Microsomal Triacylglycerol Transfer Protein Is Required for Lumenal Accretion of Triacylglycerol Not Associated with ApoB, as Well as for ApoB Lipidation

The assembly of very low density lipoproteins in hepatocytes requires the microsomal triacylglycerol transfer protein (MTP). This microsomal lumenal protein transfers lipids, particularly triacylglycerols (TG), between membranesin vitro and has been proposed to transfer TG to nascent apolipoprotein (apo) B in vivo. We examined the role of MTP in the assembly of apoB-containing lipoproteins in cultured murine primary hepatocytes using an inhibitor of MTP. The MTP inhibitor reduced TG secretion from hepatocytes by 85% and decreased the amount of apoB100 in the microsomal lumen, as well as that secreted into the medium, by 70 and 90%, respectively, whereas the secretion of apoB48 was only slightly decreased and the amount of lumenal apoB48 was unaffected. However, apoB48-containing particles formed in the presence of inhibitor were lipid-poor compared with those produced in the absence of inhibitor. We also isolated a pool of apoB-free TG from the microsomal lumen and showed that inhibition of MTP decreased the amount of TG in this pool by ∼45%. The pool of TG with was However, inhibition of MTP TG transfer from the TG pool to in the microsomal We that MTP TG in the microsomal lumen and as a of TG assembly with of MTP the of TG to assembly in murine The assembly of very low density lipoproteins in hepatocytes requires the microsomal triacylglycerol transfer protein (MTP). This microsomal lumenal protein transfers lipids, particularly triacylglycerols (TG), between membranesin vitro and has been proposed to transfer TG to nascent apolipoprotein (apo) B in vivo. We examined the role of MTP in the assembly of apoB-containing lipoproteins in cultured murine primary hepatocytes using an inhibitor of MTP. The MTP inhibitor reduced TG secretion from hepatocytes by 85% and decreased the amount of apoB100 in the microsomal lumen, as well as that secreted into the medium, by 70 and 90%, respectively, whereas the secretion of apoB48 was only slightly decreased and the amount of lumenal apoB48 was unaffected. However, apoB48-containing particles formed in the presence of inhibitor were lipid-poor compared with those produced in the absence of inhibitor. We also isolated a pool of apoB-free TG from the microsomal lumen and showed that inhibition of MTP decreased the amount of TG in this pool by ∼45%. The pool of TG with was However, inhibition of MTP TG transfer from the TG pool to in the microsomal We that MTP TG in the microsomal lumen and as a of TG assembly with of MTP the of TG to assembly in murine very low density apolipoprotein low density microsomal triacylglycerol transfer protein triacylglycerol assembly of very low density lipoproteins in the triacylglycerol the of apolipoprotein (apo) microsomal lumenal the microsomal triacylglycerol transfer protein has been in the of TG by nascent assembly and secretion of in with the a in the MTP and MTP protein and MTP transfer a in MTP has the to transfer TG and lipids, and between in vitro and has been proposed to transfer TG to nascent apoB-containing lipoproteins in vivo. This with that and MTP of assembly MTP in the and as a with a protein of the lumen that protein The MTP transfer to the in vitro the transfer of MTP that MTP transfers between a The and of and between in that MTP in the of with of the MTP transfer been that by to in and of in apoB-containing lipoproteins from the to the However, were in the MTP was in the compared with apoB100 were reduced by apoB48 were reduced by only of the that MTP TG the of apoB48 in the of was in particles that were only apoB100 secretion from cultured hepatocytes was whereas apoB48 secretion was only slightly reduced compared with that in hepatocytes that compared with very particles the and in to in the particles in the lumen of that lumenal TG as a of TG assembly with The of MTP secretion by the that of MTP in of apoB100 and as well as TG has been to in as a that the of the in the of into the lumen in the transfer of the of TG to in the lumen and in the transfer of TG into the lumen assembly into The of the role of MTP been in cultured that compared with primary in the of in The of MTP has also been in and that lipoproteins in secretion was by with MTP and of the the role of MTP in assembly to the a been the of MTP in primary hepatocytes the role of MTP has been in MTP were the of MTP in hepatocytes from of as of to the of assembly in murine the of MTP assembly and secretion in murine primary that secretion of apoB100 to inhibition of MTP and that MTP the apoB48 particles produced lipid-poor compared with those in hepatocytes with MTP We also isolated from the microsomal lumen a pool of TG that with and that inhibition of MTP the amount of TG this However, inhibition of MTP the transfer of TG from this pool to lipid-poor apoB-containing the role of MTP in the assembly of apoB-containing lipoproteins in murine that inhibition of MTP by inhibition