Statements in which the resource exists as a subject.
PredicateObject
rdf:type
lifeskim:mentions
pubmed:issue
34
pubmed:dateCreated
2000-9-25
pubmed:abstractText
The role of ADP-ribosylation factor 1 (ARF-1) in the assembly of very low density lipoproteins (VLDL) was investigated by expressing dominant-negative mutants in McA-RH7777 cells. Transient expression of ARF-1(T31N), a GDP-restrictive mutant, significantly inhibited apolipoprotein B-100 (apoB-100) VLDL production without influencing the biosynthesis of apoB-100 low density lipoproteins or total apoB production (indicating that it inhibited the second step of VLDL assembly) and without altering total protein production or biosynthesis of transferrin, phosphatidylcholine, or triglycerides. These effects were confirmed in stable inducible transfectants. In contrast, expression of an ARF-1 mutant lacking the N-terminal 17 amino acids, which has no myristoylation site and cannot interact with the microsomal membrane, did not affect VLDL assembly. Thus, active ARF-1 is needed for the second step of the process. To further explore these observations, we developed a cell-free system based on the postnuclear supernatant isolated from McA-RH7777 cells. In this system, 10-15% of the apoB-100 pool was converted to VLDL in a time- and temperature-dependent way. The assembly process was highly dependent on a heat-stable factor in the d > 1.21 g/ml infranatant of fetal calf serum; this factor was not present in low density lipoproteins or VLDL. Brefeldin A inhibited VLDL assembly in this system, as did a synthetic peptide (corresponding to N-terminal amino acids 2-17 of ARF-1) that displaces ARF-1 from the membrane. Thus, active ARF-1 is also needed for cell-free assembly of VLDL. Guanosine 5'-3-O-(thio)triphosphate also inhibited VLDL assembly in this system, indicating that the process requires ongoing hydrolysis of GTP. 1-Butanol, which inhibits the formation of phosphatidic acid (PA) and instead gives rise to phosphatidylbutanol, inhibited VLDL assembly, whereas 2-butanol, which does not inhibit PA formation, failed to do so. Thus, phospholipase D (PLD)-catalyzed formation of PA from phosphatidylcholine is essential for VLDL assembly. In support of this conclusion, exogenous PLD prevented brefeldin A from inhibiting the assembly process. Our results indicate that ARF-1 participates in the second step of VLDL assembly through a process that involves activation of PLD and production of PA.
pubmed:language
eng
pubmed:journal
pubmed:citationSubset
IM
pubmed:chemical
http://linkedlifedata.com/resource/pubmed/chemical/1-Butanol, http://linkedlifedata.com/resource/pubmed/chemical/2-butanol, http://linkedlifedata.com/resource/pubmed/chemical/ADP-Ribosylation Factor 1, http://linkedlifedata.com/resource/pubmed/chemical/Apolipoprotein B-100, http://linkedlifedata.com/resource/pubmed/chemical/Apolipoproteins B, http://linkedlifedata.com/resource/pubmed/chemical/Butanols, http://linkedlifedata.com/resource/pubmed/chemical/Guanosine 5'-O-(3-Thiotriphosphate), http://linkedlifedata.com/resource/pubmed/chemical/Guanosine Triphosphate, http://linkedlifedata.com/resource/pubmed/chemical/Lipoproteins, VLDL, http://linkedlifedata.com/resource/pubmed/chemical/Phosphatidic Acids, http://linkedlifedata.com/resource/pubmed/chemical/Phospholipase D, http://linkedlifedata.com/resource/pubmed/chemical/Transferrin
pubmed:status
MEDLINE
pubmed:month
Aug
pubmed:issn
0021-9258
pubmed:author
pubmed:issnType
Print
pubmed:day
25
pubmed:volume
275
pubmed:owner
NLM
pubmed:authorsComplete
Y
pubmed:pagination
26285-92
pubmed:dateRevised
2011-11-17
pubmed:meshHeading
pubmed:year
2000
pubmed:articleTitle
ADP-ribosylation factor 1 and its activation of phospholipase D are important for the assembly of very low density lipoproteins.
pubmed:affiliation
Department of Medical Biochemistry and the Wallenberg Laboratory for Cardiovascular Research, University of Göteborg, SE 405 30 Göteborg, Sweden.
pubmed:publicationType
Journal Article, Research Support, Non-U.S. Gov't