Long-range disruption of gene expression by a selectable marker cassette.
Recent studies have suggested that the retention of selectable marker cassettes (like PGK-Neo, in which a hybrid gene consisting of the phosphoglycerate kinase I promoter drives the neomycin phosphotransferase gene) in targeted loci can cause unexpected phenotypes in "knockout" mice due to disruption of expression of neighboring genes within a locus. We have studied targeted mutations in two multigene clusters, the granzyme B locus and the beta-like globin gene cluster. The insertion of PGK-Neo into the granzyme B gene, the most 5' gene in the granzyme B gene cluster, severely reduced the normal expression of multiple genes within the locus, even at distances greater than 100 kb from the mutation. Similarly, the insertion of a PGK-Neo cassette into the beta-globin locus control region (LCR) abrogates the expression of multiple globin genes downstream from the cassette. In contrast, a targeted mutation of the promyelocyte-specific cathepsin G gene (which lies just 3' to the granzyme genes in the same cluster) had minimal effects on upstream granzyme gene expression. Although the mechanism of these-long distance effects are unknown, the expression of PGK-Neo can be "captured" by the regulatory domain into which it is inserted. These results suggest that the PGK-Neo cassette can interact productively with locus control regions and thereby disrupt normal interactions between local and long-distance regulatory regions within a tissue-specific domain.
Pham
C T
CT
Washington University School of Medicine, Department of Medicine, St. Louis, MO 63110-1093, USA.
MacIvor
D M
DM
Hug
B A
BA
Heusel
J W
JW
Ley
T J
TJ
D000818:Animals; D056649:Cathepsin G; D002403:Cathepsins / Q000235:genetics; D004251:DNA Transposable Elements; D015870:Gene Expression; D015723:Gene Library; D005819:Genetic Markers; D005914:Globins / Q000235:genetics*; D053804:Granzymes; D019868:Kanamycin Kinase; D051379:Mice; D018345:Mice, Knockout; D008969:Molecular Sequence Data; D005810:Multigene Family; D016254:Mutagenesis, Insertional; D010735:Phosphoglycerate Kinase / Q000096:biosynthesis / Q000235:genetics; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000096:biosynthesis; D016133:Polymerase Chain Reaction; D011401:Promoter Regions, Genetic; D012045:Regulatory Sequences, Nucleic Acid; D012697:Serine Endopeptidases / Q000235:genetics*
D004251:DNA Transposable Elements; D005819:Genetic Markers; D005914:Globins; D017853:Phosphotransferases (Alcohol Group Acceptor); D019868:Kanamycin Kinase; D010735:Phosphoglycerate Kinase; D002403:Cathepsins; D053804:Granzymes; C509214:Gzmb protein, mouse; D012697:Serine Endopeptidases; D056649:Cathepsin G; C534096:Ctsg protein, mouse