of TG the secretion of whereas the amount of apoB48 secreted was the of apoB48 was were in the and of of apoB100 and apoB48 in the microsomal The in of apoB100 and apoB48 to inhibition of MTP was of the presence of We isolated a microsomal lumenal pool of TG that was with and showed that the of this pool was the pool of TG with the MTP inhibitor decreased by the amount of apoB-free and TG in the microsomal lumen, inhibition of MTP the transfer of TG to of MTP the of and of the the role of MTP in assembly to the to this MTP has been proposed to as a that in the and into the lumen that TG secretion by 85% the amount of apoB100 in the microsomal lumen reduced by whereas the amount of apoB48 unaffected. This with in the murine MTP been in the hepatocytes from apoB100 secretion was whereas apoB48 secretion was reduced only slightly in using hepatocytes from a of the secretion of apoB100 and apoB48 was The the from of of particularly apoB48 been in the of with that apoB100 was the and from in the presence of the inhibitor particularly the by MTP lipoproteins were in the lumen of the apoB100 apoB100 that with the lumenal of the the between that with the lumenal of the and that has been into the using hepatocytes from and that the in secretion of compared with inhibition of MTP to that apoB100 into the lumen that by the We that of apoB100 requires to a the were reduced by the MTP apoB100 We to this by hepatocytes with the However, been in the of the murine hepatocytes was by of the of TG as as the of of an apoB-free pool of TG from the microsomal lumen of We that this pool of TG the of by the TG pool the the lumenal were were with and to hepatocytes were with the MTP the amount of was whereas the amount of apoB-free in the lumenal was decreased by the of of the lumenal TG that of TG was with the amount of in the lumenal pool was only of that this TG pool was isolated from of the pool of apoB-free TG from the microsomal lumen with an of that were to in the particles in the lumen of the of were in the lumen of the of a assembly has been proposed in a TG in the lumen with a apoB-containing the of and to the of MTP that the MTP inhibitor decreased by the amount of microsomal lumenal TG with This with that the MTP inhibitor reduced the amount of TG with by the of in of the MTP in particles from the and that an of MTP to TG in the lumen in a pool with of that this TG from the TG between the TG with the lumenal of the a pool been in the has that with of the of hepatocytes of the that a that as that MTP transfers TG between a that a of TG to the lumenal of the that apoB48 particles with a of in the microsomal lumen and in the also with a of TG to the of of apoB48 particles the of lipid-poor particles with TG of by MTP the of TG the lumen assembly to of TG a to the of TG that with the in this been microsomal TG has been in this TG with of the TG that a the TG in and The this has been TG secretion has been that MTP TG the lumen, as well as However, in with in MTP to transfer the of TG to a the role of MTP in assembly that MTP the of TG in the lumen in the of TG assembly with We that the decreased of by inhibition of MTP a of the decreased of TG in the lumenal TG assembly of very low density lipoproteins in the triacylglycerol the of apolipoprotein (apo) microsomal lumenal the microsomal triacylglycerol transfer protein has been in the of TG by nascent assembly and secretion of in with the a in the MTP and MTP protein and MTP transfer a in MTP has the to transfer TG and lipids, and between in vitro and has been proposed to transfer TG to nascent apoB-containing lipoproteins in vivo. This with that and MTP of assembly MTP in the and as a with a protein of the lumen that protein The MTP transfer to the in vitro the transfer of MTP that MTP transfers between a The and of and between in that MTP in the of with of the MTP transfer been that by to in and of in apoB-containing lipoproteins from the to the However, were in the MTP was in the compared with apoB100 were reduced by apoB48 were reduced by only of the that MTP TG the of apoB48 in the of was in particles that were only apoB100 secretion from cultured hepatocytes was whereas apoB48 secretion was only slightly reduced compared with that in hepatocytes that compared with very particles the and in to in the particles in the lumen of that lumenal TG as a of TG assembly with The of MTP secretion by the that of MTP in of apoB100 and as well as TG MTP has been to in as a that the of the in the of into the lumen in the transfer of the of TG to in the lumen and in the transfer of TG into the lumen assembly into The of the role of MTP been in cultured that compared with primary in the of in The of MTP has also been in and that lipoproteins in secretion was by with MTP and of the the role of MTP in assembly to the a been the of MTP in primary hepatocytes the role of MTP has been in MTP were the of MTP in hepatocytes from of as of to the of assembly in murine We the of MTP assembly and secretion in murine primary that secretion of apoB100 to inhibition of