Overexpression of apolipoprotein A-IV enhances lipid secretion in IPEC-1 cells by increasing chylomicron size.
Intestinal apolipoprotein A-IV expression is highly regulated by dietary lipid in newborn swine, suggesting a role in lipid absorption. Constitutive overexpression of apoA-IV in newborn swine enterocytes enhances basolateral secretion of triacylglycerol (TG) in TG-rich lipoproteins 4.9-fold (Lu, S., Yao, Y., Meng, S., Cheng, X., and Black, D. D. (2002) J. Biol. Chem. 277, 31929-31937). To investigate the mechanism of this enhancement, IPEC-1 cells were transfected with a tetracycline-regulatable expression system (Tet-On). In cells incubated with oleic acid, a dose response relationship was observed between medium doxycycline concentration and basolateral apoA-IV and TG secretion. Similarly regulated expression of apoA-I did not enhance lipid secretion. The mean diameter of TG-rich lipoproteins secreted from doxycycline-treated cells was larger than from untreated cells (87.0 nm versus 53.4 nm). Basolateral apoB secretion decreased. Using the same expression system, full-length human apoA-IV (376 amino acids); a "pig-like" human apoA-IV, lacking the C-terminal EQQQ repeats (361 amino acids); and a "chicken-like" apoA-IV, further truncated to 343 amino acids, were expressed in IPEC-1 cells. With increasing protein secretion, cells expressing the full-length human apoA-IV displayed a 2-fold increase in TG secretion; in sharp contrast, cells expressing the pig-like human apoA-IV displayed a 25-fold increase in TG secretion and a 27-fold increase in lipoprotein diameter. When human apoA-IV was further truncated to yield a chicken-like protein, TG secretion was inhibited. We conclude that overexpression of swine apoA-IV enhances basolateral TG secretion in a dose-dependent manner by increasing the size of secreted lipoproteins. These data suggest that the region in the human apoA-IV protein from residues 344 to 354 is critical to its ability to enhance lipid secretion, perhaps by enabling the packaging of additional core TG into chylomicron particles. The EQQQ-rich region may play an inhibitory or modulatory role in chylomicron packaging in humans.
Lu
Song
S
Children's Foundation Research Center at Le Bonheur Children's Medical Center and Department of Pediatrics, University of Tennessee Health Science Center, Memphis, Tennessee 38103, USA.
Yao
Ying
Y
Cheng
Xiangying
X
Mitchell
Sonya
S
Leng
Shuangying
S
Meng
Songmei
S
Gallagher
James W
JW
Shelness
Gregory S
GS
Morris
Gabriel S
GS
Mahan
James
J
Frase
Sharon
S
Mansbach
Charles M
CM
Weinberg
Richard B
RB
Black
Dennis D
DD
D000595:Amino Acid Sequence; D000818:Animals; D000831:Animals, Newborn; D001053:Apolipoproteins / Q000737:chemistry; D001054:Apolipoproteins A / Q000096:biosynthesis* / Q000502:physiology; D015153:Blotting, Western; D002460:Cell Line; D002645:Chickens; D002914:Chylomicrons / Q000737:chemistry*; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004318:Doxycycline / Q000378:metabolism / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D047468:Immunoprecipitation; D007413:Intestinal Mucosa / Q000378:metabolism; D007422:Intestines / Q000166:cytology*; D050356:Lipid Metabolism; D008055:Lipids / Q000737:chemistry*; D008074:Lipoproteins / Q000378:metabolism; D008854:Microscopy, Electron; D046529:Microscopy, Electron, Transmission; D008969:Molecular Sequence Data; D009154:Mutation; D019301:Oleic Acid / Q000737:chemistry / Q000378:metabolism; D017434:Protein Structure, Tertiary; D012313:RNA / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013552:Swine; D013752:Tetracycline / Q000494:pharmacology; D015533:Transcriptional Activation; D014280:Triglycerides / Q000378:metabolism
D001053:Apolipoproteins; D001054:Apolipoproteins A; D002914:Chylomicrons; D018076:DNA, Complementary; D008055:Lipids; D008074:Lipoproteins; D012333:RNA, Messenger; D014280:Triglycerides; C022205:apolipoprotein A-IV; D019301:Oleic Acid; D012313:RNA; D013752:Tetracycline; D004318:Doxycycline