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D Landsman

Publications and source records attributed to D Landsman.

At least 55 records · Page 3Linked to original sources
PubMed1991

Assessment of the transcriptional activation potential of the HMG chromosomal proteins.

Chromosomal proteins HMG-14, HMG-17, and HMG-1 are among the most abundant, ubiquitous, and evolutionarily conserved nonhistone proteins. Analysis of their structure reveals features which are similar to those of certain transcription factors. The distribution of charged amino acid residues along the polypeptide chains is asymmetric: positive charges are clustered toward the N-terminal region, while negative charges are clustered toward the C-terminal region. The residues in the C-terminal region have the potential to form alpha helices with negatively charged surfaces. The abilities of HMG-14, -17, and -1 to function as transcriptional activators were studied in Saccharomyces cerevisiae cells expressing LexA-HMG fusion proteins (human HMG-14 and -17 and rat HMG-1) which bind to reporter molecules containing the beta-galactosidase gene downstream from a lexA operator. Fusion constructs expressing deletion mutants of HMG-14, -17, and -1 were also tested. Analysis of binding to the lexA operator with in vitro-synthesized fusion proteins shows that there are more sites for HMG-14, -17, and -1 binding than for LexA binding and that only the fusion constructs which contain the C-terminal, acidic domains of HMG-17 bind the lexA operator specifically. None of the LexA-HMG fusion protein constructs elevate the level of beta-galactosidase activity in transfected yeast cells. Thus, although HMG-14, -17, and -1 are structurally similar to acidic transcriptional activators, these chromosomal proteins do not function as activators in this test system.

Animals↗
PubMed1990

A single copy gene for chicken chromosomal protein HMG-14b has evolutionarily conserved features, has lost one of its introns and codes for a rapidly evolving protein.

The evolutionary origins and common features of the genes coding for the HMG-14/-17 family of chromosomal proteins have been studied by isolating and sequencing the chicken HMG-14b gene, the true homolog of the human and calf HMG-14 gene. Comparison of the structure of this gene to that of the human HMG-14 gene and to the human and chicken HMG-17 genes indicates that the HMG-14 and HMG-17 genes evolved from a common ancestor. We postulate that the ancestral gene consisted of six exons. In all genes the first exon codes for the entire 5' untranslated region and for the first four amino acids, which are invariant among all the known members of the HMG-14/-17 protein family. The last exon codes for ten to 16 amino acids and for the entire 3' untranslated region, which, for each gene, constitutes over 70% of the transcript. The DNA-binding domain of the proteins is encoded by two distinct exons. The genes are characterized by 5' regions that are highly enriched in G + C residues and have features characteristic of "housekeeping" genes. The HMG-17 genes are distinct from the HMG-14 in that the 5' regulatory region of the former has two TATA boxes while the HMG-14 genes have no such regulatory element. The chicken HMG-14b gene is a single-copy gene and produces a unique transcript. In this gene, exons II and III are fused and intron 2 is missing. The fusion of the two exons produced a codon for valine in a position that, among all HMG-14/-17 proteins, is unique to HMG-14b. The possible consequences of a valine insertion at the N-terminal end of the DNA-binding domains are discussed. The HMG-14 proteins evolve significantly faster than HMG-17, suggesting that the proteins are subject to different evolutionary pressure. However, certain amino acids are conserved among all the known members of the HMG-14/-17 protein family, suggesting that they are part of the functional domain of this family of chromosomal proteins.

Amino Acid Sequence↗
PubMed1990

Mapping the human gene coding for chromosomal protein HMG-17.

The functional gene coding for nonhistone chromosomal protein HMG-17, a nucleosomal binding protein that may confer unique properties to the chromatin structure of active genes, has been mapped to band 1p36.1. The multiple, nonfunctional, HMG-17 retropseudogenes are scattered over several chromosomes.

Chromosome Mapping↗
PubMed1989

Human non-histone chromosomal protein HMG-17: identification, characterization, chromosome localization and RFLPs of a functional gene from the large multigene family.

