The molecular basis for the aberrant production of plasminogen activator inhibitor type 2 in THP-1 monocytes.
Plasminogen activator inhibitor type 2 (PAI-2) is a urokinase inhibitor that is expressed primarily in monocytes. THP-1 monocytes, however. contain a unique defect in the production of PAI-2 in that the PAI-2 transcript is truncated and the expressed protein inactive (1). Here we describe the basis of this mutation in THP-1 cells. Southern blot analysis of THP-1-derived genomic DNA indicated that there were no obvious deletions in the structure of the PAI-2 gene. However, assessment of the THP-1-derived PAI-2 transcript by RT-PCR indicated that only exons seven and eight of the normal PAI-2 mRNA could be detected. Cloning of the 5' region of the PAI-2 mRNA by 5-'RACE indicated that the PAI-2 cDNA derived from THP-1 cells is approximately 1329 bp long and contains 180 bp of sequence derived from intron 5. followed by sequences corresponding to exons seven and eight of the normal PAI-2 mRNA. The presence of the intron five fragment in endogenous THP-1 derived PAI-2 mRNA was confirmed by Northern blotting. The absence of any wild-type PAI-2 mRNA in these cells suggests that one copy of the PAI-2 allele has been deleted. The remaining allele producing the truncated mRNA appears to have undergone a translocation event and contains a mutation that has disrupted the splicing of the PAI-2 primary transcript.
Katsikis
J
J
Monash University Department of Medicine, Box Hill Hospital, Victoria, Australia.
Yu
H
H
Maurer
F
F
Medcalf
R
R
D003001:Cloning, Molecular; D005091:Exons; D006801:Humans; D007438:Introns; D009000:Monocytes / Q000378:metabolism*; D009154:Mutation; D017396:Plasminogen Activator Inhibitor 2 / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000737:chemistry / Q000235:genetics; D017423:Sequence Analysis, RNA; D014178:Translocation, Genetic; D014407:Tumor Cells, Cultured; D020298:U937 Cells
D017396:Plasminogen Activator Inhibitor 2; D012333:RNA, Messenger