PubMed Health⌕ Search

Biomedical subjects

M Busslinger

Publications and source records attributed to M Busslinger.

At least 91 records · Page 5Linked to original sources
PubMed1987

Mutually exclusive interaction of the CCAAT-binding factor and of a displacement protein with overlapping sequences of a histone gene promoter.

The sperm histone H2B-1 gene of the sea urchin Psammechinus miliaris contains two octamer sequences (ATTTGCAT) and two CCAAT motifs upstream of its TATA box. The CCAAT-binding factors present in nuclear extracts from testis and from blastula and gastrula embryos are indistinguishable by mobility shift and methylation interference analysis. However, there is a testis-specific octamer-binding factor in addition to the ubiquitous form. In DNAase I protection experiments, the CCAAT-binding factor of only the testis extract is able to interact with the sperm H2B promoter. In the two embryonic extracts a novel factor binds with high affinity to sequences overlapping the proximal CCAAT element, thus preventing the DNA interaction of the CCAAT-binding factor in the embryo where the sperm H2B gene is not expressed. This CCAAT displacement protein may therefore act as a repressor of sperm H2B gene transcription.

Base Sequence↗
PubMed1986

Characterization of two nonallelic pairs of late histone H2A and H2B genes of the sea urchin: differential regulation in the embryo and tissue-specific expression in the adult.

Two nonallelic pairs of late H2A and H2B genes of the sea urchin Psammechinus miliaris were isolated on two different cosmid clones. The genes of cosmid PmL1 are separated by 11 kilobases of DNA and code for the late H2A-2 and H2B-2 variants. The genes of clone PmL2 are divergently transcribed with 1,060 base pairs of intergenic spacer DNA and code for novel variants of the H2A-2 and H2B-2 type. A comparison of the promoter sequences revealed little homology upstream of the TATA box with the exception of a 24-base-pair-long conserved sequence which is present at the same position in both late H2B promoters and part of which is identical with the "H2B-specific" 5' element. The mRNAs of the H2A and H2B genes of cosmid PmL1 reach their maximal levels early in the mesenchyme blastula embryo, whereas the transcripts of both genes of clone PmL2 accumulate maximally only later in the pluteus larva. In the adult sea urchin all four mRNAs are present in the tube foot but not in the intestine and lantern muscle. This pattern of differential expression in the embryo and tissue-specific expression in the adult suggests cell lineage-specific regulation of the late H2A-2 and H2B-2 genes. Another class of late histone genes represented by the H2A-3 and H2B-1 genes was shown to be expressed in all three adult tissues tested, whereas transcripts of the late H2A-1 genes could not be detected, suggesting that these genes are active exclusively during sea urchin development.

Alleles↗
PubMed1985

Synthesis of sperm and late histone cDNAs of the sea urchin with a primer complementary to the conserved 3' terminal palindrome: evidence for tissue-specific and more general histone gene variants.

We have cloned histone cDNAs from total RNA isolated from testis and from gastrula-stage embryos of the sea urchin Psammechinus miliaris. The reverse transcription of histone mRNAs was specifically primed with an oligonucleotide that is complementary to the conserved palindromic sequence present at the 3' end of nonpolyadenylated histone mRNAs. Two sperm H2B, two late H2B, and three late H2A variant cDNA clones were isolated and characterized by DNA sequence analysis. These cDNA clones were used to study the accumulation of histone mRNA during sea urchin embryogenesis. The different late H2A and H2B mRNAs are present in as few as 200 copies in the egg and each accumulate to 3-5 X 10(5) molecules in the gastrula embryo. One of the late mRNAs, the H2A-3 mRNA, is also abundant in testis RNA and codes for the H2A variant present in sperm chromatin. The late H2A-3 protein is therefore a more prevalent H2A variant of the sea urchin. In contrast, the two sperm H2B mRNAs are found in testes but not ovaries and embryos of the sea urchin, suggesting that the sperm H2B genes are expressed only during spermatogenesis. In addition, evidence for gene conversion between two late H2A gene variants is presented.

Animals↗
PubMed1984

The cDNA sequences of the sea urchin U7 small nuclear RNA suggest specific contacts between histone mRNA precursor and U7 RNA during RNA processing.

