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Biomedical subjects

R E Kaufman

Publications and source records attributed to R E Kaufman.

At least 37 records · Page 2Linked to original sources
PubMed1991

Isolation and characterization of the genomic human CD7 gene: structural similarity with the murine Thy-1 gene.

The human CD7 molecule is a 40-kDa member of the immunoglobulin gene superfamily that is expressed on T-lymphoid and myeloid precursors in fetal liver and bone marrow. CD7 is also expressed on T lymphocytes in multiple stages of T-cell development, including a major subset of mature peripheral T cells. In this paper we report the isolation and characterization of the human CD7 gene and 5' flanking region. Sequence analysis revealed that the CD7 gene comprises four exons that span 3.5 kilobases. The 5' flanking region (506 base pairs) has a high G + C content and no "TATA" or "CCAAT" elements. DNase I sensitivity analysis of chromatin from the CD7+ progenitor cell leukemia line, DU528, and the CD7-, CD4+, CD8+, TCR alpha beta + T-cell line, DU980 (where TCR is the T-cell receptor), revealed two distinct hypersensitive sites 5' of the CD7 gene. Hypersensitive site 1, present in the DU980 T-cell line, was located 4.5 kilobases upstream of the presumed CD7 transcription initiation site. Only DNase I hypersensitive site 2, which mapped to the promoter region, was found in the DU528 line. Comparison of the organization of the CD7 gene with that of other members of the immunoglobulin gene superfamily revealed that the human CD7 gene most closely resembles the murine Thy-1 gene. Both CD7 and Thy-1 are encoded by small genes with four exons, contain TATA-less promoters, and have a similar functional organization. These structural similarities suggest that human CD7 and murine Thy-1 may be functional homologues.

Amino Acid Sequence↗
PubMed1990

Dissociation of thymidine incorporation and transferrin receptor expression from cell growth and c-myc accumulation in alpha-interferon-treated cells.

Alpha-interferon is capable of altering the pattern of growth of both normal and neoplastic cells, but the pathways essential to sensitivity and resistance to alpha-interferon are unknown. To explore the growth inhibition induced by alpha-interferon, we examined the interferon-sensitive cell line Daudi and the resistant cell line HL-60. In Daudi, alpha-interferon induced a fall in c-myc mRNA accumulation at 24 h, inhibited tritiated thymidine ([3H]Thd) uptake at 48-72 h, and inhibited proliferation at 72-96 h. The half-life of c-myc mRNA was shortened from 31 to 13 min by alpha-interferon treatment. In HL-60, no alteration in c-myc accumulation or cell growth was observed, but [3H]Thd uptake was inhibited by 49%. Exogenous thymidine partially reversed the effects of alpha-interferon on [3H]Thd incorporation. The number of transferrin receptors, as measured by immunofluorescence, was unaffected by alpha-interferon in both cell lines. We conclude that the growth inhibitory effects of alpha-interferon are neither dependent upon inhibition of thymidine metabolism nor on expression of the transferrin receptor, but may be linked to control of c-myc.

Cell Division↗
PubMed1989

Hodgkin's disease presenting as myelofibrosis.

Four patients with Hodgkin's disease and bone marrow fibrosis are presented in whom the clinical presentation was dominated by cytopenias; this was associated with a delayed diagnosis for an average of 20 months. Despite marrow involvement, chemotherapy resulted in complete remissions and two patients appear to have been cured. Marrow fibrosis resolved at least partially after chemotherapy. The medical literature relevant to bone marrow involvement by Hodgkin's disease is reviewed. Hodgkin's disease should be considered in the differential diagnosis of idiopathic myelofibrosis.

Adult↗
PubMed1988

Antifibrinolytic therapy in the management of the Kasabach Merritt syndrome.

The Kasabach Merritt syndrome consists of thrombocytopenia, microangiopathic hemolytic anemia, and a localized consumption coagulopathy that develops within the abnormal vascular channels of a hemangioma. In general, these patients demonstrate only mild abnormalities of screening clotting tests, but they can potentially develop life-threatening complications. We present a patient who developed a severe anemia that was refractory to erythrocyte transfusions. Treatment with epsilon-aminocaproic acid to inhibit fibrinolysis and cryoprecipitate to replenish his deficient circulating fibrinogen interrupted the cycle of his systemic coagulopathy and enabled us to transfuse him to a normal hematocrit.

