A 700-bp fragment of the human antithrombin III promoter is sufficient to confer high, tissue-specific expression on human apolipoprotein A-II in transgenic mice.
The human antithrombin III-encoding gene (hAT-III) promoter (phAT-III) was used to generate transgenic mice producing a human hepatic apolipoprotein, apolipoprotein A-II (hApoA-II). Integration of the transgene into the mouse genome resulted in the efficient production of hApoA-II in plasma, reaching up to 0.40 g/l in two transgenic lines. The human ApoA-II mRNA was detected at high levels, both in the liver and in the kidney of transgenic mice. The rat AT-III gene shows the same expression pattern. In contrast, as previously described, the same promoter permitted the expression of the SV40 large T antigen only in the liver of transgenic mice. In view of the extra-hepatic distribution of the ApoA-II mRNA, a preliminary characterization of the hAT-III proximal promoter (phAT-III), driving the expression of the transgene, was realized. Using DNase I footprinting analysis with liver nuclear extracts, four protected regions (I-IV) were identified in the first 175 bp of the 5' region of hAT-III. Electrophoretic mobility shift assays performed with liver and kidney nuclear extracts indicate that region III (nucleotides (nt) -67 to -90) interacts with the liver-enriched HNF4 nuclear factor. Furthermore, our data suggest that region I (nt -123 to -138) interacts with the liver-enriched HNF3 transcription factor family, both in liver and kidney. Taken together, these results demonstrate that phAT-III is a useful tool to create transgenic mice producing high plasma levels of a human apolipoprotein due to expression of the transgene in liver and kidney.(ABSTRACT TRUNCATED AT 250 WORDS)
Tremp
G L
GL
Laboratoire de Transgenèse Animale, Rhône-Poulenc Rorer S.A., Centre de Recherche de Vitry-Alfortville, Vitry sur Seine, France.
Duchange
N
N
Branellec
D
D
Cereghini
S
S
Tailleux
A
A
Berthou
L
L
Fievet
C
C
Touchet
N
N
Schombert
B
B
Fruchart
J C
JC
D000818:Animals; D000990:Antithrombin III / Q000235:genetics*; D016633:Apolipoprotein A-II / Q000096:biosynthesis* / Q000097:blood / Q000235:genetics*; D001483:Base Sequence; D005260:Female; D005786:Gene Expression Regulation; D018390:Gene Targeting / Q000379:methods*; D006801:Humans; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008808:Mice, Inbred CBA; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D011401:Promoter Regions, Genetic / Q000235:genetics*; D012333:RNA, Messenger / Q000032:analysis; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis; D014018:Tissue Distribution; D014157:Transcription Factors / Q000378:metabolism
D016633:Apolipoprotein A-II; D012333:RNA, Messenger; D011993:Recombinant Fusion Proteins; D014157:Transcription Factors; D000990:Antithrombin III