May 1, 1997Open Access

Normal Activity of Microsomal Triglyceride Transfer Protein Is Required for the Oleate-induced Secretion of Very Low Density Lipoproteins Containing Apolipoprotein B from McA-RH7777 Cells

YWYuwei WangZhejiang University RMRoger S. McLeodDalhousie University ZYZemin YaoChina Academy of Space Technology

Key Points

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Abstract

The requirement of the activity of microsomal triglyceride transfer protein (MTP) for very low density lipoprotein (VLDL) secretion was determined using McA-RH7777 cells stably transfected with human apoB48 (hB48). Secretion of VLDL containing hB48 (hB48-VLDL) by the transfected cells was induced by exogenous oleate (0.4 mm), and oleate-dependent VLDL secretion was selectively inhibited by brefeldin A (0.2 μg/ml). Two protocols were used to determine the effect of MTP inhibition on VLDL secretion. In the first protocol, cell protein and lipid were labeled with radioactive amino acids and oleate prior to MTP inhibition (using 5 μm of the photoaffinity inhibitor BMS-192951 to reduce MTP activity by 65–70%), and secretion of prelabeled apoB and triacylglycerol (TG) associated with lipoproteins was monitored during oleate-supplemented chase. In control cells, a 6-fold increase in incorporation of prelabeled TG into hB48-VLDL was observed after oleate supplement, while incorporation of prelabeled TG into VLDL containing endogenous rat apoB100 (rB100-VLDL) was unaffected. Inhibition of MTP activity abolished the oleate-induced utilization of prelabeled TG (by 80%) and hB48 (by 70%) for hB48-VLDL secretion but decreased utilization of pre-existing TG (by <25%) and B100 (by 45%) for rB100-VLDL secretion to a lesser extent. Inhibition of MTP did not affect incorporation of prelabeled TG or hB48 into high density lipoproteins containing hB48 (hB48-HDL). In the second protocol, MTP was inactivated prior to metabolic labeling of protein and lipid, and secretion of newly labeled apoB and TG as lipoproteins was monitored after oleate supplement. Under this condition, MTP inhibition decreased incorporation of newly labeled TG (by 80%) and hB48 (80%) into hB48-VLDL but did not affect their incorporation into hB48-HDL. Additionally, MTP inhibition decreased incorporation of newly labeled TG (by 50%) and rB100 (by 90%) into rB100-VLDL. Thus, normal activity of MTP is required for the oleate-induced secretion of hB48-VLDL from McA-RH7777 cells. The requirement of the activity of microsomal triglyceride transfer protein (MTP) for very low density lipoprotein (VLDL) secretion was determined using McA-RH7777 cells stably transfected with human apoB48 (hB48). Secretion of VLDL containing hB48 (hB48-VLDL) by the transfected cells was induced by exogenous oleate (0.4 mm), and oleate-dependent VLDL secretion was selectively inhibited by brefeldin A (0.2 μg/ml). Two protocols were used to determine the effect of MTP inhibition on VLDL secretion. In the first protocol, cell protein and lipid were labeled with radioactive amino acids and oleate prior to MTP inhibition (using 5 μm of the photoaffinity inhibitor BMS-192951 to reduce MTP activity by 65–70%), and secretion of prelabeled apoB and triacylglycerol (TG) associated with lipoproteins was monitored during oleate-supplemented chase. In control cells, a 6-fold increase in incorporation of prelabeled TG into hB48-VLDL was observed after oleate supplement, while incorporation of prelabeled TG into VLDL containing endogenous rat apoB100 (rB100-VLDL) was unaffected. Inhibition of MTP activity abolished the oleate-induced utilization of prelabeled TG (by 80%) and hB48 (by 70%) for hB48-VLDL secretion but decreased utilization of pre-existing TG (by <25%) and B100 (by 45%) for rB100-VLDL secretion to a lesser extent. Inhibition of MTP did not affect incorporation of prelabeled TG or hB48 into high density lipoproteins containing hB48 (hB48-HDL). In the second