Adult↗
PubMed1988

Adenosine deaminase (ADA) deficiency due to deletion of the ADA gene promoter and first exon by homologous recombination between two Alu elements.

In 15-20% of children with severe combined immunodeficiency (SCID), the underlying defect is adenosine deaminase (ADA) deficiency. The goal of this study was to determine the precise molecular defect in a patient with ADA-deficient SCID whom we previously have shown to have a total absence of ADA mRNA and a structural alteration of the ADA gene. By detailed Southern analysis, we now have determined that the structural alteration is a deletion of approximately 3.3 kb, which included exon 1 and the promoter region of the ADA gene. DNA sequence analysis demonstrates that the deletion created a novel, complete Alu repeat by homologous recombination between two existing Alu repeats that flanked the deletion. The 26-bp recombination joint in the Alu sequence includes the 10-bp "B" sequence homologous to the RNA polymerase III promoter. This is the first example of homologous recombination involving the B sequence in Alu repeats. Similar recombination events have been identified involving Alu repeats in which the recombination joint was located between the A and B sequences of the polymerase III split promoter. The nonrandom location of these events suggests that these segments may be hot spots for recombination.

Adenosine Deaminase↗
PubMed1987

Differences in the regulation of messenger RNA for housekeeping and specialized-cell ferritin. A comparison of three distinct ferritin complementary DNAs, the corresponding subunits, and identification of the first processed in amphibia.

The ferritin family is a widespread group of proteins that maintain iron in a soluble form and also protect against the toxic effects of excess iron. The structure and sequence of the proteins are highly conserved. However, the cell-specific features of structure which occur within the same organism indicate cell specificity of gene expression and may be related to variations in types of iron storage, i.e. specialized-cell ferritin (stored iron is for other cell types) versus housekeeping ferritin (stored iron is for intracellular purposes related to normal or stress metabolism); the protein structure may also affect rates of iron turnover. Iron induces ferritin synthesis and accumulation by recruiting stored ferritin mRNA that is efficiently translated in cells specialized for iron storage. For the first time we show the occurrence of three different cDNAs from bullfrog tadpoles, corresponding to three subunits of the protein: H, M, and L. Thus, ferritin can be encoded by at least three different mRNAs and probably three different genes, in contrast to the older idea of two, H and L; the subunits maintain the conserved sequences of known ferritins and have similar predicted masses, 20.5, 20.6, and 19.9 kDa, but have distinct mobilities in denaturing gels. Ferritin subunit expression is cell specific; more of the H and L chain mRNAs are expressed in red cells than in liver. Ferritin expression is regulated by transcription (or mRNA stability) in adult red cells; cellular levels of ferritin mRNA were 20% that of embryonic red cells, and L subunit mRNA increased 2.5 times with excess iron. Ferritin expression is also regulated during translation in adult red cells; iron recruits stored ferritin mRNA, but only during certain stages of red cell maturation, in contrast to embryonic red cells. The developmental differences in ferritin expression are discussed in relation to the shift from specialized-cell ferritin to housekeeping ferritin in red cells of the embryonic versus adult lines.

Amino Acid Sequence↗
PubMed1987

Identification of a deletion in the adenosine deaminase gene in a child with severe combined immunodeficiency.

A patient with adenosine deaminase-deficient severe combined immunodeficiency is described whose defect is secondary to deletion of a portion of the ADA structural gene. In Southern analyses, DNA from this patient does not hybridize to a genomic probe that includes the 3' end of exon 1. This implies that both his parents are heterozygous for deletions of exon 1 sequences. Consistent with this finding, the patient has no detectable adenosine deaminase mRNA by Northern analysis. This is the first report of a deletion mutation as the cause of adenosine deaminase deficiency.

Adenosine Deaminase↗
PubMed1986

Treatment of murine peritoneal macrophages with bacterial lipopolysaccharide alters expression of c-fos and c-myc oncogenes.