The multigene family of chromosomal protein HMG-17 is the largest known human retropseudogene family. A functional gene was identified and isolated by screening cDNA-selected genomic clones with a set of 5 oligonucleotides whose sequence corresponded to regions in which the sequence of the retropseudogenes differed from that of the cDNA and which did not span previously identified exon/intron junctions. A 7195 bp genomic fragment containing 6 exons, ranging in size from 30 to 817 bp, two of which encode the entire DNA binding domain of the protein, was sequenced. The gene has features which are typical to "housekeeping" genes and is characterized by a very high content of G + C residues in a 1.4 kb fragment starting 500 bp from the cap site and by an "HTF" island in the 5' region. Transcriptional regulatory signals, exon/intraon boundaries and features characteristic of "housekeeping" genes are evolutionary conserved between the human and chicken genes. The HMG-17 gene was localized to human chromosome 1p12-34. RFLP's useful for further mapping were detected. The experimental evidence presented leads to the assumption that the gene characterized is the only functional human HMG-17 gene.

Amino Acid Sequence↗
PubMed1989

Chromosomal protein HMG-14. Identification, characterization, and chromosome localization of a functional gene from the large human multigene family.

The human HMG-14 multigene family is one of the largest retropseudogene families known. To identify and isolate a functional human HMG-14 gene, genomic clones, selected with the cDNA, were screened with a set of 6 oligonucleotides. A single genomic clone was isolated suggesting that the human genome contains few, and perhaps only one, functional genes. An 8882-base pair (bp) genomic clone containing the complete, 6804-bp-long human gene together with 850 bp 5' to the start of transcription and 1228 bp 3' to the end of transcription was sequenced. The gene is comprised of 6 exons ranging in size from 30 to 839 bp, two of which code for the entire DNA binding site of the protein, and has several features typical of "housekeeping" genes. Using human-rodent somatic cell hybrids, the HMG-14 gene was localized to human chromosome 21. A restriction fragment length polymorphism, useful for further analysis and mapping, has been detected. The present article, which describes the first isolation and characterization of a gene coding for chromosomal protein HMG-14, indicates that genes coding for HMG-14 and HMG-17 may share several distinctive characteristics. Comparison with the human and chicken HMG-17 genes reveals that all contain 6 exons, that all have exons of similar size, that all have 5' regions highly enriched in GC residues and that all have features typical of housekeeping genes.

Animals↗
PubMed1988

Cloning of the chicken chromosomal protein HMG-14 cDNA reveals a unique protein with a conserved DNA binding domain.

The isolation and sequencing of a cDNA clone coding for the entire sequence of chicken chromosomal protein HMG-14 is described. The open reading frame constitutes only 25% of the transcript; the 5'-untranslated region is extremely rich in GC residues; and the 3'-untranslated region is highly enriched in AT residues. Comparison with other cDNAs coding for HMG-14 and HMG-17 reveals that the transcripts of genes coding for this family of chromosomal proteins have a characteristic structure. The deduced amino acid sequence is unique and different from all other known HMG-14 and -17 sequences. Analysis of amino acid position identity between the chicken HMG-14 and other HMG-14/-17 proteins revealed that the protein has 37% similarity to the HMG-17 group and 69% similarity to the HMG-14 group; therefore, the protein is classified as belonging to the HMG-14 group. Additional analysis leads to the conclusion that the chicken cDNA described here codes for the true homolog of calf and human HMG-14 protein. Comparison of all the known HMG-14 sequences reveals the DNA binding domain is conserved and contains the invariant dodecapeptide PKRRSARLSAKP. The HMG-14 proteins have a distinct charge distribution along the polypeptide chain: while the central region is positively charged the C-terminal domain is negatively charged.

Animals↗
PubMed1988

Aberrant DNase I digestion kinetics of nucleosomal core particles from sea urchin sperm.

The pancreatic deoxyribonuclease (DNase I) digestion rates at the susceptible sites on nucleosomal core particles from blastula, gastrula and sperm cells of the sea urchin, Parechinus angulosus, have been determined. Although there are differences in their isohistone composition, the rates of digestion are similar for both embryonic stages. The rates of digestion for sperm core particles are 3-5 times lower than for embryo core particles at the more, and up to 2.5 times lower at the less susceptible sites. An explanation for these differences could be sought in the sperm isohistones H2B which are characterized by N-terminal extensions of 20-25 amino acid residues.

Animals↗
PubMed1988

Single copy gene for the chicken non-histone chromosomal protein HMG-17.

