Secretion of apolipoprotein B-containing lipoproteins from HeLa cells is dependent on expression of the microsomal triglyceride transfer protein and is regulated by lipid availability.

Gordon, David; Jamil, Haris; Sharp, Daru; Mullaney, David; Yao, Zemin; Gregg, Richard E.; Wetterau, John R.

doi:10.1073/pnas.91.16.7628

Cited by 191 publications

(158 citation statements)

References 28 publications

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“…Previous studies have demonstrated that MTP is essential for the assembly of apoB-containing lipoproteins (12,13). Moreover, it has been shown that mutations in the MTP gene give rise to the phenotype of abetalipoproteinemia (10,11).…”

Section: Discussionmentioning

confidence: 99%

“…Mutations that inactivate or abolish MTP give rise to the phenotype of abetalipoproteinemia, an almost total absence of apoB-containing lipoproteins in the plasma (10,11). Thus MTP is essential for the assembly and secretion of apoB-containing lipoproteins (12,13). Very recent results indicate that MTP is important for the early events in the assembly of endogenous rat apoB-48 VLDL.…”

mentioning

confidence: 99%

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The Microsomal Triglyceride Transfer Protein Catalyzes the Post-translational Assembly of Apolipoprotein B-100 Very Low Density Lipoprotein in McA-RH7777 Cells

Rustaeus

Stillemark

Lindberg

et al. 1998

Journal of Biological Chemistry

118

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In cells in which the lipoprotein assembly process had been inactivated by brefeldin A (BFA), membrane-associated apoB-100 disappeared without forming lipoproteins or being secreted, indicating that it was degraded. Reactivation of the assembly process by chasing the cells in the absence of BFA, gave rise to a quantitative recovery of the membrane-associated apoB-100 in the very low density lipoprotein (VLDL) fraction in the medium. These results indicate that the membrane-associated apoB-100 can be converted to VLDL.A new method was developed by which the major amount (88%) of microsomal apoB-100 but not integral membrane proteins could be extracted. The major effect of this method was to increase the recovery of apoB-100 that banded in the LDL and HDL density regions, suggesting that the membrane-associated form of apoB-100 is partially lipidated. We also investigated the role of the microsomal triglyceride transfer protein (MTP) in the assembly of apoB-100 VLDL using a photoactivatable MTP inhibitor (BMS-192951). This compound strongly inhibited the assembly and secretion of apoB-100 VLDL when present during the translation of the protein. To investigate the importance of MTP during the later stages in the assembly process, the cells were preincubated with BFA (to reversibly inhibit the assembly of apoB-100 VLDL) and pulse-labeled (؉BFA) and chased (؉BFA) for 30 min to obtain full-length apoB-100 associated with the microsomal membrane. Inhibition of MTP after the 30-min chase blocked assembly of VLDL. This indicates that MTP is important for the conversion of full-length apoB-100 into VLDL. Results from experiments in which a second chase (؊BFA) was introduced before the inactivation of MTP indicated that only early events in this conversion of full-length apoB-100 into VLDL were blocked by the MTP inhibitor. Together these results indicate that there is a MTP-dependent "window" in the VLDL assembly process that occurs after the completion of apoB-100 but before the major amount of lipids is added to the VLDL particle. Thus the assembly of apoB-100 VLDL from membrane-associated apoB-100 involves an early MTP-dependent phase and a late MTP-independent phase, during which the major amount of lipid is added.

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Section: Discussionmentioning

confidence: 99%

mentioning

confidence: 99%

The Microsomal Triglyceride Transfer Protein Catalyzes the Post-translational Assembly of Apolipoprotein B-100 Very Low Density Lipoprotein in McA-RH7777 Cells

Rustaeus

Stillemark

Lindberg

et al. 1998

Journal of Biological Chemistry

118

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“…Inactivation of MTP in hepatic or intestinal cells invariably impairs apoB assembly and secretion (7,20,21). On the other hand, coexpression of the MTP 97-kDa subunit with recombinant apoB facilitates assembly of small, dense lipoproteins containing apoB in heterologous cells (22)(23)(24)(25). Although these studies have provided strong evidence that MTP is essential for apoB secretion as lipoproteins, our knowledge on the mechanism by which MTP facilitates VLDL assembly is incomplete and controversial.…”

mentioning

confidence: 92%

“…Initial transfection studies with non-hepatic cell lines (22)(23)(24)(25) suggested that the major function of MTP was to assist translocation of apoB across the membrane of ER. Since it was believed that the formation of primordial dense particles took place during apoB translocation, some researchers postulated that MTP activity was essential only in the early stage of apoB lipidation (26).…”

mentioning

confidence: 99%

The Activity of Microsomal Triglyceride Transfer Protein Is Essential for Accumulation of Triglyceride within Microsomes in McA-RH7777 Cells