protocol, MTP was inactivated prior to metabolic labeling of protein and lipid, and secretion of newly labeled apoB and TG as lipoproteins was monitored after oleate supplement. Under this condition, MTP inhibition decreased incorporation of newly labeled TG (by 80%) and hB48 (80%) into hB48-VLDL but did not affect their incorporation into hB48-HDL. Additionally, MTP inhibition decreased incorporation of newly labeled TG (by 50%) and rB100 (by 90%) into rB100-VLDL. Thus, normal activity of MTP is required for the oleate-induced secretion of hB48-VLDL from McA-RH7777 cells. Two forms of apolipoprotein B (apoB) 1The abbreviations used are: apo B, apolipoprotein B; VLDL, very low density lipoprotein(s); B48-VLDL, VLDL containing apoB48; ER, endoplasmic reticulum; HDL, high density lipoprotein(s); B48-HDL, HDL containing apoB48; TG, triacylglycerol; MTP, microsomal triglyceride transfer protein; TLC, thin layer chromatography; hB48, human apoB48; DMEM, Dulbecco's modified Eagle's medium; rB100-VLDL, VLDL containing rB100; hB48-VLDL, VLDL containing hB48; hB48-HDL, HDL containing hB48; rB48, rat apoB48; PC, phosphatidylcholine. 1The abbreviations used are: apo B, apolipoprotein B; VLDL, very low density lipoprotein(s); B48-VLDL, VLDL containing apoB48; ER, endoplasmic reticulum; HDL, high density lipoprotein(s); B48-HDL, HDL containing apoB48; TG, triacylglycerol; MTP, microsomal triglyceride transfer protein; TLC, thin layer chromatography; hB48, human apoB48; DMEM, Dulbecco's modified Eagle's medium; rB100-VLDL, VLDL containing rB100; hB48-VLDL, VLDL containing hB48; hB48-HDL, HDL containing hB48; rB48, rat apoB48; PC, phosphatidylcholine.are synthesized by the rat liver, the full-length apoB100 and apoB48, which represents the N-terminal 48% of apoB100 (1Chan L. J. Biol. Chem. 1992; 267: 25621-25624Abstract Full Text PDF PubMed Google Scholar). Although the physiological significance of having two forms of apoB in rat liver is not clear (2Farese Jr., R.V. Véniant M.M. Cham C.M. Flynn L.M. Pierotti V. Loring J.F. Ruland S. Stokowski R.S. Huszar D. Young S.G. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 6393-6398Crossref PubMed Scopus (112) Google Scholar), both forms of apoB have the ability to assemble very low density lipoproteins (VLDL) (3Elovson J. Chatterton J.E. Bell G.T. Schumaker V.N. Reuben M.A. Puppione D.L. Reeve Jr., J.R. Young N.L. J. Lipid Res. 1988; 29: 1461-1473Abstract Full Text PDF PubMed Google Scholar). The mechanism by which hepatic VLDL is synthesized has not been completely defined. However, significant progress has been made over the past several years concerning the formation and secretion of VLDL containing apoB48 (B48-VLDL) (4Innerarity T.L. Borén J. Yamanaka S. Olofsson S.-O. J. Biol. Chem. 1996; 271: 2353-2356Abstract Full Text Full Text PDF PubMed Scopus (122) Google Scholar). Biochemical evidence has been obtained through studies with primary rat hepatocytes (5Elovson, J., and Doolittle, M. H. (1994) Circulation, 90, Suppl. I, 185 (abstr.).Google Scholar, 6Swift L.L. J. Lipid Res. 1995; 36: 395-406Abstract Full Text PDF PubMed Google Scholar) and the rat hepatoma cell line McA-RH7777 (7Borén J. Rustaeus S. Olofsson S.-O. J. Biol. Chem. 1994; 269: 25879-25888Abstract Full Text PDF PubMed Google Scholar) that B48-VLDL is assembled via two discontinuous lipidation stages in the endoplasmic reticulum (ER). In the first stage, apoB48 is associated with a small amount of lipid to form a primordial particle with high buoyant density. These high density lipoprotein (HDL) particles (designated B48-HDL) may be secreted from the cells if further lipid recruitment does not occur. Alternatively, the B48-HDL particle can undergo a second lipidation stage, expanding its lipid content, primarily triacylglycerol (TG), to form VLDL. This “two-step” assembly model is consistent with the early immunohistochemical studies of hepatic VLDL assembly in rats (8Alexander C.A. Hamilton R.L. Havel R.J. J. Cell Biol. 1976; 69: 241-263Crossref PubMed Scopus (252) Google Scholar). In rat hepatoma cells, the conversion of B48-HDL into B48-VLDL is associated with increased synthesis of cellular lipid, and the process can be inhibited by brefeldin A (9Rustaeus S. Lindberg K. Borén J. Olofsson S.