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Nerve Growth Factor‐Induced Changes in the Structure of Sulfated Proteoglycans in PC 12 Pheochromocytoma Cells
Author(s) -
KatohSemba Ritsuko,
Oohira Atsuhiko,
Kashiwamata Shigeo
Publication year - 1992
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1111/j.1471-4159.1992.tb08902.x
Subject(s) - nerve growth factor , molecular mass , sulfation , cell culture , chondroitin sulfate , chemistry , microbiology and biotechnology , biochemistry , glycosaminoglycan , biology , receptor , enzyme , genetics
Structural changes in proteoglycans (PGs) were examined during the neuritogenesis of PC12 cells induced by nerve growth factor (NGF). (1) A heparan sulfate (HS) PG and a chondroitin sulfate (CS) PG were synthesized by PC 12 cells, irrespective of the presence of NGF or the duration of culture. PGs released from PC12 cells into the culture medium were mostly CSPGs. (2) In the absence of NGF, the apparent molecular mass of HSPG prepared from PC 12 cells after 3 days of culture was in the range of 90–190 kDa for the intact form ( K av = 0.38 on Sepharose CL‐6B), 12 kDa for HS, and 61 kDa for the core protein. In the presence of NGF, these values were 90–190 kDa, 10 kDa, and 51 kDa and 61 kDa, respectively. The intact forms of cell‐associated CSPG had apparent molecular mass ranges of 120–150 kDa and 120–190 kDa ( K av = 0.38 and 0.34), with CSs of 15 kDa and 20 kDa in the presence and absence of NGF, respectively. The apparent molecular mass of the core protein of cell‐associated CSPG was 92 kDa, irrespective of the presence of NGF. The molecular sizes of cell‐associated PGs and their glycosaminoglycans remained unchanged during culture. (3) CSPGs released by PC12 cells into the culture medium were separated into two peaks (I and II) by column chromatography on DEAE‐cellulose. The peak II fraction prepared from the medium with NGF after 3 days of culture consisted of CSPG with K av = 0.22 on Sephacryl S‐300 [40–84 kDa by sodium dodecyl sulfate‐polyacrylamide gel electrophoresis (SDS‐PAGE)]. The CS and core protein of this CSPG had apparent molecular masses of 26 kDa and 15–17 kDa, respectively. The peak I fraction was separated further into three fractions of different molecular sizes on Sephadex G‐200 (I‐1,I‐2,I‐3). In the presence of NGF, peak I‐1 prepared after 3 days of culture consisted of mostly CSPG (130–170 kDa by SDS‐PAGE; K av = 0.13 on Sephacryl S‐300) with CS of 32 kDa and a core protein of 105 kDa. Peaks 1–2 and 1–3 contained, respectively, CSPGs of 38–74 kDa and 31–56 kDa for their intact forms, with CSs of 25 kDa and 15 kDa. The apparent molecular masses of the core proteins of their CSPGs were 15–17 kDa. These CSPGs had similar molecular sizes, irrespective of the presence of NGF or the duration of culture. (4) These results indicate that structural changes in PGs during the neuritogenesis in PC12 cells induced by NGF occurred mostly in cell‐associated PGs and preceded elongation of neurites.