3' Processing of sea urchin H3 histone pre-mRNA depends on a small nuclear RNP which contains an RNA of nominally 60 nucleotide length, referred to below as U7 RNA. The U7 RNA can be enriched by precipitation of sea urchin U-snRNPs with human systematic lupus erythematosus antiserum of the Sm serotype. We have prepared cDNA clones of U7 RNA and determined by hybridization techniques that this RNA is present in sea urchin eggs at 30-fold lower molar concentration than U1 RNA. The RNA sequences derived from an analysis of eight U7 cDNA clones show neither homologies nor complementarities to any other know U-RNAs. The 3' portion of the presumptive RNA sequence can be folded into a stem-loop structure. The 5'-terminal sequences would be largely unstructured as free RNA. Their most striking feature is their base complementarity to the 3' conserved sequences of histone pre-mRNAs. Six out of nine bases of the conserved CAAGAAAGA sequence of the histone mRNA precursor and 13 out of 16 nucleotides from the conserved palindrome can be base paired with presumptive U7 RNA sequence, suggesting a unique hybrid structure for a processing intermediate formed from histone precursor and U7 RNA.

Animals↗
PubMed1983

The sequence GGCmCGG is resistant to MspI cleavage.

MspI essentially fails to cut the sequence GGCmCGG at enzyme concentrations which give total digestion of CCGG, CmCGG and GGCCGG sites. This result explains why certain sites in mammalian DNA are resistant to both MspI and HpaII and shows that this results from an idiosynchracy of MspI rather than a novel form of DNA methylation at this site in mammalian cells.

Animals↗
PubMed1983

DNA methylation and the regulation of globin gene expression.

We have studied the effect of DNA methylation on human gamma-globin gene expression, using a novel in vitro DNA methylation technique. With this method we have methylated specific segments of the gamma-globin gene and its 5'-flanking region, which were cloned for this purpose in the bacteriophage vector M13mp8. The vitro methylated DNA was introduced into mouse L-cells by cotransfer using the herpes simplex virus thymidine kinase gene as a selective marker. Transformed cell lines were isolated to examine the inheritance of the segmental methylation pattern and its effect on the expression of the gamma-globin gene. Cells transformed with DNA methylated throughout the M13 vector and gamma-globin DNA sequences do not express the gamma-globin gene. Methyl-C residues in either the M13 DNA sequences or the gamma-globin structural gene have, however, no inhibitory effect on globin transcription. In contrast, methylation in the 5' region of the gamma-globin gene (from nucleotides -760 to +100) prevents transcription. These data indicate that DNA methylation in the 5' region of a gene might play a direct role in the regulation of gene expression.

5-Methylcytosine↗
PubMed1982

An unusual evolutionary behaviour of a sea urchin histone gene cluster.

DNA sequences of cloned histone coding sequences and spacers of sea urchin species that diverged long ago in evolution were compared. The highly repeated H4 and H3 genes active during early embryogenesis had evolved (in their silent sites) at a rate (0.5-0.6% base changes/Myr) similar to single-copy protein-coding genes and nearly as fast as spacer DNA (0.7% base changes/Myr) and unique DNA. Thus, evolution in the major histone genes conforms to a universal evolutionary clock based on the rate of base sequence change. By contrast, the H4 and H3 coding sequences and a non-transcribed spacer of the DNA clone h19 of Psammechinus miliaris show an exceptionally low rate of sequence evolution only 1/100 to 1/200 that predicted from the clock hypothesis. According to the classical model of gene inheritance, the h19 DNA sequences in the Psammechinus genome require unusual conservation mechanisms by selection at the level of the gene and spacer sequences. An alternative explanation could be recent horizontal gene transfer of a histone gene cluster from the very distantly related Strongylocentrotus dröbachiensis to the P. miliaris genome.

Journal Article↗
PubMed1981

Beta + thalassemia: aberrant splicing results from a single point mutation in an intron.

