Molecular Cloning and Characterization of Chick SPACRCAN*

MY-174, a monoclonal antibody that reacts with specific sialylated O-linked glycoconjugates of chick SPACR (sialoprotein associated with cones and rods), also recognizes another molecule of 300 kDa. Here, we verified that this 300-kDa molecule is chick SPACRCAN (sialoproteoglycan associated with cones and rods), another member of a novel interphotoreceptor matrix molecule family. Screening for chick SPACRCAN was carried out by plaque hybridization using a probe for chick SPACR. Specific polyclonal antibodies raised against chick SPACRCAN were used for the following experiments. To determine whether the 300-kDa molecule detected by MY-174 was identical to 300-kDa chick SPACRCAN, the migrations of these bands were examined after various glycosidase digestions. Furthermore, the expression levels were measured during retinal development and compared with those of chick SPACR. The results demonstrated that the 300-kDa molecule recognized by MY-174 was chick SPACRCAN, and we further identified it as a proteoglycan with chondroitin sulfate chains. SPACRCAN had heavily sialylated N- and O-linked glycoconjugates, and its MY-174 antigenicity was abolished by O-glycanase treatment after neuraminidase treatment, as observed for chick SPACR. During retinal development, the mRNA and core protein expression levels, MY-174 antigenicity, and hyaluronan binding ability of SPACRCAN peaked around embryonic day 17 and then gradually decreased, whereas the corresponding expression levels of SPACR simply increased, but not its hyaluronan binding ability. The MY-174 reactivity of SPACRCAN in the adult retina was decreased compared with that in the newborn retina, whereas that of SPACR was increased. The decreased hyaluronan binding of SPACR was induced by an inhibitory effect of the excess of sialic acids in the adult stage. Thus, with similar core protein structures and specific sialylated glycoconjugates but distinct chondroitin sulfate chains, SPACRCAN and SPACR may have separate roles in the retina due to their differing expression profiles during development.

[1]  G. Prestwich,et al.  Spacrcan Binding to Hyaluronan and Other Glycosaminoglycans , 2004, Journal of Biological Chemistry.

[2]  M. Yamagata,et al.  Tissue variation of two large chondroitin sulfate proteoglycans (PG-M/versican and PG-H/aggrecan) in chick embryos , 1993, Anatomy and Embryology.

[3]  Qiuyun Chen,et al.  SPACRCAN in the interphotoreceptor matrix of the mouse retina: molecular, developmental and promoter analysis. , 2003, Experimental eye research.

[4]  Yasuhiko Suzuki,et al.  Molecular Cloning and Characterization of Chick Sialoprotein Associated with Cones and Rods, a Developmentally Regulated Glycoprotein of Interphotoreceptor Matrix* , 2002, The Journal of Biological Chemistry.

[5]  W. Young,et al.  SPACRCAN in the developing retina and pineal gland of the rat: Spatial and temporal pattern of gene expression and protein synthesis , 2001, The Journal of comparative neurology.

[6]  M. Miyagi,et al.  Interphotoreceptor matrix in the fovea and peripheral retina of the primate Macaca mulatta: distribution and glycoforms of SPACR and SPACRCAN. , 2001, Experimental eye research.

[7]  A. Oohira,et al.  Neuroglycan C, a neural tissue-specific transmembrane chondroitin sulfate proteoglycan, in retinal neural network formation. , 2000, Investigative ophthalmology & visual science.

[8]  I. Rodriguez,et al.  SPACR in the interphotoreceptor matrix of the mouse retina: molecular, biochemical and immunohistochemical characterization. , 2000, Experimental eye research.

[9]  W. Young,et al.  SPACRCAN, a Novel Human Interphotoreceptor Matrix Hyaluronan-binding Proteoglycan Synthesized by Photoreceptors and Pinealocytes* , 2000, The Journal of Biological Chemistry.

[10]  R. Midura,et al.  Chondroitin sulfate proteoglycan core proteins in the interphotoreceptor matrix: a comparative study using biochemical and immunohistochemical analysis. , 1999, Experimental eye research.

[11]  R. Midura,et al.  SPACR, a Novel Interphotoreceptor Matrix Glycoprotein in Human Retina That Interacts with Hyaluronan* , 1998, The Journal of Biological Chemistry.

[12]  J. Hollyfield,et al.  Characterization of SPACR, a sialoprotein associated with cones and rods present in the interphotoreceptor matrix of the human retina: immunological and lectin binding analysis. , 1998, Glycobiology.

[13]  M. Tammi,et al.  Hyaluronan in the interphotoreceptor matrix of the eye: species differences in content, distribution, ligand binding and degradation. , 1998, Experimental eye research.

[14]  J. Hollyfield,et al.  Hyaluronan localization in tissues of the mouse posterior eye wall: absence in the interphotoreceptor matrix. , 1997, Experimental eye research.

[15]  K. Kimata,et al.  Transient Expression of PG‐M/Versican, a Large Chondroitin Sulfate Proteoglycan in Developing Chicken Retina , 1997, Journal of neurochemistry.

[16]  M. Brownstein,et al.  Sequence analysis of PG10.2, a gene expressed in the pineal gland and the outer nuclear layer of the retina. , 1996, Brain research. Molecular brain research.

[17]  E. Turley,et al.  The characterization of a human RHAMM cDNA: conservation of the hyaluronan-binding domains. , 1996, Gene.

[18]  P. Bork,et al.  The SEA module: A new extracellular domain associated with O‐glycosylation , 1995, Protein science : a publication of the Protein Society.

[19]  W. Halfter,et al.  Two chondroitin sulfate proteoglycans differentially expressed in the developing chick visual system. , 1995, Developmental biology.

[20]  K. Ito,et al.  Expression of PG-M(V3), an Alternatively Spliced Form of PG-M without a Chondroitin Sulfate Attachment Region in Mouse and Human Tissues (*) , 1995, The Journal of Biological Chemistry.

[21]  W. Knudson,et al.  Hyaluronan‐binding proteins in development, tissue homeostasis, and disease , 1993, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[22]  J. Silver,et al.  Chondroitin sulfate as a regulator of neuronal patterning in the retina. , 1992, Science.

[23]  V. Hamburger,et al.  A series of normal stages in the development of the chick embryo. 1951. , 2012, Developmental dynamics : an official publication of the American Association of Anatomists.

[24]  A. Adler,et al.  Proteins and glycoproteins of the bovine interphotoreceptor matrix: composition and fractionation. , 1982, Experimental eye research.

[25]  A. Adler,et al.  Proteins of the bovine interphotoreceptor matrix: tissues of origin. , 1981, Experimental eye research.

[26]  R. Hill,et al.  Systematic purification of five glycosidases from Streptococcus (Diplococcus) pneumoniae. , 1977, The Journal of biological chemistry.