MTP and that MTP the apoB48 particles produced lipid-poor compared with those in hepatocytes with MTP We also isolated from the microsomal lumen a pool of TG that with and that inhibition of MTP the amount of TG this However, inhibition of MTP the transfer of TG from this pool to lipid-poor apoB-containing the role of MTP in the assembly of apoB-containing lipoproteins in murine that inhibition of MTP by inhibition of TG the secretion of whereas the amount of apoB48 secreted was the of apoB48 was were in the and of of apoB100 and apoB48 in the microsomal The in of apoB100 and apoB48 to inhibition of MTP was of the presence of We isolated a microsomal lumenal pool of TG that was with and showed that the of this pool was the pool of TG with the MTP inhibitor decreased by the amount of apoB-free and TG in the microsomal lumen, inhibition of MTP the transfer of TG to of MTP the of and of the the role of MTP in assembly to the to this MTP has been proposed to as a that in the and into the lumen that TG secretion by 85% the amount of apoB100 in the microsomal lumen reduced by whereas the amount of apoB48 unaffected. This with in the murine MTP been in the hepatocytes from apoB100 secretion was whereas apoB48 secretion was reduced only slightly in using hepatocytes from a of the secretion of apoB100 and apoB48 was The the from of of particularly apoB48 been in the of with that apoB100 was the and from in the presence of the inhibitor particularly the by MTP lipoproteins were in the lumen of the apoB100 apoB100 that with the lumenal of the the between that with the lumenal of the and that has been into the using hepatocytes from and that the in secretion of compared with inhibition of MTP to that apoB100 into the lumen that by the We that of apoB100 requires to a the were reduced by the MTP apoB100 We to this by hepatocytes with the However, been in the of the murine hepatocytes was by of the of TG as as the of of an apoB-free pool of TG from the microsomal lumen of We that this pool of TG the of by the TG pool the the lumenal were were with and to hepatocytes were with the MTP the amount of was whereas the amount of apoB-free in the lumenal was decreased by the of of the lumenal TG that of TG was with the amount of in the lumenal pool was only of that this TG pool was isolated from of the pool of apoB-free TG from the microsomal lumen with an of that were to in the particles in the lumen of the of were in the lumen of the of a assembly has been proposed in a TG in the lumen with a apoB-containing the of and to the of MTP that the MTP inhibitor decreased by the amount of microsomal lumenal TG with This with that the MTP inhibitor reduced the amount of TG with by the of in of the MTP in particles from the and that an of MTP to TG in the lumen in a pool with of that this TG from the TG between the TG with the lumenal of the a pool been in the has that with of the of hepatocytes of the that a that as that MTP transfers TG between a that a of TG to the lumenal of the that apoB48 particles with a of in the microsomal lumen and in the also with a of TG to the of of apoB48 particles the of lipid-poor particles with TG of by MTP the of TG the lumen assembly to of TG a to the of TG that with the in this been microsomal TG has been in this TG with of the TG that a the TG in and The this has been TG secretion has been that MTP TG the lumen, as well as However, in with in MTP to transfer the of TG to a the role of MTP in assembly that MTP the of TG in the lumen in the of TG assembly with We that the decreased of by inhibition of MTP a of the decreased of TG in the lumenal TG We the role of MTP in the assembly of apoB-containing lipoproteins in murine that inhibition of MTP by inhibition of TG the secretion of whereas the amount of apoB48 secreted was the of apoB48 was were in the and of of apoB100 and apoB48 in the microsomal The in of apoB100 and apoB48 to inhibition of MTP was of the presence of We isolated a microsomal lumenal pool of TG that was with and showed that the of this pool was the pool of TG with the MTP inhibitor decreased by the amount of apoB-free and TG in the microsomal lumen, inhibition of MTP the transfer of TG to of MTP the of and of the the role of MTP in assembly to the to this MTP has been proposed to as a that in the and into the lumen that TG secretion by 85% the amount of apoB100 in the microsomal lumen reduced by whereas the amount of apoB48 unaffected. This with in the murine MTP been in the hepatocytes from apoB100 secretion was whereas apoB48 secretion was reduced only slightly in using hepatocytes from a of the secretion of apoB100 and apoB48 was The the from of of particularly apoB48 been in the of with that apoB100 was the and from in the presence of the inhibitor particularly the by MTP lipoproteins were in the lumen of the apoB100 apoB100 that with the lumenal of the the between that with the lumenal of the and that has been into the using hepatocytes from and that the in secretion of compared with inhibition of MTP to that apoB100 into the lumen that by the We that