Synthesis, processing, and secretion of apolipoprotein B by the chick liver cell.

Siuta-Mangano, P; Howard, Sarah; Lennarz, William J.; Lane, M. Daniel

doi:10.1016/s0021-9258(18)34720-3

Cited by 50 publications

(6 citation statements)

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“…Abbreviations used in this paper: apoAI, apolipoprotein AI; apoB, apolipoprotein B; HDL, high density lipoprotein; VLDL, very low density lipoprotein. and Vance, 1988), and carbohydrate (Siuta-Mangano et al, 1982) moieties of the lipoproteins have been determined. There are some studies on the nature of the nascent lipid protein complexes in the various organelles (Banerjee and Redman, 1983;Banerjee and Redman, 1984;Howell and Palade, 1982), but detailed understanding of the structure of the nascent lipoprotein particles is not available.…”

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confidence: 99%

Biosynthesis of lipoprotein: location of nascent apoAI and apoB in the rough endoplasmic reticulum of chicken hepatocytes

Dixon

Chattapadhyay

Huima

et al. 1992

The Journal of Cell Biology

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Abstract. Our previous studies showed that in hepatic RER of young chickens, nascent apoAI is not associated with lipoprotein particles and only becomes part of these lipoprotein structures in the Golgi. In this study, we have used three different methodologies to determine the locations of apoAI and apoB in the RER and compared them to that of albumin. Immunoelectron microscopic examination of the RER cell fractions showed that both apoAI and apoB were associated only with the RER membrane whereas albumin was located both within the lumen and on the limiting membrane of the vesicles. To examine the possibility of membrane integration of nascent apoAI and apoB in the RER, we administered L-[3H]leucine to young chickens for 10 min, isolated RER, treated this cell fraction with buffers of varying pH, and measured the release of radioactive albumin, apoAI, and apoB. The majority of nascent apoAI (64%), nascent apoB (100%), and nascent albumin (97%) was released from RER vesicles at pH 11.2, suggesting that, like albumin, apolipoproteins are not integrated within the membrane. To determine if nascent apoproteins are exposed to the cytoplasmic surface, we administered L-[3H]leucine to young chickens and at various times isolated RER and Golgi cell fractions. Radioactive RER and Golgi cell fractions were treated with exogenous protease and the percent of nascent apoAI and apoB accessible to proteolysis was determined and compared to that of albumin. At 5, 10, and 20 min of labeling, 35-56% of nascent apoAI and 60-75% of apoB in RER were degraded, while albumin was refractive to this treatment. At all times both apolipoproteins and albumin present in Golgi cell fractions were protected from proteolysis. These biochemical and morphological findings indicate that apoAI and apoB are associated with the rough microsomal membrane and are partially exposed to the cytoplasmic surface at early stages of secretion. They may later enter the luminal side of the ER and, on entering the Golgi, form lipoprotein particles.

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confidence: 99%

Biosynthesis of lipoprotein: location of nascent apoAI and apoB in the rough endoplasmic reticulum of chicken hepatocytes

Dixon

Chattapadhyay

Huima

et al. 1992

The Journal of Cell Biology

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“…Nascent apolipoproteins are complexed intracellularly with lipids prior to secretion into the blood. The site of synthesis of the apoprotein, lipid (1, 14), and carbohydrate (34) moieties oflipoproteins have been studied but there is little information on the nature of the nascent lipid protein complexes in the various organelles and on the elapsed time of transfer of nascent Apo A-I from its site of synthesis in the rough endoplasmic reticulum (RER) to the Golgi apparatus. Recently we showed that newly synthesized Apo A-I, even though it is present within the RER and the smooth endoplasmic reticulum, failed to float between densities 1.063-1.21 g/ml, whereas the Apo A-I, which is present within the Golgi complex, is capable of floating at a buoyant density similar to that of plasma HDL (3).…”

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Biosynthesis of high density lipoprotein by chicken liver: intracellular transport and proteolytic processing of nascent apolipoprotein A-1.