We have analyzed the molecular basis of beta + thalassemia by studying the expression of a cloned beta-globin gene in HeLa cells. This beta-globin gene was isolated from a beta + thalassemic patient and differs from the normal beta-globin genes by only a single point mutation within the first intron. The beta + thalassemic and the normal beta-globin genes were cloned into an SV40-pBR328 vector and introduced into HeLa cells by calcium phosphate coprecipitation. We assayed the RNA from these transfected HeLa cells by S1 nuclease mapping and cDNA sequencing to detect the nature of the defect in beta-globin gene expression. While the transcripts of the normal beta-globin gene are processed correctly, the first intron of the beta + thalassemic beta-globin gene is incorrectly spliced in about 90% of the mRNA because of an additional 3; splice site that has been created by the point mutation. This incorrectly spliced mRNA is effectively exported to the cytoplasm, where it would conceivably be translated to give a truncated globin chain of 35 amino acids. The remaining 10% of the mRNA transcribed from the beta+ thalassemic globin gene is correctly spliced and can therefore be translated to give normal beta-globin. In addition to the incorrect splicing of the first intron, the splicing of both introns is retarded, which results in the accumulation of unspliced pre-mRNA. This suggests that removal of the first intron might facilitate splicing of the second intron.

Base Sequence↗
PubMed1980

Ubiquitous and gene-specific regulatory 5' sequences in a sea urchin histone DNA clone coding for histone protein variants.

The DNA sequences of the entire structural H4, H3, H2A and H2B genes and of their 5' flanking regions have been determined in the histone DNA clone h19 of the sea urchin Psammechinus miliaris. In clone h19 the polarity of transcription and the relative arrangement of the histone genes is identical to that in clone h22 of the same species. The histone proteins encoded by h19 DNA differ in their primary structure from those encoded by clone h22 and have been compared to histone protein sequences of other sea urchin species as well as other eukaryotes. A comparative analysis of the 5' flanking DNA sequences of the structural histone genes in both clones revealed four ubiquitous sequence motifs; a pentameric element GATCC, followed at short distance by the Hogness box GTATAAATAG, a conserved sequence PyCATTCPu, in or near which the 5' ends of the mRNAs map in h22 DNA and lastly a sequence A, containing the initiation codon. These sequences are also found, sometimes in modified version, in front of other eukaryotic genes transcribed by polymerase II. When prelude sequences of isocoding histone genes in clone h19 and h22 are compared areas of homology are seen to extend beyond the ubiquitous sequence motifs towards the divergent AT-rich spacer and terminate between approximately 140 and 240 nucleotides away from the structural gene. These prelude regions contain quite large conservative sequence blocks which are specific for each type of histone genes.

Animals↗
PubMed1979

A regulatory sequence near the 3' end of sea urchin histone genes.

The 3' flanking sequences of all five histone genes have been sequenced in the histone DNA clone h19 of the sea urchin Psammechinus miliaris. A large (23 bp) and a small (10 bp) conserved sequence was found by sequence comparison, some 29-40 bp downstream from the termination codon. 12 bases of the larger homology block show a dyad symmetry. The available sequences of clone h22 of the same species and those of the histone clones pSp2 and pSp17 of Strongylocentrotus purpuratus, another sea urchin species, fit well into this comparison. Two types of sequences are involved in the dyad symmetry; one is H1, H3 and H4 specific, the other is H2A and H2B specific. If these conserved sequences are transcribed, a hairpin loop could form in the RNA molecules. This secondary structure might serve as a recognition signal for a regulatory protein.

Animals↗
PubMed1978

Aspects of the regulation of histone genes.

Sequencing of cloned histone DNA of the sea urchin Psammechinus miliaris has confirmed the map of the histone genes obtained earlier by rather less refined techniques. Sequencing of spacer has revealed that it is unlikely to code for protein. Some interesting sequences in the prelude regions to the structural genes have been found. The technique of injecting DNA into the germinal vesicle of the Xenopus oocyte has been greatly simplified, so that now many of the parameters governing the transcription of the injected genes can be investigated. Some mRNA-like molecules appear when circular histone DNA is inserted into the oocyte nucleus. We are cautiously optimistic that the technique can be further developed and will provide a useful tool for the study of the molecular mechanisms governing the expression of structural genes coding for proteins.

Animals↗