of apoB100 requires to a the were reduced by the MTP apoB100 We to this by hepatocytes with the However, been in the of the murine hepatocytes was by of the of TG as as the of of an apoB-free pool of TG from the microsomal lumen of We that this pool of TG the of by the TG pool the the lumenal were were with and to hepatocytes were with the MTP the amount of was whereas the amount of apoB-free in the lumenal was decreased by the of of the lumenal TG that of TG was with the amount of in the lumenal pool was only of that this TG pool was isolated from of the pool of apoB-free TG from the microsomal lumen with an of that were to in the particles in the lumen of the of were in the lumen of the of a assembly has been proposed in a TG in the lumen with a apoB-containing the of and to the of MTP that the MTP inhibitor decreased by the amount of microsomal lumenal TG with This with that the MTP inhibitor reduced the amount of TG with by the of in of the MTP in particles from the and that an of MTP to TG in the lumen in a pool with of that this TG from the TG between the TG with the lumenal of the a pool been in the has that with of the of hepatocytes of the that a that as that MTP transfers TG between a that a of TG to the lumenal of the that apoB48 particles with a of in the microsomal lumen and in the also with a of TG to the of of apoB48 particles the of lipid-poor particles with TG of by MTP the of TG the lumen assembly to of TG a to the of TG that with the in this been microsomal TG has been in this TG with of the TG that a the TG in and The this has been TG secretion has been that MTP TG the lumen, as well as However, in with in MTP to transfer the of TG to a the role of MTP in assembly that MTP the of TG in the lumen in the of TG assembly with We that the decreased of by inhibition of MTP a of the decreased of TG in the lumenal TG of MTP the of and of the the role of MTP in assembly to the to this MTP has been proposed to as a that in the and into the lumen that TG secretion by 85% the amount of apoB100 in the microsomal lumen reduced by whereas the amount of apoB48 unaffected. This with in the murine MTP been in the hepatocytes from apoB100 secretion was whereas apoB48 secretion was reduced only slightly in using hepatocytes from a of the secretion of apoB100 and apoB48 was The the from of of particularly apoB48 been in the of with that apoB100 was the and from in the presence of the inhibitor particularly the by MTP lipoproteins were in the lumen of the apoB100 apoB100 that with the lumenal of the the between that with the lumenal of the and that has been into the using hepatocytes from and that the in secretion of compared with inhibition of MTP to that apoB100 into the lumen that by the We that of apoB100 requires to a the were reduced by the MTP apoB100 We to this by hepatocytes with the However, been in the of the murine hepatocytes was by of the of TG as as the of of an apoB-free pool of TG from the microsomal lumen of We that this pool of TG the of by the TG pool the the lumenal were were with and to hepatocytes were with the MTP the amount of was whereas the amount of apoB-free in the lumenal was decreased by the of of the lumenal TG that of TG was with the amount of in the lumenal pool was only of that this TG pool was isolated from of the pool of apoB-free TG from the microsomal lumen with an of that were to in the particles in the lumen of the of were in the lumen of the of a assembly has been proposed in a TG in the lumen with a apoB-containing the of and to the of MTP that the MTP inhibitor decreased by the amount of microsomal lumenal TG with This with that the MTP inhibitor reduced the amount of TG with by the of in of the MTP in particles from the and that an of MTP to TG in the lumen in a pool with of that this TG from the TG between the TG with the lumenal of the a pool been in the has that with of the of hepatocytes of the that a that as that MTP transfers TG between a that a of TG to the lumenal of the that apoB48 particles with a of in the microsomal lumen and in the also with a of TG to the of of apoB48 particles the of lipid-poor particles with TG of by MTP the of TG the lumen assembly to of TG a to the of TG that with the in this been microsomal TG has been in this TG with of the TG that a the TG in and The this has been TG secretion has been that MTP TG the lumen, as well as However, in with in MTP to transfer the of TG to a the role of MTP in assembly that MTP the of TG in the lumen in the of TG assembly with We that the decreased of by inhibition of MTP a of the decreased of TG in the lumenal TG of MTP the of and of the the role of MTP in assembly to the to this MTP has been proposed to as a that in the and into the lumen that TG secretion by 85% the amount of apoB100 in the microsomal lumen reduced by whereas the amount of apoB48 unaffected. This with in the murine MTP been in the hepatocytes from apoB100 secretion was whereas apoB48 secretion was reduced only slightly in using hepatocytes from a of the secretion of apoB100 and apoB48 was The the from