A chicken genomic library was screened with the human cDNA encoding the non-histone chromosomal protein HMG-17 and a 4565-base pair fragment containing the entire gene encoding this protein was isolated and characterized. Sequence analysis of the fragment revealed that from the start to end of transcription, the HMG-17 gene is 3293 base pairs long and is comprised of 6 exons ranging in size from 30 to 890 base pairs. Upstream of the putative cap site are both a CAAT box and a TATA box as well as several Sp1 binding sites. The gene has an extremely high content of G and C residues (75%) in a 1150-base pair fragment starting 500 base pairs from the putative cap site. This region satisfies the definition of an HpaII tiny fragment island. Southern analysis indicated that there is a single copy of this gene in chickens, whereas Northern analysis revealed that a single transcript is being synthesized from this gene. A comparison of the chicken and human cDNA and protein sequences and subsequent calculation of the evolutionary rates indicated that HMG-17 is a slowly evolving gene. The present article, which is the first study on the isolation and characterization of a complete gene coding for a high mobility group non-histone protein, indicates that the gene has features characteristic of housekeeping genes.

Amino Acid Sequence↗
PubMed1987

Retropseudogenes for human chromosomal protein HMG-17.

The human genome contains multiple copies of sequences homologous to the cDNA coding for non-histone chromosomal protein HMG-17. To study the mechanism of generation and dispersion of the HMG-17 multigene family a human genomic library was screened and 70 clones isolated and studied by Southern transfer and restriction site analysis. The results suggest that most of the clones contain unique sequences. Sequence analysis of two genomic clones indicates that they contain elements typical of processed retropseudogenes. Even though both sequences contained open reading frames the sequences lacked introns, were flanked by short, direct repeats and lacked elements associated with functional genes. The sequences of the two pseudogenes were 85% homologous to each other and each was 90% homologous to the human cDNA. Based on the sequence difference in the open reading frame between the pseudogenes and the cDNA it can be estimated that the sequences arose approximately ten million years ago from a common precursor. The present paper, which is the first study on genes coding for this nucleosomal binding protein, indicates that the HMG-17 multigene family is the largest known human retropseudogene family.

Base Sequence↗
PubMed1987

Modulation of the cellular ratio of chromosomal high mobility group proteins 14 to 17 in transfected cells.

The cDNAs coding for human nonhistone chromosomal high mobility group (HMG) proteins 14 and 17 have been introduced into the eukaryotic expression vector pSVL under the transcriptional control of the SV40 late promoter and the constructs used to transfect COS cells. Transfection with plasmid pSVL14s, containing the HMG-14 cDNA in the sense orientation, increased the endogenous levels of HMG-14 mRNA 50-fold and the levels of HMG-14 protein 3-fold. Transfection with pSVL17s, which contains the HMG-17 mRNA in the sense orientation, resulted in a 19-fold increase in mRNA levels and a 3-fold increase in the protein level. Transfection with pSVL17as, containing the HMG-17 in the antisense orientation, resulted in a noticeable decrease in the protein levels. The overproduction of HMG mRNAs does not affect the level of other cellular mRNAs and the increase in the cellular level of either HMG-14 or -17 did not affect the level of the other HMG or that of any other cellular protein examined. The results suggest that COS cells can tolerate large excess of HMG mRNAs and protein, that the relative amounts of HMG-14 and HMG-17 and their mRNAs are not constant, and that neither the transcription nor the translation of the proteins is coordinately regulated.

Animals↗
PubMed1987

Cell cycle regulated synthesis of an abundant transcript for human chromosomal protein HMG-17.

The abundance and cell cycle dependent expression of the mRNA for human nonhistone protein HMG-17 were studied in synchronized HeLa cells. Slot blot analysis indicates that the HMG-17 mRNA is a very abundant message, significantly more so than histone or actin mRNA. RNA prepared from tissue culture cells contains higher amounts of HMG-17 transcripts than RNA prepared from liver suggesting a correlation between the rate of cell division and HMG-17 mRNA levels. HMG-17 mRNA is present in the cells throughout the cell cycle however there is a significant increase in the mRNA levels late in S phase suggesting that the protein is deposited on chromatin after nucleosome assembly. Synthesis of the HMG-17 transcript is not coupled to DNA replication suggesting that the cell cycle related expression during late S phase is regulated in a different manner from that of the nucleosomal histones.