Wang¹,

Tran²,

Yao³

1999

Journal of Biological Chemistry

128

140

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Previously, based on distinct requirement of microsomal triglyceride transfer protein (MTP) and kinetics of triglyceride (TG) utilization, we concluded that assembly of very low density lipoproteins (VLDL) containing B48 or B100 was achieved through different paths (Wang, Y., McLeod, R. S., and Yao, Z. (1997) J. Biol. Chem. 272, 12272-12278). To test if the apparent dual mechanisms were accounted for by apolipoprotein B (apoB) length, we studied VLDL assembly using transfected cells expressing various apoB forms (e.g. B64, B72, B80, and B100). For each apoB, enlargement of lipoprotein to form VLDL via bulk TG incorporation was induced by exogenous oleate, which could be blocked by MTP inhibitor BMS-197636 treatment. While particle enlargement was readily demonstrable by density ultracentrifugation for B64-and B72-VLDL, it was not obvious for B80-and B100-VLDL unless the VLDL was further resolved by cumulative rate flotation into VLDL 1 (S f > 100) and VLDL 2 (S f 20 -100). BMS-197636 diminished B100 secretion in a dose-dependent manner (0.05-0.5 M) and also blocked the particle enlargement from small to large B100-lipoproteins. These results yield a unified model that can accommodate VLDL assembly with all apoB forms, which invalidates our previous conclusion. To gain a better understanding of the MTP action, we examined the effect of BMS-197636 on lipid and apoB synthesis during VLDL assembly. While BMS-197636 (0.2 M) entirely abolished B100-VLDL 1 assembly/secretion, it did not affect B100 translation or translocation across the microsomal membrane, nor did it affect TG synthesis and cell TG mass. However, BMS-197636 drastically decreased accumulation of [ 3 H]glycerol-labeled TG and TG mass within microsomal lumen. The decreased TG accumulation was not a result of impaired B100-VLDL assembly, because in cells treated with brefeldin A (0.2 g/ml), the assembly of B100-VLDL was blocked yet lumenal TG accumulation was normal. Thus, MTP plays a role in facilitating accumulation of TG within microsomes, a prerequisite for the post-translational assembly of TG-enriched VLDL.

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“…43 Assuming that these effects occur in vivo, a reduced degradation of apo B caused by increased FFA, together with increased MTP activity, would accelerate the secretion of VLDL particles from the liver. 44,45 In 30-week-old OLETF rats, plasma triglyceride levels were over eightfold higher and plasma cholesterol levels 1.5-fold higher than in LETO rats of the same age. Insulin resistance and NIDDM are manifested at this stage.…”

Section: Figmentioning

confidence: 99%

Enhanced expression of hepatic acyl-coenzyme A synthetase and microsomal triglyceride transfer protein messenger RNAs in the obese and hypertriglyceridemic rat with visceral fat accumulation

et al. 1998

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The liver plays a central role in lipoprotein metabolism. In particular, very-low density lipoprotein (VLDL) is assembled in the hepatocytes and secreted into the blood circulation. The VLDL is then catabolized to low-density lipoprotein by lipoprotein lipase and hepatic triglyceride lipase. Obese subjects, especially those with visceral fat accumulation, are frequently associated with hyperlipidemia, non-insulin-dependent diabetes mellitus (NIDDM), and hypertension. The mechanism of hyperlipidemia in visceral fat obesity has not yet been elucidated. Otsuka Long-Evans Tokushima Fatty (OLETF) rat is an animal model of NIDDM, characterized by obesity with visceral fat accumulation, hyperlipidemia, and late-onset insulin resistance. To elucidate the mechanism of hyperlipidemia observed in OLETF rats, we focused on the production of VLDL by the liver and investigated hepatic messenger RNA (mRNA) levels of microsomal triglyceride transfer protein (MTP), acyl-coenzyme A synthetase (ACS), and apolipoprotein B (apo B), which play important roles in VLDL synthesis and secretion. In 6-week-old OLETF rats, in which insulin resistance had not been manifested, visceral fat weight was already higher and portal free fatty acid (FFA) and VLDL-triglyceride levels were elevated compared with the control rats. Hepatic ACS activity and mRNA levels, and MTP mRNA levels were also increased in OLETF rats, whereas apo B mRNA levels were similar; these results suggest that the enhanced expression of both ACS and MTP genes associated with visceral fat accumulation before developing insulin resistance may be involved in the pathogenesis of hyperlipidemia in obese animal models with NIDDM. (HEPATOLOGY 1998;27:557-562.)

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Secretion of apolipoprotein B-containing lipoproteins from HeLa cells is dependent on expression of the microsomal triglyceride transfer protein and is regulated by lipid availability.

Cited by 191 publications

References 28 publications

The Microsomal Triglyceride Transfer Protein Catalyzes the Post-translational Assembly of Apolipoprotein B-100 Very Low Density Lipoprotein in McA-RH7777 Cells

The Microsomal Triglyceride Transfer Protein Catalyzes the Post-translational Assembly of Apolipoprotein B-100 Very Low Density Lipoprotein in McA-RH7777 Cells

The Activity of Microsomal Triglyceride Transfer Protein Is Essential for Accumulation of Triglyceride within Microsomes in McA-RH7777 Cells

Enhanced expression of hepatic acyl-coenzyme A synthetase and microsomal triglyceride transfer protein messenger RNAs in the obese and hypertriglyceridemic rat with visceral fat accumulation

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