular Proteolytic Cleavage of Myocilin and Identification of Calpain II as a Myocilin-processing Protease

Cleavage and Oligomerization of Gliomedin, a Transmembrane Collagen Required for Node of Ranvier Formation

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