Kilobase Pairs In Length Research Articles

Differentiation-induced Gene Expression in 3T3-L1 Preadipocytes: A Second Differentially Expressed Gene Encoding Stearoyl-CoA Desaturase

Abstract Previously we isolated and characterized a differentially expressed gene from mouse 3T3-L1 preadipocytes that encodes stearoyl-CoA desaturase (SCD1; Ntambi, J. M., Buhrow, S. A., Kaestner, K. H., Christy, R. J., Sibley, E., Kelly, T. J., Jr., and Lane, M. D. (1988) J. Biol. Chem. 263, 17291-17300). Genomic Southern blot analysis indicated the existence of another closely related gene. Here we report the isolation and characterization of this gene and the corresponding cDNA which encode a second stearoyl-CoA desaturase, SCD2, 3T3-L1 adipocytes. SCD2 cDNA is 5 kilobase pairs in length and encodes a protein of 358 amino acids with greater than 87% amino acid sequence identity to SCD1. RNase protection analysis reveals a 10-fold increase in the expression of SCD2 mRNA during 3T3-L1 preadipocyte differentiation. SCD2 mRNA is expressed constitutively at a high level in brain, is not expressed in liver, and its expression in kidney, adipose, and lung tissue is increased greatly by shifting mice from a diet containing unsaturated fatty acids to a diet devoid of fat. The tissue distribution and the dietary alteration of SCD1 mRNA expression differs markedly from that of SCD2 mRNA being absent from brain, constitutive in adipose tissue, and subject to negative control in liver by feeding a diet containing unsaturated fatty acids. The SCD2 gene spans approximately 15 kilobase pairs and consists of six exons and five introns, with intron/exon junctions similar to those of SCD1. As determined by primer extension analysis the start site of transcription maps 300 nucleotides upstream of the initiator methionine codon. Unlike the SCD1 gene, SCD2 lacks a typical TATA box in the 5'-flanking region, but has two boxes at positions -90 and -135 relative to the transcription initiation site. The SCD2 promoter contains a 140-base pair sequence (located between nucleotides -54 and -201) which possesses 77% sequence identity to a region (located between nucleotides -472 and -325) in the SCD1 promoter. There is a GC-rich sequence in the SCD2 promoter (at nucleotide -175) similar to the binding site for the nuclear transcription factor Sp1 as well as an element with homology to the core consensus sequence for the glucocorticoid regulatory element position -500 and a potential CCAAT box/enhance binding protein sequence at position -540. The SCD gene family provides a new model system for the study of differentiation-induced as well as tissue-specific metabolite controlled gene expression.

Sequence and expression of chicken and mouse rsk: homologs of Xenopus laevis ribosomal S6 kinase.

We have previously reported the isolation of cDNAs encoding two closely related Xenopus ribosomal S6 kinases, S6KII alpha and -beta (S. W. Jones, E. Erikson, J. Blenis, J. L. Maller, and R. L. Erikson, Proc. Natl. Acad. Sci. USA 85:3377-3381, 1988). We report here the molecular cloning of one chicken and two mouse homologs of the Xenopus laevis cDNAs. As described for the Xenopus proteins, these cDNAs were found to predict polypeptides that contain two distinct kinase domains, of which one is most closely related to the catalytic subunit of cyclic AMP-dependent protein kinase and the other is most closely related to the catalytic subunit of phosphorylase b kinase. The three predicted proteins were more than 79% identical to the Xenopus S6KII alpha protein. The chicken and one of the mouse cDNAs were, respectively, 3.7 and 3.1 kilobase pairs in length, predicted proteins of 752 and 724 amino acids with molecular weights of 84.4 and 81.6 kilodaltons, and hybridized to mRNAs in fibroblasts and tissues of approximately 3.6 and 3.4 kilobases (kb). The second mouse cDNA was approximately 6.1 kilobase pairs and was not full length but predicted the C-terminal 633 amino acids of a protein that is similar to the C-terminal portion of Xenopus S6KII alpha. This clone hybridized to mRNA transcripts of 7.6 and 3.4 kb. In vitro transcription and translation of the chicken and the mouse cDNAs that predict complete proteins produced major products with apparent molecular weights of 96 and 84 kilodaltons. Analysis of mRNA levels in chicken tissues showed significant quantities of the 3.6-kb transcript in small and large intestine, spleen, and bursa. Both mouse cDNA were similarly expressed at significant levels in intestine, thymus, and lung; however, the 7.6-kb mRNA was differentially and more highly expressed in heart and brain. The two mouse cDNAs represent two different S6 kinase genes, as shown by comparison of their protein sequences, mRNA transcript sizes, genomic organizations, and nucleic acid sequences. We propose that this family of genes be named rsk, for ribosomal S6 kinase.