-O. J. Biol. Chem. 1995; 270: 28879-28886Abstract Full Text Full Text PDF PubMed Scopus (66) Google Scholar) or cycloheximide (7Borén J. Rustaeus S. Olofsson S.-O. J. Biol. Chem. 1994; 269: 25879-25888Abstract Full Text PDF PubMed Google Scholar). These results suggest that in addition to active TG synthesis, other factors involved in vesicular trafficking or lipid mobilization may participate in the second stage of B48-VLDL formation. The microsomal triglyceride transfer protein (MTP) is a heterodimeric protein consisting of a 97-kDa catalytic subunit noncovalently linked to protein-disulfide isomerase (10Gordon D.A. Wetterau J. Gregg R.E. Trends Cell Biol. 1995; 5: 317-321Abstract Full Text PDF PubMed Scopus (118) Google Scholar). The ability of MTP to transfer TG between lipid membranes has been demonstrated in vitro (11Jamil H. Dickson Jr., J.K. Chu C.-H. Lago M.W. Rinehart J.K. Biller S.A. Gregg R.E. Wetterau J.R. J. Biol. Chem. 1995; 270: 6549-6554Abstract Full Text Full Text PDF PubMed Scopus (143) Google Scholar), and deficiency in the MTP activity is associated with human abetalipoproteinemia (12Wetterau J.R. Aggerbeck L.P. Bouma M.-E. Eisenberg C. Munck A. Hermier M. Schmitz J. Gay G. Rader D.J. Gregg R.E. Science. 1992; 258: 999-1001Crossref PubMed Scopus (626) Google Scholar, 13Sharp D. Blinderman L. Combs K.A. Kienzle B. Ricci B. Wager-Smith K. Gil C.M. Turck C.W. Bouma M.-E. Rader D.J. Aggerbeck L.P. Gregg R.E. Gordon D.A. Wetterau J.R. Nature. 1993; 365: 65-69Crossref PubMed Scopus (394) Google Scholar). In HepG2 cells, a physical association between the hydrophobic sequences of apoB and MTP has been detected (14Wu X. Zhou M. Huang L.-S. Wetterau J. Ginsberg H.N. J. Biol. Chem. 1996; 271: 10277-10281Abstract Full Text Full Text PDF PubMed Scopus (122) Google Scholar). It is unclear, however, whether the direct interaction between MTP and apoB is essential for the recruitment of lipid. The functional role of MTP in the secretion of lipoproteins containing apoB has been demonstrated by co-expression of MTP and apoB in heterologous cells that normally produce neither protein (15Gordon D.A. Jamil H. Sharp D. Mullaney D. Yao Z. Gregg R.E. Wetterau J. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 7628-7632Crossref PubMed Scopus (189) Google Scholar, R.J. D.J. J. J. Biol. Chem. 1994; 269: Full Text PDF PubMed Google Scholar, J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar, S. R.S. Gordon D.A. Yao Z. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar). from that MTP role in the assembly and secretion of lipoproteins containing However, cell used for the the ability to and VLDL, the requirement for MTP activity in VLDL in the second not be Thus, to the of MTP in VLDL synthesis is to that can MTP in of MTP using MTP has been by several to apoB secretion from cells of hepatic or D.A. Jamil H. Gregg R.E. Olofsson S.-O. Borén J. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar, H. Gordon D.A. C.M. Dickson Jr., J.K. Ricci B. Chu C.-H. Jr., Biller S.A. Gregg R.E. Wetterau J.R. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: PubMed Scopus Google Scholar, J. 1996; PubMed Scopus Google Scholar, M. D. J. Lipid Res. 1996; Full Text PDF PubMed Google Scholar). the inhibition of MTP activity decreased secretion of the full-length However, the effect of MTP inhibition on apoB48 secretion was In cells, while MTP inhibition in decreased secretion of secretion of apoB48 was M. D. J. Lipid Res. 1996; Full Text PDF PubMed Google Scholar). In McA-RH7777 cells, of MTP to affect formation of B48-HDL the of the first but effect on the conversion of B48-HDL into B48-VLDL D.A. Jamil H. Gregg R.E. Olofsson S.