http://repub.eur.nl/res/aut/596/ List of Publications en http://repub.eur.nl/static-eur/img/logo.png http://repub.eur.nl http://repub.eur.nl/res/pub/8801/ 1998-01-01T00:00:00Z Murine ZFP-37 is a member of the large family of C2H2 type zinc finger proteins. It is characterized by a truncated NH2-terminal Kruppel-associated box and is thought to play a role in transcriptional regulation. During development Zfp-37 mRNA is most abundant in the developing central nervous system, and in the adult mouse expression is restricted largely to testis and brain. Here we show that at the protein level ZFP-37 is detected readily in neurons of the adult central nervous system but hardly in testis. In brain ZFP-37 is associated with nucleoli and appears to contact heterochromatin. Mouse and human ZFP-37 have a basic histone H1-like linker domain, located between KRAB and zinc finger regions, which binds double-stranded DNA. Thus we suggest that ZFP-37 is a structural protein of the neuronal nucleus which plays a role in the maintenance of specialized chromatin domains. http://repub.eur.nl/res/pub/2497/ 1996-01-01T00:00:00Z Single-copy human beta-globin transgenes are very susceptible to suppression by position effects of surrounding closed chromatin. However, these position effects are overcome by a 20 kbp DNA fragment containing the locus control region (LCR). Here we show that the 6.5 kbp microlocus LCR cassette reproducibly directs full expression from independent single-copy beta-globin transgenes. By testing individual DNase I-hypersensitive sites (HS) present in the microlocus cassette, we demonstrate that the 1.5 kbp 5'HS2 enhancer fragment does not direct beta-globin expression from single-copy transgenes. In contrast, the 1.9 kbp 5'HS3 fragment directs beta-globin expression in five independent single-copy transgenic mouse lines. Moreover, the 5'HS3 core element and beta-globin proximal promoter sequences are DNase I hypersensitive in fetal liver nuclei of these expressing transgenic lines. Taken together, these results demonstrate that LCR activity is the culmination of at least two separable functions including: (i) a novel activity located in 5'HS3 that dominantly opens and remodels chromatin structure; and (ii) a recessive enhancer activity residing in 5'HS2. We postulate that the different elements of the LCR form a 'holocomplex' that interacts with the individual globin genes. http://repub.eur.nl/res/pub/2506/ 1996-01-01T00:00:00Z The murine Zfp-37 gene encodes a protein with 12 zinc fingers at its C-terminus (Nelki et al., 1990, Nucleic Acids Res. 18: 3655; Burke and Wolgemuth, 1992, Nucleic Acids Res. 20: 2827-2834). Contrary to the published data, our Northern blot analysis demonstrates not only that the Zfp-37 gene is expressed as 2.3, 2.6, and 4.2 kb mRNAs in testis, but also that there is a 3.7-kb message in the adult mouse brain. Using a partial cDNA as a probe, we have isolated a brain-specific Zfp-37 cDNA clone of 3.3 kb, whose sequence was extended to full length using 5' end RACE. This revealed that the 3.7-kb mRNA is in fact a collection of transcripts with heterogenous 5' ends. Comparison of cDNA and genomic sequences shows that the Zfp-37 gene is spread over a region of approximately 20 kb and consists of six exons, the large 3' end exon containing the complete zinc finger domain, and 3' UTR. Our data show that the Zfp-37 gene utilizes different promoters, alternative splicing, and differential polyadenylation to generate the distinct transcripts of brain and testis. Several protein isoforms are encoded by these mRNAs, some of which contain a truncated form of a conserved domain (Kruppel-associated box) found in other zinc finger genes. In situ hybridization analysis of postnatal brain sections indicates that the Zfp-37 gene is expressed in all neurons of the central nervous system. Together, these results suggest that ZFP-37 is a transcriptional regulator predominantly present in postmitotic cells from two different lineages. http://repub.eur.nl/res/pub/2504/ 1995-01-01T00:00:00Z Erythroid Kruppel-like factor (EKLF) was originally isolated from erythroid cell RNA by differential screening and shown to be erythroid-specific, although a low level of EKLF was found in mast cell lines. EKLF contains three zinc-fingers homologous to those found in the Kruppel family of transcription factors. Because it binds the sequence CCACACCCT, EKLF may affect erythroid development as a result of its ability to bind to the CAC box in the promoter of the beta-globin gene. Mutation of this element leads to reduced beta-globin expression and it appears to mediate the effect of the globin locus control region on the promoter. Here we inactivate the EKLF gene through insertion of a lacZ reporter gene by homologous recombination in embryonic stem (ES) cells. Heterozygous EKLF+/- mice show that the reporter gene is expressed in a developmentally specific manner in all types of erythroblasts in the fetal liver and adult bone marrow. Homozygous EKLF-/- mice appear normal during the embryonic stage of haematopoiesis in the yolk sac, but develop a fatal anaemia during early fetal life when haematopoiesis has switched to the fetal liver. Enucleated erythrocytes are formed but these do not contain the proper amount of haemoglobin. We conclude that the transcription factor EKLF is essential for the final steps of definitive erythropoiesis in fetal liver.