Heterogeneous metabolism and subcellular localization of a potentially leukemogenic membrane glycoprotein encoded by Friend erythroleukemia virus. Isolation of viral and cellular processing mutants

M. Ruta, S. Clarke, B. Boswell, David Kabat

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Abstract

We have analyzed the metabolism of gp55, a membrane glycoprotein with an apparent M(r) of 55,000 which is encoded by Friend spleen focus-forming virus (SFFV) and which has recently been implicated in causation of viral erythroleukemia. A small proportion of gp55 (3 to 5%) which has a larger size than the major component is located on the outer surface of plasma membranes and is called gp55(P), whereas the remainder is irreversibly situated in membranous intracellular organelle(s). Although both of these glycoprotein components contain asparagine-linked oligosaccharides, intracellular gp55 contains small high mannose oligosaccharides of the type formed in the endoplasmic reticulum, whereas gp55(P) contains larger complex oligosaccharides with galactose, fucose, and sialic acid, indicative of transit through the Golgi apparatus. To analyze the causes for its heterogeneous metabolism, we studied viral and cellular mutants with defects in gp55 processing. An SFFV mutant with an abnormal gp55 structural gene was obtained by an extensive survey of independently cloned SFFVs. Cellular mutants were selected by immune killing of SFFV-infected normal rat kidney fibroblasts with an antiserrum that reacts with gp55. The resulting immunoresistant cell lines lack gp55(P). Furthermore, the intracellular gp55 in these mutants contains immature forms of high mannose oligosaccharides which have glucose and are larger and more homogeneous than the processed or partially degraded forms which predominate in wild type cells. These results support the idea that gp55 is processed along a single pathway from the rough endoplasmic reticulum via the Golgi apparatus to the plasma membrane. However, the processing of most gp55 molecules ceases in a stochastic fashion en route and the rate of this cessation is increased in the immunoresistant mutants, thereby blocking gp55 processing at an early stage and reducing gp55(P) formation. We propose that heterogeneous cessation of gp55 processing occurs by spontaneous, random denaturation and is caused by a folding instability of the polypeptide chain. This unusual metabolism could possibly be involved in the leukemogenic mechanism.

Original languageEnglish (US)
Pages (from-to)126-132
Number of pages7
JournalJournal of Biological Chemistry
Volume257
Issue number1
StatePublished - 1982

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Spleen Focus-Forming Viruses
Friend murine leukemia virus
Leukemia, Erythroblastic, Acute
Membrane Glycoproteins
Oligosaccharides
Viruses
Metabolism
Golgi Apparatus
Cell membranes
Mannose
Processing
Cell Membrane
Denaturation
Fucose
Rough Endoplasmic Reticulum
Asparagine
N-Acetylneuraminic Acid
Fibroblasts
Galactose
Endoplasmic Reticulum

ASJC Scopus subject areas

  • Biochemistry

Cite this

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title = "Heterogeneous metabolism and subcellular localization of a potentially leukemogenic membrane glycoprotein encoded by Friend erythroleukemia virus. Isolation of viral and cellular processing mutants",
abstract = "We have analyzed the metabolism of gp55, a membrane glycoprotein with an apparent M(r) of 55,000 which is encoded by Friend spleen focus-forming virus (SFFV) and which has recently been implicated in causation of viral erythroleukemia. A small proportion of gp55 (3 to 5{\%}) which has a larger size than the major component is located on the outer surface of plasma membranes and is called gp55(P), whereas the remainder is irreversibly situated in membranous intracellular organelle(s). Although both of these glycoprotein components contain asparagine-linked oligosaccharides, intracellular gp55 contains small high mannose oligosaccharides of the type formed in the endoplasmic reticulum, whereas gp55(P) contains larger complex oligosaccharides with galactose, fucose, and sialic acid, indicative of transit through the Golgi apparatus. To analyze the causes for its heterogeneous metabolism, we studied viral and cellular mutants with defects in gp55 processing. An SFFV mutant with an abnormal gp55 structural gene was obtained by an extensive survey of independently cloned SFFVs. Cellular mutants were selected by immune killing of SFFV-infected normal rat kidney fibroblasts with an antiserrum that reacts with gp55. The resulting immunoresistant cell lines lack gp55(P). Furthermore, the intracellular gp55 in these mutants contains immature forms of high mannose oligosaccharides which have glucose and are larger and more homogeneous than the processed or partially degraded forms which predominate in wild type cells. These results support the idea that gp55 is processed along a single pathway from the rough endoplasmic reticulum via the Golgi apparatus to the plasma membrane. However, the processing of most gp55 molecules ceases in a stochastic fashion en route and the rate of this cessation is increased in the immunoresistant mutants, thereby blocking gp55 processing at an early stage and reducing gp55(P) formation. We propose that heterogeneous cessation of gp55 processing occurs by spontaneous, random denaturation and is caused by a folding instability of the polypeptide chain. This unusual metabolism could possibly be involved in the leukemogenic mechanism.",
author = "M. Ruta and S. Clarke and B. Boswell and David Kabat",
year = "1982",
language = "English (US)",
volume = "257",
pages = "126--132",
journal = "Journal of Biological Chemistry",
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T1 - Heterogeneous metabolism and subcellular localization of a potentially leukemogenic membrane glycoprotein encoded by Friend erythroleukemia virus. Isolation of viral and cellular processing mutants