-O. Borén J. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar). The effect of MTP inhibition on apoB48 secretion in cells has not been Z. Young S.G. J. Biol. Chem. Full Text PDF PubMed Google Scholar, D.L. R.J. J. PubMed Scopus Google Scholar, D.J. Chatterton J.E. J. Schumaker V.N. J. Biol. Chem. 1992; 267: Full Text PDF PubMed Google Scholar, R.S. J. Yao Z. J. Biol. Chem. 1994; 269: Full Text PDF PubMed Google Scholar) have evidence that the of lipoproteins is with the of the associated apoB during the first However, the of apoB does not to a role in the conversion of into during the second In McA-RH7777 cells stably human apoB or HDL that apoB as as or containing a of apoB as as of were into VLDL in the of exogenous oleate R.S. S. A. Yao Z. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar). Thus, conversion of into in the second may be determined primarily by synthesis of lipid and by protein factors that the lipid during assembly apoB MTP role in lipid that the MTP activity is required for the mobilization of lipid that is for the oleate-induced B48-VLDL This was in the and were obtained from for were obtained from apoB was obtained from and protein were obtained from was obtained from was obtained from of and and were obtained from and were obtained from thin layer and used for were obtained from A was obtained from The MTP inhibitor BMS-192951 was a of D. Gordon for human apoB was obtained from and of McA-RH7777 cells stably transfected with the human apoB48 were and in Dulbecco's modified Eagle's as M.M. Yao Z. Biol. 1995; PubMed Scopus Google Scholar). Inhibition of MTP in McA-RH7777 cells with the BMS-192951 was to D.A. Jamil H. Gregg R.E. Olofsson S.-O. Borén J. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar). The cells were with of in of oleate for to The were to 5 with and to density as (7Borén J. Rustaeus S. Olofsson S.-O. J. Biol. Chem. 1994; 269: 25879-25888Abstract Full Text PDF PubMed Google Scholar). were from the of the The apoB were and by and as R.S. S. A. Yao Z. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar). were with of or of for and during the were with of In after labeling a or was prior to the of to and of MTP with BMS-192951 was during the VLDL and HDL were by and lipoproteins containing rat B100 or hB48 or were by associated with particles were with of and was on as Yao Z. Biol. 1995; PubMed Scopus Google Scholar) using as a Cell were and by The associated with lipid was by was to to the obtained from of the labeling were with and of for to was the was with to lipoproteins containing endogenous rat with or were with of by lipoproteins were to density and and rB100-VLDL, and were to lipid The of hB48-VLDL from the by was and the hB48-VLDL endogenous rB100-VLDL as determined by and by of not In McA-RH7777 cells transfected with hB48, the secretion of hB48-VLDL is oleate R.S. S. A. Yao Z. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar). This is to the oleate-induced secretion of VLDL containing rat in McA-RH7777 cells (7Borén J. Rustaeus S. Olofsson S.-O. J. Biol. Chem. 1994; 269: 25879-25888Abstract Full Text PDF PubMed Google Scholar). of hB48 endogenous secretion M.M. Yao Z. Biol. 1995; PubMed Scopus Google Scholar). further that hB48-VLDL secretion was with endogenous the of the transfected cell line to brefeldin A. It has been that the oleate-induced assembly second was to low of brefeldin A (9Rustaeus S. Lindberg K. Borén J. Olofsson S.