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A R Kornblihtt

Publications and source records attributed to A R Kornblihtt.

47 records · Page 3Linked to original sources
PubMed1984

Human fibronectin: molecular cloning evidence for two mRNA species differing by an internal segment coding for a structural domain.

Two different fibronectin (FN) mRNA species were detected in the human cell line Hs578T. One species, mRNA I, contains an additional 270 nucleotide long insert (ED) that encodes exactly one of the internally repeated structural domains of the protein. The 90 amino acid extra domain belongs to the so-called type III homology and it is located in the carboxy-terminal half of FN, in between the cell attachment and the heparin binding sites of the protein. The evidence of two mRNAs is provided by the isolation and characterisation of four independent cDNA clones from a library prepared with a synthetic oligonucleotide primer, and it was confirmed by S1 nuclease analysis of cDNA/mRNA hybrids. This kind of analysis also showed that in the human cell line, mRNA I is present at a lower level than mRNA II (the mRNA species without the ED), whilst in human liver, mRNA I is virtually undetectable. Since liver tissue has recently been reported to be the source of plasma FN, our results indicate that the presence of the ED insert could be a particular feature of cellular FN.

Amino Acid Sequence↗
PubMed1984

Expression of a human alpha-globin/fibronectin gene hybrid generates two mRNAs by alternative splicing.

We have isolated genomic clones for human fibronectin (FN), by screening a human gene library with previously isolated FN cDNA clones. We have recently reported two different FN mRNAs, one of them containing an additional 270 nucleotide insert coding for a structural domain ED. Restriction mapping and DNA sequencing of the genomic clones show that the ED type III unit corresponds to exactly one exon in the gene, whilst the two flanking type III units are split in two exons at variable positions. When an alpha-globin/FN gene hybrid construct, containing the ED exon, flanking introns and neighbouring FN exons, is transfected into HeLa cells, two hybrid mRNAs differing by the ED exon are synthesized. These experiments confirmed that the two FN mRNAs observed in vivo arise from the same gene by alternative splicing.

Bacteriophage lambda↗
PubMed1983

Reconstitution of a hormone-sensitive adenylate cyclase with membrane extracts from Neurospora and avian erythrocytes.

Adenylate cyclase activity associated to wild type Neurospora membranes is highly dependent on Mn2+ and insensitive to fluoride, guanyl nucleotides, and cholera toxin. These membranes are able to interact with components of detergent extracts from turkey erythrocyte ghosts. The reconstituted cyclase system is catalytically active in the presence of Mg2+ and it is activated by guanyl-5'-yl imidodiphosphate plus isoproterenol and fluoride. When detergent extracts were prepared from avian erythrocyte membranes treated with cholera toxin, the reconstituted system was stimulated by guanyl-5'-yl imidodiphosphate in the absence of isoproterenol and cyclase activities were higher than those observed with extracts from membranes not treated with the toxin. Dose-response curves for isoproterenol and fluoride in the reconstituted system were similar to those reported for avian erythrocyte and liver membranes, respectively.

Adenylyl Cyclases↗
PubMed1983

Gene structure of human apolipoprotein A1.

Apolipoprotein A1 is the major polypeptide of the human plasma high density lipoprotein (HDL). The structure and function of the apo A1 gene are of interest because of the inverse correlation shown between HDL levels and coronary heart disease. We have determined the nucleotide sequence of the apo A1 gene previously isolated. Its coding sequence is interrupted by two introns of 185 and 588 base pairs. As there may be one or more introns in the 5' non coding region, the 5' end of the gene could not be precisely located. The human apo A1 has an unusual propeptide segment very similar to its rat counterpart. The data reported here provide an essential basis for future studies of structural and functional alleles of the apo A1 gene.

Amino Acid Sequence↗
PubMed1983

Isolation and characterization of cDNA clones for human and bovine fibronectins.

A bovine fibronectin (FN) cDNA clone (pFB1) was isolated by screening a cDNA library of calf testis fibroblasts with a synthetic oligonucleotide probe. The probe was a mixture of eight 14-base-long oligonucleotides designed from the amino acid sequence Glu-Cys-Phe-Met-Pro present in the Mr 3,000 COOH-terminal fragment of bovine plasma FN [Petersen, T.E., Thøgersen, H.C., Skorstengaard, K., Vibe-Pedersen, K., Sahl, P., Sottrup-Jensen, L. & Magnusson. S. (1983) Proc. Natl. Acad. Sci. USA 80. 137-141]. pFB1 contained a 1,000 base-pair (bp) insert comprising the complete 3' noncoding sequence (690 bp) and approximately equal to 300 bp of the coding region. The clone pFB1 was used as a radioactive probe in the screening of a human cell line (Hs 578T) cDNA library. Eleven positive cDNA clones were detected, one of which, named pFH1, contained a 2,000-bp insert comprising the complete 3' noncoding region (693 bp) and approximately equal to 1,300 bp of the coding region of human FN. The sequences of the clone pFB1 insert and of the homologous region in clone pFH1 were determined. The nucleotide sequences are 90% homologous. Six amino acid changes were found, clustered in an area connecting two structural domains described in bovine plasma FN. Furthermore, the 204 COOH-terminal amino acid sequence of bovine FN was completed by overlapping two peptide fragments (MrS 3,000 and 23,000). Clone pFH1 was used in estimating the size of human fibronectin mRNA (7,900 bases) through blot hybridization analysis. Southern blot studies suggest that human FN is coded by a unique gene.

