Liang Y, Häring M, Roughley P J, Margolis R K, Margolis R U
Department of Pharmacology, New York University Medical Center, New York 10016, USA.
J Cell Biol. 1997 Nov 17;139(4):851-64. doi: 10.1083/jcb.139.4.851.
We have investigated the expression patterns and subcellular localization in nervous tissue of glypican, a major glycosylphosphatidylinositol-anchored heparan sulfate proteoglycan that is predominantly synthesized by neurons, and of biglycan, a small, leucine-rich chondroitin sulfate proteoglycan. By laser scanning confocal microscopy of rat central nervous tissue and C6 glioma cells, we found that a significant portion of the glypican and biglycan immunoreactivity colocalized with nuclear staining by propidium iodide and was also seen in isolated nuclei. In certain regions, staining was selective, insofar as glypican and biglycan immunoreactivity in the nucleus was seen predominantly in a subpopulation of large spinal cord neurons. The amino acid sequences of both proteoglycans contain potential nuclear localization signals, and these were demonstrated to be functional based on their ability to target beta-galactosidase fusion proteins to the nuclei of transfected 293 cells. Nuclear localization of glypican beta-galactosidase or Fc fusion proteins in transfected 293 cells and C6 glioma cells was greatly reduced or abolished after mutation of the basic amino acids or deletion of the sequence containing the nuclear localization signal, and no nuclear staining was seen in the case of heparan sulfate and chondroitin sulfate proteoglycans that do not possess a nuclear localization signal, such as syndecan-3 or decorin (which is closely related in structure to biglycan). Transfection of COS-1 cells with an epitope-tagged glypican cDNA demonstrated transport of the full-length proteoglycan to the nucleus, and there are also dynamic changes in the pattern of glypican immunoreactivity in the nucleus of C6 cells both during cell division and correlated with different phases of the cell cycle. Our data therefore suggest that in certain cells and central nervous system regions, glypican and biglycan may be involved in the regulation of cell division and survival by directly participating in nuclear processes.
我们研究了磷脂酰肌醇蛋白聚糖（一种主要由神经元合成的糖基磷脂酰肌醇锚定硫酸乙酰肝素蛋白聚糖）和双糖链蛋白聚糖（一种富含亮氨酸的小分子硫酸软骨素蛋白聚糖）在神经组织中的表达模式和亚细胞定位。通过对大鼠中枢神经组织和C6胶质瘤细胞进行激光扫描共聚焦显微镜观察，我们发现，磷脂酰肌醇蛋白聚糖和双糖链蛋白聚糖的显著部分免疫反应性与碘化丙啶的核染色共定位，并且在分离的细胞核中也可见。在某些区域，染色具有选择性，因为细胞核中的磷脂酰肌醇蛋白聚糖和双糖链蛋白聚糖免疫反应性主要见于大脊髓神经元的一个亚群中。两种蛋白聚糖的氨基酸序列都包含潜在的核定位信号，基于它们将β-半乳糖苷酶融合蛋白靶向转染的293细胞核的能力，这些信号被证明是有功能的。在碱性氨基酸发生突变或包含核定位信号的序列被缺失后，转染的293细胞和C6胶质瘤细胞中磷脂酰肌醇蛋白聚糖β-半乳糖苷酶或Fc融合蛋白的核定位大大减少或消失，并且对于不具有核定位信号的硫酸乙酰肝素和硫酸软骨素蛋白聚糖（如多配体蛋白聚糖-3或核心蛋白聚糖（其结构与双糖链蛋白聚糖密切相关）），未观察到核染色。用表位标记的磷脂酰肌醇蛋白聚糖cDNA转染COS-1细胞证明全长蛋白聚糖向细胞核的转运，并且在细胞分裂期间以及与细胞周期的不同阶段相关时，C6细胞核中磷脂酰肌醇蛋白聚糖免疫反应性模式也存在动态变化。因此，我们的数据表明，在某些细胞和中枢神经系统区域，磷脂酰肌醇蛋白聚糖和双糖链蛋白聚糖可能通过直接参与核过程而参与细胞分裂和存活的调节。
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