of of particularly apoB48 been in the of with that apoB100 was the and from in the presence of the inhibitor particularly the by MTP lipoproteins were in the lumen of the apoB100 apoB100 that with the lumenal of the the between that with the lumenal of the and that has been into the using hepatocytes from and that the in secretion of compared with inhibition of MTP to that apoB100 into the lumen that by the We that of apoB100 requires to a the were reduced by the MTP apoB100 We to this by hepatocytes with the However, been in the of the murine hepatocytes was by of the of the the role of MTP in assembly to the to this MTP has been proposed to as a that in the and into the lumen that TG secretion by 85% the amount of apoB100 in the microsomal lumen reduced by whereas the amount of apoB48 unaffected. This with in the murine MTP been in the hepatocytes from apoB100 secretion was whereas apoB48 secretion was reduced only slightly in using hepatocytes from a of the secretion of apoB100 and apoB48 was The the from of of particularly apoB48 been in the of with The that apoB100 was the and from in the presence of the inhibitor particularly the by MTP lipoproteins were in the lumen of the apoB100 apoB100 that with the lumenal of the the between that with the lumenal of the and that has been into the using hepatocytes from and that the in secretion of compared with inhibition of MTP to that apoB100 into the lumen that by the We that of apoB100 requires to a the were reduced by the MTP apoB100 We to this by hepatocytes with the However, been in the of the murine hepatocytes was by of the MTP of TG as as the of of an apoB-free pool of TG from the microsomal lumen of We that this pool of TG the of by the TG pool the the lumenal were were with and to hepatocytes were with the MTP the amount of was whereas the amount of apoB-free in the lumenal was decreased by the of of the lumenal TG that of TG was with the amount of in the lumenal pool was only of that this TG pool was isolated from of the pool of apoB-free TG from the microsomal lumen with an of that were to in the particles in the lumen of the of were in the lumen of the of a assembly has been proposed in a TG in the lumen with a apoB-containing the of and to the of MTP that the MTP inhibitor decreased by the amount of microsomal lumenal TG with This with that the MTP inhibitor reduced the amount of TG with by the of in of the MTP in particles from the and that an of MTP to TG in the lumen in a pool with of that this TG from the TG between the TG with the lumenal of the a pool been in the has that with of the of hepatocytes of the that a that as that MTP transfers TG between a that a of TG to the lumenal of the that apoB48 particles with a of in the microsomal lumen and in the also with a of TG to the of of apoB48 particles the of lipid-poor particles with TG of by MTP the of TG the lumen assembly to of TG a to the of TG that with the in this been microsomal TG has been in this TG with of the TG that a the TG in and The this has been TG secretion has been that MTP TG the lumen, as well as However, in with in MTP to transfer the of TG to a the role of MTP in assembly that MTP the of TG in the lumen in the of TG assembly with We that the decreased of by inhibition of MTP a of the decreased of TG in the lumenal TG the of of an apoB-free pool of TG from the microsomal lumen of We that this pool of TG the of by the TG pool the the lumenal were were with and to hepatocytes were with the MTP the amount of was whereas the amount of apoB-free in the lumenal was decreased by the of of the lumenal TG that of TG was with the amount of in the lumenal pool was only of that this TG pool was isolated from of the pool of apoB-free TG from the microsomal lumen with an of that were to in the particles in the lumen of the of were in the lumen of the of a assembly has been proposed in a TG in the lumen with a apoB-containing the of and to the of MTP that the MTP inhibitor decreased by the amount of microsomal lumenal TG with This with that the MTP inhibitor reduced the amount of TG with by the of in of the MTP in particles from the and that an of MTP to TG in the lumen in a pool with of that this TG from the TG between the TG with the lumenal of the a pool been in the has that with of the of hepatocytes of the that a that as that MTP transfers TG between a that a of TG to the lumenal of the that apoB48 particles with a of in the microsomal lumen and in the also with a of TG to the of of apoB48 particles the of lipid-poor particles with TG of The by MTP the of TG the lumen assembly to of TG a to the of TG that with the in this been microsomal TG has been in this TG with of the TG that a the TG in and The this has been TG secretion has been that MTP TG the lumen, as well as However, in with in MTP to transfer the of TG to a the role of MTP in assembly that MTP the of TG in the lumen in the of TG assembly with We that the decreased of by inhibition of MTP a of the decreased of TG in the lumenal TG We and the

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