Cell Cycle↗
PubMed1986

Chromosomal protein HMG-14. Complete human cDNA sequence and evidence for a multigene family.

The isolation and sequencing of cDNA clone coding for the entire sequence of human nonhistone chromosomal protein HMG-14 is described. Sequence analysis reveals that the open reading frame constitutes only 25% of the transcript, that the 5'-untranslated region is extremely rich in GC residues (75%), and that the 3'-untranslated region is highly enriched in AT residues. The amino acid sequence, deduced from the reading frame, is 94% homologous to the calf thymus protein suggesting evolutionary constraints on the conformation of the protein. The human genome contains 60-90 HMG-14 gene copy equivalents which, as suggested by Southern analysis, are not tandemly arranged. Northern analysis of RNA isolated from several sources reveals that a single-sized mRNA codes for this protein. Southern analysis reveals that cross-hybridizing sequences are present in the genome of several different species indicating that the evolutionary origin of this gene was over 350 million years ago. The overall features of the human HMG-14 cDNA are very similar to those of the human HMG-17 cDNA, and the number of gene equivalents present in the human genome is similar for the two proteins. However, their nucleotide sequence is significantly different indicating that the multigene family coding for HMG-14 is distinct from that coding for HMG-17.

Base Sequence↗
PubMed1986

Chromosomal proteins HMG-14 and HMG-17. Distinct multigene families coding for similar types of transcripts.

Human nonhistone chromosomal proteins HMG-14 and HMG-17 are encoded by genes which are part of multigene families. Southern analysis of human, mouse, and rat genomic restriction digests reveals that the two families are distinct. Although the cDNAs of HMG-14 and HMG-17 do not cross-hybridize, they have several similar structural features: the open reading frame comprises only 23% of the transcripts, the 5'-untranslated region is extremely GC rich whereas the 3'-untranslated region is unusually long and AT rich. The overall sequence homology between the two transcripts is highest (71%) in the 90 nucleotides coding for the DNA-binding domains of the proteins. The sequence of the human HMG-14 and HMG-17 proteins, deduced from the open reading frame, differs by more than 50%; the DNA-binding domains of the proteins show 74% sequence homology. However, even in this 30-residue long peptide there are significant differences between the proteins as the proline content of HMG-17 (8 residues) is twice that of HMG-14. The two proteins have different hydropathy index profiles and are serologically distinct. The multigene families may have evolved independently from similar genetic elements or from a shared ancestral gene in which the nucleotide sequence coding for the DNA-binding domain of the protein is the most conserved region. The structural differences between the molecules and the differences in their DNA-binding domains suggest that the proteins may be involved in distinguishable cellular functions.

Amino Acid Sequence↗
PubMed1986

Immunofractionation of chromatin regions associated with histone H1o.

Two monoclonal antibodies, which were elicited against histone H5, bind to purified rat liver chromatin and to rat liver H1o but not to rat liver H1. The monoclonal antibodies were immobilized on CNBr-Sepharose and the resulting immunoaffinity column was used to fractionate rat liver oligonucleosomes. Enzyme-linked immunoabsorbant assay (ELISA) and immunoblotting experiments indicate that the nucleosomes bound to the column were tenfold enriched in their content of H1o. Oligonucleosomes, prepared from the livers of either untreated or 3-methylcholanthrene-treated adult rats, were fractionated on the anti-H1o affinity column. The DNA purified from the unfractionated nucleosomes, from the unbound nucleosomes and from the nucleosomes which were bound to the column was examined with various 32P-labeled probes. A slight enrichment in H1o was detected in the coding region of the rat albumin gene. In contrast DNA which was bound to the column was significantly depleted in sequences hybridizing with total cellular RNA (which contains mostly ribosomal RNA) and with sequences hybridizing to the 3'-terminal region of a cytochrome P-450 gene, which is inducible by the chemical carcinogen 3-methylcholanthrene, regardless of whether isolated from control or from carcinogen-treated rat livers. Our experiments clearly demonstrate that chromatin can be efficiently immunofractionated. The results suggest that the H1o content of chromatin regions containing genes which are constitutively transcribed is not necessarily different from that of regions containing non-transcribed genes and that highly inducible genes may be segregated into chromatin regions which are depleted of H1o.

Animals↗