-O. J. Biol. Chem. 1995; 270: 28879-28886Abstract Full Text Full Text PDF PubMed Scopus (66) Google Scholar). that in McA-RH7777 cells, secretion of hB48-VLDL was to brefeldin A. A of that were secreted as lipoproteins from the cells with or brefeldin and B the of apolipoprotein secretion. In the of brefeldin hB48 was secreted as both VLDL and HDL In the of brefeldin secretion of hB48-VLDL was decreased by with secretion of was not decreased endogenous rB100-VLDL secretion was decreased by by the brefeldin A In the amount of associated with rB100 was in the brefeldin cells control a labeling but the associated with hB48 was not of TG and was not by brefeldin A as by the incorporation of during a labeling not These results that for the second hB48-VLDL assembly and secretion in the transfected McA-RH7777 cells. Thus, in the the cells were used to the requirement of MTP activity for the oleate-induced B48-VLDL secretion. lipid incorporation into secreted B48-VLDL, first labeled the lipid with this labeling oleate was not the cells did not produce the cells were and with containing oleate to the second was not in the of control cells. In was in both to newly synthesized lipid. A that incorporation of TG into secreted rB100-VLDL, hB48-VLDL, or was increased by and by exogenous incorporation of into secreted rB100-VLDL and hB48-VLDL not was increased by the oleate These results suggest that oleate newly synthesized TG is for both hB48-VLDL and rB100-VLDL secretion. However, a was observed between incorporation of prelabeled into secreted hB48-VLDL or rB100-VLDL. oleate effect on incorporation of into rB100-VLDL, increased incorporation of into hB48-VLDL 6-fold of associated with hB48-VLDL be of secretion of prelabeled TG, of the with oleate TG synthesis and decreased the activity of the was a in the incorporation of into the of a The effect of oleate on the incorporation of prelabeled into the secreted however, was to that for newly synthesized These results that the associated with the hB48-VLDL secretion is the utilization of pre-existing In the used incorporation of pre-existing TG into hB48-VLDL as a to the requirement of MTP activity in the second A MTP BMS-192951 D.A. Jamil H. Gregg R.E. Olofsson S.-O. Borén J. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar), was used to MTP in McA-RH7777 cells. with cells for and for BMS-192951 5 or μm the MTP activity by The effect for not The effect of MTP inhibition on the recruitment of prelabeled TG during oleate-induced second was determined by labeling of the cells with MTP with BMS-192951 and lipoprotein secretion during oleate-supplemented chase. In that a or between and did not affect the oleate-induced secretion of pre-existing TG as a or incorporation of prelabeled TG into secreted hB48-VLDL was increased by oleate while incorporation of prelabeled TG into rB100-VLDL was TG synthesis during oleate-supplemented did not the of the prelabeled TG or PC, a small increase in labeled TG and a in labeled were observed and These results that the recruitment of pre-existing TG for hB48-VLDL secretion is not after a or However, recruitment of pre-existing TG for hB48-VLDL secretion was abolished by MTP inhibition 5 μm incorporation of rB100 and hB48 into secreted VLDL decreased by and as with cells with inhibitor and MTP inhibition effect on secretion of hB48 with of MTP decreased the incorporation of lipid into VLDL. Secretion of prelabeled and associated with hB48-VLDL was decreased by and the of a Inhibition of MTP did not affect secretion of associated with or with rB100-VLDL, and decreased the secretion of with rB100-VLDL or with These results evidence that utilization of pre-existing TG for the oleate-induced hB48-VLDL secretion is to MTP The small effect on secretion of pre-existing TG as rB100-VLDL that a amount of rB100-VLDL particles the oleate-induced second The of prelabeled and in cells during oleate-supplemented was to that in cells and that MTP did not the of prelabeled lipid. did MTP inhibition affect secretion of endogenous rat as HDL not the effect of MTP inhibition on VLDL secretion by MTP prior to metabolic labeling of apoB and lipid. of MTP the secretion of apoB associated with hB48-VLDL (by 80%) or rB100-VLDL (by 90%) as with cells inhibitor A and MTP inhibition decreased secretion of associated with