Animals↗
PubMed1982

Soluble adenylate cyclase activity in Neurospora crassa.

A soluble form of adenylate cyclase was extracted from mycelia of Neurospora crassa wild-type strains. This enzyme activity was purified by chromatography on hexyl-amino-Sepharose, agarose and Blue Sepharose and preparative polyacrylamide-gel electrophoresis. On sodium dodecyl sulphate/polyacrylamide-gel electrophoresis, peak fractions from the later purification steps showed a main polypeptide band with an apparent molecular weight of about 66 000. The following hydrodynamic and molecular parameters were established for the Neurospora soluble adenylate cyclase activity: sedimentation coefficient, 6.25 S; Stokes radius, 7.3 nm; partial specific volume, 0.74 ml/g; molecular weight, 202 000; frictional ratio, 1.65. The isoelectric point of this enzyme activity was 4.65. The enzyme was not activated by GTP, [beta gamma-imido]GTP, fluoride or cholera toxin.

Adenylyl Cyclases↗
PubMed1981

Association of triiodothyronine binding activity to soluble adenylate cyclase in testicular preparations.

Cytosolic adenylate cyclase activity from rat seminiferous tubules was purified by chromtography in DEAE-cellulose, hydroxylapatite and Bio-Gel A-0.5 m as well as by centrifugation in sucrose gradients. In all these purification steps, fractions with adenylate cyclase activity also contained binding activity for L-T3. Binding studies indicate the existence of two L-T3 receptor components associated to adenylate cyclase activity. The component exhibiting the highest hormone affinity has the lowest binding capacity.

Adenylyl Cyclases↗
PubMed1981

Manganese ion dependent adenylate cyclase activity in rat testes: purification and properties.

Testicular, soluble adenylate cyclase has been purified by anion-exchange chromatography, gel filtration, and isoelectric focusing. Upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis, peak fractions from the latter purification step showed only one polypeptide band with an apparent molecular weight of about 69 000. The following hydrodynamic and molecular parameters have been established for this enzyme: sedimentation constant, 4.3; Stokes radius, 3.95 nm; partial specific volume, 0.74 mL.g(-1); molecular weight, 74 000; fractional ratio, 1.4.

Adenylyl Cyclases↗
PubMed1981

Inhibition of cyclic 3',5' monophosphate synthesis in rat testis by L-triiodothyronine.

Cytosolic adenylate cyclase activity from rat seminiferous tubules is inhibited by L-triiodothyronine (L-T3). In a typical dose-response curve, using Mn-ATP as substrate, no effect is observed at 10(-10) M L-T3; about 15 to 25% inhibition is found in the range between 10(-9) and 10(-6) M L-T3 and finally a sharp enzyme inhibition is evident at increasing hormone concentrations from 10(-6) to 10(-4) M. Incubation of decapsulated testes with L-T3 leads to a decrease of intracellular cyclic AMP levels. Dose-response relationships for such effect are similar to those found for adenylate cyclase activity. In this case a clear response is observed at 10(-8) M L-T3.

Adenylyl Cyclase Inhibitors↗
PubMedDate not supplied

MRNA encoding a putative RNA helicase of the DEAD-box gene family is up-regulated in trypomastigotes of Trypanosoma cruzi.

Differential display of mRNAs from Trypanosoma cruzi epimastigote and metacyclic trypomastigote stages showed several mRNA species differing in their expression level. The cDNA corresponding to one of these mRNAs was used as a probe in Northern blots and identified a RNA product of 2.6 kb with an expression level eight or more times higher in trypomastigotes than in epimastigotes. This probe was also used to screen a genomic library of T. cruzi CL Brener clone prepared in lambda FIX. A clone of about 15 kb was selected that, after partial sequencing, revealed an open reading frame of 688 amino acids encoding a deduced protein with similarity to RNA helicases of the DEAD-box gene family. The presence of the eight conserved motifs characteristic of the DEAD protein family was observed in the T. cruzi sequence, indicating that it corresponds to a putative RNA helicase gene, which we named HelTc. Southern blot analysis indicated that HelTc is a single-copy gene. Pulsed-field gel electrophoresis separation of chromosomes of several isolates of T. cruzi showed that this gene was localized in one or two chromosomal bands.

Amino Acid Motifs↗