Expression of the c-fos, c-myc, and c-fms proto-oncogenes has been studied in thioglycollate-elicited murine peritoneal macrophages after exposure to lipopolysaccharide (LPS). After incubation with LPS (20 ng/ml), a transient and rapid induction of the expression of c-fos and c-myc oncogenes could be observed, whereas the RNA levels for c-fms were not affected. Treatment with lipid A, the active moiety of the LPS molecule, increased the c-fos and c-myc expression to a comparable degree. Similar induction of c-fos and c-myc was observed after treatment with phorbol myristate acetate, suggesting that this effect of LPS on murine macrophages might be mediated through stimulation of protein kinase C. Under similar experimental conditions, LPS treatment of macrophages did not trigger DNA synthesis. Treatment with LPS blocked DNA synthesis in macrophages treated with L cell-conditioned medium containing colony-stimulating factor. Thus changes in c-fos and c-myc expression may be elements in the complex series of biochemical events that contribute to macrophage activation and are not necessarily related to induction or priming for cellular proliferation.

Animals↗
PubMed1986

Multiple red cell ferritin mRNAs, which code for an abundant protein in the embryonic cell type, analyzed by cDNA sequence and by primer extension of the 5'-untranslated regions.

Ferritin maintains iron in a bioavailable, nontoxic form for vertebrates and invertebrates, higher plants, fungi, and bacteria; the protein is formed from two classes of subunits (H and L) in ratios which vary in different cell types. Ferritin may be an abundant, differentiation-specific protein or a "housekeeping" protein. The red cells of embryos are specialized for iron storage and have abundant ferritin; iron regulates the synthesis of ferritin in such cells translationally by recruitment of stored, ferritin mRNA and by translational competition. To characterize mRNA regulated in such a manner, we prepared cDNA from reticulocytes of bullfrog tadpoles, a readily available source of embryonic red cells; moreover, no protein sequence information was available for nonmammalian ferritin. An almost full-length (817 base pairs) cDNA (pJD5F12) was isolated and sequenced, the 5' end was analyzed by primer extension, and the cloned DNA was used as a hybridization probe. We have shown that ferritin mRNA is stored in the cytoplasm and that the 5' end of the mRNA is heterogeneous. The 5'-untranslated region of ferritin mRNA consisted of 143 nucleotides in the major (65%) species and 146 or 152 in the minor species (approximately 17% each). (Heterogeneity is characteristic of some other abundant mRNAs, e.g. globin, which is also translationally regulated.) Since excess iron had no detectable effect on the heterogeneity of the 5' end of ferritin mRNA, the feature is more likely associated with mRNA abundance and/or cell specialization than translational control. In the bullfrog, as in humans and rats, ferritin is encoded by multiple genomic sequences (four to eight) which specify proteins of considerable homology. For example, 75 of the 81 amino acids present in all mammalian ferritins sequenced are also present in the frog; the overall homology between frogs and humans or rats is 59-66%. Ferritin H and L subunits in humans are distinct (overall homology 56%) and appear to have diverged from a common precursor relatively recently. In contrast, ferritin H and L subunits have high homology in tadpole red cells, determined by hybrid select translation, which suggests that bullfrog red cell ferritin may be close to the primordial sequence.

Amino Acid Sequence↗
PubMed1986

Regulation of nuclear and mitochondrial gene expression by contractile activity in skeletal muscle.

Increased contractile activity of skeletal muscle augments the volume fraction and enzymatic capacity of mitochondria and suppresses the enzymatic capacity of several cytoplasmic enzymes of glycolysis. To examine the biochemical mechanisms underlying these effects, we measured the concentrations of cytochrome b mRNA and aldolase A mRNA in tibialis anterior muscles of adult rabbits that had been stimulated via the motor nerve to contract continuously at 10 Hz for 5 or 21 days; these were compared with the corresponding levels in the unstimulated limbs of the same animals. After 21 days of stimulation aldolase mRNA had fallen to one-fourth of control levels, while cytochrome b mRNA had increased by 5-fold. A reduction in aldolase mRNA was already evident after only 5 days of stimulation, whereas the level of cytochrome b mRNA was not elevated at this stage. Mitochondrial DNA was unchanged after 5 days but had increased by 4-fold after 21 days. We conclude that contractile activity in skeletal muscle produces reciprocal changes in the expression of these two genes at a transcriptional level but via different regulatory mechanisms. Enhancement of the expression of the mitochondrial cytochrome b gene appears to be proportional to the increase in its copy number and may not, therefore, depend upon changes in transcriptional or translational efficiency. The reduction in aldolase A mRNA occurs at an earlier stage in the response to contractile activity and is probably mediated by a reduced transcriptional efficiency.

Animals↗
PubMed1986

Dibutyryl cyclic adenosine monophosphate reduces expression of c-myc during HL-60 differentiation.