hB48-VLDL (by or rB100-VLDL (by The in rB100 in TG in the rB100-VLDL that the amount of secreted rB100-VLDL is with newly labeled However, to in secretion of or associated with was by MTP inhibition and These results of the effect of brefeldin A on hB48-VLDL secretion and further that MTP activity is required for the oleate-induced secretion of Under incorporation of into cellular TG was decreased by of during a labeling in the cells with cells, incorporation of into was not of In the have whether or not MTP activity is required for B48-VLDL secretion using two MTP was inactivated or after metabolic labeling to secretion of pre-existing or newly synthesized apoB and lipid as of the that hB48-VLDL be from the of McA-RH7777 cells, which the ability to hB48-VLDL oleate of the that pre-existing TG was for hB48-VLDL which be used to the oleate-induced second In the that MTP activity was by secretion of newly synthesized or prelabeled hB48 and TG as hB48-VLDL was their secretion as was unaffected. secretion of pre-existing TG as hB48-VLDL that was during the oleate-induced second and was to MTP inhibition These results suggest that expanding the lipid during hB48-VLDL assembly be with normal MTP that secretion of VLDL containing other apoB or was induced by oleate R.S. S. A. Yao Z. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar). secretion of VLDL containing or by the transfected McA-RH7777 cells oleate was abolished by MTP S. and Z. These results evidence that the oleate-induced VLDL secretion is not determined by apoB and that MTP is of the second VLDL assembly and secretion. two between hB48-VLDL and rB100-VLDL secretion in the lipid recruitment and its to MTP The first was in the of incorporation of pre-existing TG oleate Although both newly synthesized and pre-existing TG be for rB100-VLDL and hB48-VLDL pre-existing TG to be into hB48-VLDL oleate-induced VLDL secretion. Thus, while was a 6-fold increase in the incorporation of pre-existing TG into hB48-VLDL, was in secretion of pre-existing TG as rB100-VLDL oleate and These results that assembly and secretion of B48-VLDL and be through The second was observed in the of pre-existing TG recruitment to MTP Although of MTP (by decreased secretion of both newly synthesized and pre-existing TG as B48-VLDL and the effect of MTP inhibition was to the incorporation of pre-existing TG into B48-VLDL during oleate-induced second Thus, while secretion of pre-existing TG as hB48-VLDL was decreased by incorporation of pre-existing TG into rB100-VLDL was by MTP inhibition The incorporation of pre-existing TG into hB48-VLDL and its to MTP inhibition evidence to the assembly that lipid is to a primordial B48-HDL particle in the oleate-induced second (4Innerarity T.L. Borén J. Yamanaka S. Olofsson S.-O. J. Biol. Chem. 1996; 271: 2353-2356Abstract Full Text Full Text PDF PubMed Scopus (122) Google Scholar). In to the of B100 to pre-existing TG in to oleate and its to MTP inhibition the that is assembled primarily through a process oleate (7Borén J. Rustaeus S. Olofsson S.-O. J. Biol. Chem. 1994; 269: 25879-25888Abstract Full Text PDF PubMed Google Scholar, J. L. M. J. A. Olofsson S.-O. J. Biol. Chem. 1992; 267: Full Text PDF PubMed Google Scholar), and that lipid recruitment may not a role in The that MTP activity results in decreased hB48-VLDL secretion is of the effect of brefeldin A on VLDL secretion It has been that B48-VLDL secretion induced by exogenous oleate in McA-RH7777 cells can be inhibited by a low of brefeldin A (9Rustaeus S. Lindberg K. Borén J. Olofsson S.-O. J. Biol. Chem. 1995; 270: 28879-28886Abstract Full Text Full Text PDF PubMed Scopus (66) Google Scholar). brefeldin A with the formation of essential for vesicular J.E. Nature. 