ABSTRACT
Testican is a highly conserved, differentially expressed gene product of unknown function. Since testican is expressed by human endothelial cells and includes a signal sequence, it was our hypothesis that testican protein would be present in blood. We have developed chicken antibodies specific for testican sequence near the N-terminal and identified a 130-kDa form of testican in human plasma. This is much larger than the calculated molecular weight of the encoded polypeptide, suggesting glycosylation of this plasma protein, and large forms of recombinant testican produced in culture were found to include chondroitin sulfate. The 130-kDa form of testican is unstable in plasma. It is converted to smaller stable forms by separable plasma factors that can be blocked by certain serine protease inhibitors. Testican size conversion may be important in its functional activation or decay. One testican domain has strong homology to thyropin-type cysteine protease-inhibitors. Thus, testican may have a function related to protease inhibition in the blood.
Subject(s)
Proteoglycans/blood , Amino Acid Sequence , Animals , Antibodies , Chickens , Electrophoresis, Polyacrylamide Gel , Glycosaminoglycans/chemistry , Humans , Immunoblotting , Models, Molecular , Molecular Sequence Data , Molecular Weight , Open Reading Frames , Peptide Fragments/chemistry , Peptide Fragments/immunology , Protein Structure, Secondary , Proteoglycans/chemistry , Proteoglycans/genetics , Recombinant Proteins/chemistry , Recombinant Proteins/immunology , Sequence Alignment , Sequence Homology, Amino Acid , Testicular Hormones/blood , Thyroglobulin/chemistryABSTRACT
Testican is a putative extracellular heparan/ chondroitin sulfate proteoglycan of unknown function that is expressed in a variety of human tissues at widely different levels but is most abundant in the brain. In mice, testican mRNA has been detected only in brain and it is therefore likely to have an important function in the central nervous system. RNA blot analysis reveals the relative intensity of testican in various regions of the human brain. Levels of testican message are most pronounced in the thalamus, hippocampus, occipital lobe, nucleus accumbens, temporal lobe, and caudate nucleus, with somewhat lower levels in the cerebral cortex, medulla oblongata, frontal lobe, amygdala, putamen, spinal cord, substantia nigra, and cerebellum. In situ hybridization reveals the cellular distribution of the mRNA within these areas to be highest in neurons and in choroid plexus epithelium, and moderately lower in ependymal cells lining the ventricles and in vascular endothelial cells. Testican mRNA is not detected in oligodendrocytes or in most astrocytes. However, astrocytes in regions of reactive gliosis do express testican mRNA. These findings, along with a cysteine-rich pattern similarity to neurocan, brevican, versican, and other proteoglycans found in brain, suggest that testican may be a part of the specialized extracellular matrix of the brain.
Subject(s)
Brain/metabolism , Proteoglycans/metabolism , Basal Ganglia/metabolism , Brain/anatomy & histology , Brain Stem/metabolism , Cerebellum/metabolism , Cerebral Cortex/metabolism , Choroid Plexus/metabolism , Extracellular Matrix Proteins/genetics , Extracellular Matrix Proteins/metabolism , Extracellular Matrix Proteins/physiology , Hippocampus/metabolism , Humans , Hypothalamus/metabolism , In Situ Hybridization , Pituitary Gland/metabolism , Proteoglycans/genetics , Proteoglycans/physiology , RNA, Messenger/biosynthesis , Transcription, GeneticABSTRACT
By screening random cDNAs from a continuous vascular endothelial cell line, EA.hy926, we identified a 5 kb mRNA that is expressed at high levels by this human cell line and by an early passage umbilical vein endothelial cell line. It is detected at lower levels in certain stromal cell lines, but it is not detected in most other cell lines tested, indicating that it represents a differentially expressed function rather than a ubiquitous or housekeeping function. This mRNA was readily detected in samples derived from most human organs as might be expected for a gene expressed in the vascular wall. Sequencing of the 5 kb mRNA reveals its identity with 3.5 kb of previously published testis-derived cDNA sequence called testican (Alliel et al., 1993). Differential expression of this gene by endothelial cells contributes a new perspective on the potential function of testican.