The human promyelocytic leukemia cell line HL-60 is induced to differentiate along a myelocytic pathway by dibutyryl cyclic adenosine monophosphate (dbcAMP). Other cAMP analogs are ineffective as inducing agents. The effect of these compounds on expression of c-myc was investigated using a DNA probe for c-myc to detect RNA transcripts. The dose response and time to commitment for reduction in c-myc expression with dbcAMP was similar to the findings for phenotypic changes. Bromo-cyclic AMP and butyrate alone caused no changes in c-myc expression in 24 hours, but demonstrated dramatic synergism together, suggesting that butyrate contributes in part to the effects of dbcAMP. Evidence for mechanisms of action of cAMP other than activation of the cAMP-dependent protein kinase is reviewed.

Bucladesine↗
PubMed1985

Beta-thalassemia resulting from a single nucleotide substitution in an acceptor splice site.

Beta-globin gene mutations which alter normal globin RNA splicing have confirmed the necessity of invariant nucleotides GT at donor splice sites. Functional consequences of point mutations in the invariant AG acceptor splice site have not been determined. We have isolated and characterized a beta-globin gene from a Black patient with beta-thalassemia intermedia which has an A-G transition at the usual intervening sequence 2 (IVS2) acceptor splice site. Functional analysis of transcripts produced by this mutant gene in a transient expression vector indicates that the mutation inactivates the normal acceptor splice site and results in some utilization of a cryptic splice site near position 580 of IVS2. This mutation would be expected to produce a beta-globin gene which results in no normal beta-globin mRNA.

Base Sequence↗
PubMed1985

Polymorphism of DNA sequence in the human pro alpha 2(I) collagen gene.

The human pro alpha 2(I) collagen gene was analysed for the presence of restriction fragment length polymorphisms. DNA from randomly selected unrelated persons of three Southern African populations was cleaved with one of eight different restriction enzymes, electrophoresed, blotted, and hybridised with cDNA and genomic probes specific for the pro alpha 2(I) gene. An MspI polymorphism was detected which results from the loss of a cleavage site within the 3' half of the gene. In two of the populations studied, the polymorphism occurred at significant frequencies, and should therefore prove useful as a genetic marker for the study of inherited disorders of connective tissue involving collagen structure or biosynthesis.

Chromosome Mapping↗
PubMed1984

Isolation and characterization of a human pro alpha 2(I) collagen gene segment.

Over 20 kilobase pairs of the human pro alpha 2(I) collagen gene have been isolated and characterized by restriction endonuclease mapping, cell-free translation of hybrid-selected RNA, and DNA sequence analysis. We have sequenced an exon and determined its length to be 108 base pairs (bp). This is consistent with the organization of chick and sheep collagen genes in that exons are multiples of 9 bp in length, frequently being 54 and 108 bp. The sequenced exon was bordered by a GT (guanine-thymine) at its 3' end and an AT (adenine-thymine) at its 5' end. This pattern has been found at all normal intron-exon junctions in eukaryotic cells. The amino acid sequence derived from DNA sequencing of this 108 bp exon revealed 88% homology compared to the amino acid sequence of bovine pro alpha 2(I). The bases encoded 12 Gly-X-Y triplets characteristic of the helical portion of collagen. A unique sequence Gly-Gly-Lys-Gly-Glu-Lys identified this fragment as alpha 2(I) collagen.

Amino Acid Sequence↗
PubMed1981

Analysis of globin gene structure in patients with beta thalassemia by restriction endonuclease mapping.

Twenty-six DNA samples from individuals either heterozygous or homozygous for beta thalassemia were analyzed by restriction endonuclease digestion, agarose gel electrophoresis, and Southern blot analysis to define DNA fragments containing portions or all of the beta globin gene. A total of twenty-seven genes affected by a beta thalassemia mutation and twenty-seven genes affected by a beta thalassemia mutation and twenty-two normal beta globin genes were examined in Italian, Greek, or Asian individuals. With all four restriction endonucleases used, the fragments generated from DNA of thalassemic individuals were identical to those found in DNA from normal. Thus, gross rearrangement or deletion within the genomic region containing the beta globin gene is not characteristic of mutations which cause a thalassemia. A third patient homozygous for pancellular hereditary persistence of fetal hemoglobin was shown to have complete deletion of the delta and beta globin genes.

Base Sequence↗