Restricted β-galactosidase expression of a hygromycin-lacZ gene targeted to the β-actin locus and embryonic lethality of β-actin mutant mice

β-actin is a cytoskeletal protein that is ubiquitously expressed. To exploit the regulation the β-actin gene, a promoterless hygromycin-lacZ fusion gene with a splice acceptor was introduced into the first intron of the β-actin locus by homologous recombination in mouse embryonic stem (ES) cells. The targeted ES cells were hygromycin resistant and expressed β-galactosidase (β-gal) activity. However, no β-gal activity was detected in heterozygous embryos. In adult heterozygotes, β-gal activity was detected only in testes. RT-PCR analysis demonstrated the presence of both β-actin exon 1-hygromycin- and exon 1-exon 2-containing transcripts in homozygous mutant embryos. LacZ-containing transcripts were detected in adult heterozygous testes and, surprisingly, in homozygous mutant embryos. These results demonstrate that the integration of the hygromycin-lacZ gene into the first intron of the β-actin locus was not productive for the ubiquitous expression of β-gal activity. Because this integration mimics certain types of gene trap events, it suggests that caution should be used when interpreting β-gal expression patterns in genetic screens using gene trap strategies. In addition, mice homozygous for the β-actin mutation developed normally up to embryonic day 8.5 (E8.5) but became growth retarded at E9.5 and subsequently died. The RT-PCR data indicate that this targeted mutation is a hypomorphic allele of β-actin.