1994; PubMed Scopus Google Scholar), this that of or other trafficking may be of the second the effect of a low of MTP inhibitor on lipoprotein secretion was to that of brefeldin secretion of hB48-VLDL secretion of and Thus, MTP may a role in formation of TG the of the second by mobilization of the cellular TG, that be to brefeldin A. the was not to the between and hB48-VLDL, the effect of MTP on this conversion was not The between brefeldin vesicular and TG mobilization during the second VLDL assembly further Although their on hB48-VLDL secretion were brefeldin A and MTP inhibitor on the synthesis of brefeldin A (0.2 effect on cell TG synthesis, of MTP by the inhibitor BMS-192951 decreased (by the incorporation of into cell TG metabolic labeling was after MTP The in TG synthesis not be by increased of the labeled TG, the that MTP inhibition effect on the of the prelabeled cell TG was the decreased TG synthesis the of of lipid synthesis in incorporation of into cell was not is for the TG synthesis by MTP In the the of a low of MTP inhibitor of the cellular MTP the MTP activity and the active TG synthesis were to normal secretion of the of first as hB48-HDL. or not the decreased TG synthesis (by that was associated with MTP to the second hB48-VLDL assembly to be for human abetalipoproteinemia the activity of the MTP from the functional is for normal lipoprotein using MTP in as human apoB (2Farese Jr., R.V. Véniant M.M. Cham C.M. Flynn L.M. Pierotti V. Loring J.F. Ruland S. Stokowski R.S. Huszar D. Young S.G. Proc. Natl. Acad. Sci. U. S. A. 1996; 93: 6393-6398Crossref PubMed Scopus (112) Google Scholar) or using MTP into the requirement of MTP in VLDL in The model for the of MTP in hepatic B48-VLDL synthesis but the role that MTP in the early stage of lipoprotein assembly D.A. Jamil H. Gregg R.E. Olofsson S.-O. Borén J. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar). In cell as and cells that not normally apoB or MTP, of MTP and apoB in secretion of apoB (15Gordon D.A. Jamil H. Sharp D. Mullaney D. Yao Z. Gregg R.E. Wetterau J. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 7628-7632Crossref PubMed Scopus (189) Google Scholar, R.J. D.J. J. J. Biol. Chem. 1994; 269: Full Text PDF PubMed Google Scholar, J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar, S. R.S. Gordon D.A. Yao Z. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar). In cells, the requirement of MTP and oleate-induced to be a of apoB that is between lipid MTP and the hydrophobic of In transfected cells that MTP the were to the microsomal or were after J. S. Proc. Natl. Acad. Sci. U. S. A. 1992; PubMed Scopus Google Scholar). Thus, of MTP in McA-RH7777 cells be to apoB synthesis, by of or by of newly synthesized S. R.S. Gordon D.A. Yao Z. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar). The of MTP inhibition on apoB synthesis and on the secretion of the first and to be a of apoB D. and Z. Suppl. I, (abstr.).Google Scholar). In cells with MTP secretion of was not secretion of B100 was abolished M. D. J. Lipid Res. 1996; Full Text PDF PubMed Google Scholar). in a cell line that MTP assembly and secretion of the transfected N-terminal of human apoB on lipoproteins has been observed H. A. V. Proc. Natl. Acad. Sci. U. S. A. 1995; PubMed Scopus Google Scholar). These the that the requirement of MTP activity for apoB secretion is with the of apoB and with the of lipid recruitment S. R.S. Gordon D.A. Yao Z. J. Biol. Chem. 1996; 271: Full Text Full Text PDF PubMed Scopus Google Scholar). The evidence that the activity of MTP, with the brefeldin and other protein may the second of B48-VLDL assembly and B48-VLDL secretion. These induced by exogenous oleate in McA-RH7777 cells, may mobilization of pre-existing TG for the of the lipid of VLDL. In the lipid mobilization may the first lipid assembly by a that has been to be by Thus, MTP activity is required for the VLDL assembly both and addition of lipid. to D. Gordon for in protocols during the early stage of this and to H. Jamil for the MTP activity X. and J. for and and of for

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