Banerjee¹,

Mukherjee²,

Redman³

1985

The Journal of Cell Biology

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To study the in vivo processing and secretion of Apolipoprotein A-I (Apo A-I), young chickens were administered individual L-[3H]amino acids intravenously and the time of intracellular transport of nascent Apo A-I from rough endoplasmic reticulum (RER) to the Golgi apparatus was measured. Within 3 to 9 min there was maximal incorporation of radioactivity into Apo A-I in both the RER and the Golgi cell fractions. By contrast, the majority of radioactive albumin was also present in the RER by 3 to 9 min, but did not reach peak amounts in the Golgi fraction until 9 to 25 min. Both radioactive Apo A-I and albumin appeared in the blood at about the same time (between 20 and 30 min). NH2-terminal amino acid sequence analysis of nascent intracellular Apo A-I showed that it contains a pro-hexapeptide extension identical to that of human Apo A-I. After 30 min of administration of radioactive amino acids radioactive Apo A-I was isolated by immunoprecipitation from the liver and serum. NH2-terminal sequence analysis of 20 amino acids indicated that chicken liver contained an equal mixture of nascent pro-Apo A-I and fully processed Apo A-I, whereas the serum only contained processed Apo A-I. Further studies showed that the RER only contained pro-Apo A-I, whereas a mixture of pro-Apo A-I and processed Apo A-I was found in the Golgi complex. These results indicate that, in chicken hepatocytes, there is a more rapid transport of Apo A-I than of albumin from the RER to the Golgi cell fractions, and that Apo A-I remains in the Golgi apparatus for a longer period of time before it is secreted into the blood. In addition these studies show that the in vivo proteolytic processing of chicken pro-Apo A-I to Apo A-I occurs in the Golgi cell fractions.

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“…The role of the carbohydrate moiety in apo B is unknown. Studies carried out with the inhibitor tunicamycin have shown that glycosylation is not required for apo secretion in chick [39] and rat [40] liver, or for the assembly of the protein with its major lipid constituents [40]. A cluster of oligosaccharides has been localized to the putative LDL receptor domain and to some heparin-binding sites of human apo B-100 [4], and it is possible that oligosaccharides protect the precise function of this region of the molecule.…”

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confidence: 99%

Human small-intestinal apolipoprotein B-48 oligosaccharide chains

Sasak

Lown

Colburn

1991

Biochemical Journal

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Hepatic apolipoprotein (apo) B-100 isolated from human plasma is known to contain N-linked oligosaccharides of high-mannose-type and complex-type structures. Sequencing data have revealed that apo B-48 of small-intestinal origin, which represents about 48% of apo B-100 polypeptide from the N-terminus, possesses six potential sites for N-linked oligosaccharides, of which five are likely to be glycosylated. The characterization of the carbohydrate moiety of apo B-48 is the focus of this study. Apo B-48 was labelled with L-[35S]methionine and D-[3H]glucosamine in organ culture of human small-intestinal explants. N-Glycanase treatment resulted in loss of radioactivity from D-[3H]glucosamine-labelled but not L-[35S]methionine-labelled apo B-48 secreted into the medium, and caused no distinct change in mobility of apo B-48 upon electrophoresis on 5% polyacrylamide gel. Analysis of monosaccharide content revealed the presence of 16.8, 17.8, 13.4, 3.4, 2.4 and 2.3 residues of N-acetylglucosamine, mannose, galactose, fucose, xylose and N-acetylgalactosamine respectively. Small-intestinal apo B-48 from human lymph chylomicrons bound to [14C]concanavalin A, and the binding could be inhibited with methyl alpha-D-mannoside. In addition, wheat-germ, peanut, Limulus, soya-bean and Ulex lectins bound apo B-48 specifically. To characterize the carbohydrate moiety further, N-linked oligosaccharides were released by N-Glycanase treatment and reduced with NaB3H4. Labelled oligosaccharides were separated on a concanavalin A-Sepharose column. The majority (78%) were biantennary complex-type structures, 16% were high-mannose type and 6% (not retained by the column) most probably represented higher-branched oligosaccharides. These results suggest the presence of one high-mannose-type and four biantennary complex-type oligosaccharides, as well as probable O-linked sugars in apo B-48. By the use of h.p.l.c., exoglycosidase treatments and ion-exchange chromatography, a mixture of high-mannose-type species with predominant Man8GlcNAc2 as well as monosialylated, desialylated and fucosylated forms of complex-type oligosaccharides were detected.

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Synthesis, processing, and secretion of apolipoprotein B by the chick liver cell.

Cited by 50 publications

References 44 publications

Biosynthesis of lipoprotein: location of nascent apoAI and apoB in the rough endoplasmic reticulum of chicken hepatocytes

Biosynthesis of lipoprotein: location of nascent apoAI and apoB in the rough endoplasmic reticulum of chicken hepatocytes

Biosynthesis of high density lipoprotein by chicken liver: intracellular transport and proteolytic processing of nascent apolipoprotein A-1.

Human small-intestinal apolipoprotein B-48 oligosaccharide chains

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