AU - Ruta, M.

AU - Clarke, S.

AU - Boswell, B.

AU - Kabat, David

PY - 1982

Y1 - 1982

N2 - We have analyzed the metabolism of gp55, a membrane glycoprotein with an apparent M(r) of 55,000 which is encoded by Friend spleen focus-forming virus (SFFV) and which has recently been implicated in causation of viral erythroleukemia. A small proportion of gp55 (3 to 5%) which has a larger size than the major component is located on the outer surface of plasma membranes and is called gp55(P), whereas the remainder is irreversibly situated in membranous intracellular organelle(s). Although both of these glycoprotein components contain asparagine-linked oligosaccharides, intracellular gp55 contains small high mannose oligosaccharides of the type formed in the endoplasmic reticulum, whereas gp55(P) contains larger complex oligosaccharides with galactose, fucose, and sialic acid, indicative of transit through the Golgi apparatus. To analyze the causes for its heterogeneous metabolism, we studied viral and cellular mutants with defects in gp55 processing. An SFFV mutant with an abnormal gp55 structural gene was obtained by an extensive survey of independently cloned SFFVs. Cellular mutants were selected by immune killing of SFFV-infected normal rat kidney fibroblasts with an antiserrum that reacts with gp55. The resulting immunoresistant cell lines lack gp55(P). Furthermore, the intracellular gp55 in these mutants contains immature forms of high mannose oligosaccharides which have glucose and are larger and more homogeneous than the processed or partially degraded forms which predominate in wild type cells. These results support the idea that gp55 is processed along a single pathway from the rough endoplasmic reticulum via the Golgi apparatus to the plasma membrane. However, the processing of most gp55 molecules ceases in a stochastic fashion en route and the rate of this cessation is increased in the immunoresistant mutants, thereby blocking gp55 processing at an early stage and reducing gp55(P) formation. We propose that heterogeneous cessation of gp55 processing occurs by spontaneous, random denaturation and is caused by a folding instability of the polypeptide chain. This unusual metabolism could possibly be involved in the leukemogenic mechanism.

AB - We have analyzed the metabolism of gp55, a membrane glycoprotein with an apparent M(r) of 55,000 which is encoded by Friend spleen focus-forming virus (SFFV) and which has recently been implicated in causation of viral erythroleukemia. A small proportion of gp55 (3 to 5%) which has a larger size than the major component is located on the outer surface of plasma membranes and is called gp55(P), whereas the remainder is irreversibly situated in membranous intracellular organelle(s). Although both of these glycoprotein components contain asparagine-linked oligosaccharides, intracellular gp55 contains small high mannose oligosaccharides of the type formed in the endoplasmic reticulum, whereas gp55(P) contains larger complex oligosaccharides with galactose, fucose, and sialic acid, indicative of transit through the Golgi apparatus. To analyze the causes for its heterogeneous metabolism, we studied viral and cellular mutants with defects in gp55 processing. An SFFV mutant with an abnormal gp55 structural gene was obtained by an extensive survey of independently cloned SFFVs. Cellular mutants were selected by immune killing of SFFV-infected normal rat kidney fibroblasts with an antiserrum that reacts with gp55. The resulting immunoresistant cell lines lack gp55(P). Furthermore, the intracellular gp55 in these mutants contains immature forms of high mannose oligosaccharides which have glucose and are larger and more homogeneous than the processed or partially degraded forms which predominate in wild type cells. These results support the idea that gp55 is processed along a single pathway from the rough endoplasmic reticulum via the Golgi apparatus to the plasma membrane. However, the processing of most gp55 molecules ceases in a stochastic fashion en route and the rate of this cessation is increased in the immunoresistant mutants, thereby blocking gp55 processing at an early stage and reducing gp55(P) formation. We propose that heterogeneous cessation of gp55 processing occurs by spontaneous, random denaturation and is caused by a folding instability of the polypeptide chain. This unusual metabolism could possibly be involved in the leukemogenic mechanism.

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