Nucleotide sequence of the transcriptional control region of the osmotically regulated proU operon of Salmonella typhimurium and identification of the 5' endpoint of the proU mRNA

Southern blot analysis of 15 proU transposon insertions in Salmonella typhimurium indicated that this operon is at least 3 kilobase pairs in length. The nucleotide sequence of 1.5-kilobase-pair fragment that contains the transcriptional control region of the proU operon and the coding sequences specifying 290 amino acids of the first structural gene of the operon was determined. The predicted amino acid sequence of the product of this gene shows extensive similarity to the HisP, MalK, and other proteins that are inner membrane-associated components of binding protein-dependent transport systems. S1 mapping and primer extension analysis of the proU mRNAs revealed several species with different 5' ends. Two of these endpoints are sufficiently close to sequences that have weak similarities to the consensus -35 and -10 promoter sequences that they are likely to define two transcription start sites. However, we cannot rule out the possibility that some or all of the 5' endpoints detected arose as a result of the degradation of a longer mRNA. The expression of proU-lacZ operon fusions located on plasmids was normal in S. typhimurium regardless of the plasmid copy number. The sequences mediating normal, osmoregulated expression of the proU operon were shown by subcloning to be contained on an 815-base-pair fragment. A 350-base-pair subclone of this fragment placed onto a lacZ expression vector directed a high-level constitutive expression of beta-galactosidase, suggesting that there is a site for negative regulation in the proU transcriptional control region which has been deleted in the construction of this plasmid.

The gene coding for the 190,000-dalton iron-regulated protein of Yersinia species is present only in the highly pathogenic strains

A genomic library containing DNA fragments of 0.5 to 2 kilobase pairs in length from Yersinia enterocolitica serovar O:8 was constructed in a bacteriophage lambda gt11 expression vector. Mouse antibodies specific for the iron-regulated high-molecular-weight proteins (HMWPs) were used to screen the library. Two positive clones of 1 and 0.5 kilobase pairs, designated A13 and D7, respectively, were detected and isolated. They coded for beta-galactosidase fusion proteins of 151,000 and 138,000 daltons (Da). Antibodies affinity purified on the two recombinant lambda gt11 vectors specifically recognized the smaller HMWP (190,000 Da) and not the larger (240,000 Da). The two cloned DNA fragments were used to construct recombinant amplification plasmid pUC13 and to obtain large amounts of purified A13 and D7 inserts. Southern hybridizations performed with the inserts used as probes revealed that: (i) the two cloned DNA fragments overlap; (ii) only one gene hybridizes with the A13 and D7 inserts; (iii) the gene coding for the HMWP is conserved among all highly pathogenic Yersinia species studied; (iv) this gene is missing in the low-virulence and nonvirulent strains; and (v) transcription of the HMWP gene is induced by iron starvation.

Characterization of the glycinin gene family in soybean.

We characterized the structure, organization, and expression of genes that encode the soybean glycinins, a family of storage proteins synthesized exclusively in seeds during embryogenesis. Five genes encode the predominant glycinin subunits found in soybeans, and they have each been cloned, sequenced, and compared. The five genes have diverged into two subfamilies that are designated as Group-I and Group-II glycinin genes. Each glycinin gene contains four exons and three introns like genes that encode related proteins in other legumes. Two other genes have been identified and designated as "glycinin-related" because they hybridize weakly with the five glycinin genes. Although not yet characterized, glycinin-related genes could encode other glycinin subunit families whose members accumulate in minor amounts in seeds. The three Group-I glycinin genes are organized into two chromosomal domains, each about 45 kilobase pairs in length. The two domains have a high degree of homology, and contain at least five genes each that are expressed either in embryos or in mature plant leaves. Gel blot studies with embryo mRNA, as well as transcription studies with 32P-RNA synthesized in vitro from purified embryo nuclei, indicate that glycinin and glycinin-related genes become transcriptionally activated in a coordinated fashion early in embryogenesis, and are repressed coordinately late in seed development. In addition to transcriptional control processes, posttranscriptional events also are involved in regulating glycinin and glycinin-related mRNA levels during embryogenesis.

The REV1 gene of Saccharomyces cerevisiae: isolation, sequence, and functional analysis

The REV1 gene of Saccharomyces cerevisiae is required for normal induction of mutations by physical and chemical agents. We have determined the sequence of a 3,485-base-pair segment of DNA that complements the rev1-1 mutant. Gene disruption was used to confirm that this DNA contained the REV1 gene. The sequenced segment contains a single long open reading frame, which can encode a polypeptide of 985 amino acid residues. The REV1 transcript is 3.1 kilobase pairs in length. Frameshift mutations introduced into the open reading frame yielded a Rev-phenotype. A base substitution, encoding Gly-193 to Arg-193, was found in this open reading frame in rev1-1. Deletion mutants, lacking segments of the 5' region of REV1, had intermediate mutability relative to REV1 and rev1-1; a complete deletion exhibited lower mutability than rev1-1. REV1 is not an essential gene. An in-frame fusion of the 5' end of the REV1 open reading frame to the lacZ gene produced beta-galactosidase activity constitutively. The predicted REV1 protein is hydrophilic, with a predicted pI of 9.82. No homologies to RAD1, RAD2, RAD3, RAD7, or RAD10 proteins were noted. A 152-residue internal segment displayed 25% identity with UMUC protein.

Human ornithine-delta-aminotransferase. cDNA cloning and analysis of the structural gene.

Ornithine-delta-aminotransferase (OAT) is a nuclear-encoded, mitochondrial matrix enzyme which, in rat, is expressed as basal levels in most tissues but is induced in liver by high dietary protein and in kidney by estrogen and thyroxine administration. In man, the hereditary deficiency of OAT results in ornithine accumulation and the blinding disease gyrate atrophy of the choroid and retina. We cloned near full length rat and human liver OAT cDNAs and demonstrated OAT expression in a variety of tissues from each species. We mapped the human OAT structural gene to chromosome 10, cloned 40 kilobase pairs of genomic DNA containing the complete OAT structural gene, and determined its organization. It is 21 kilobase pairs in length, and contains 11 exons. Exon 2 has been absent from all cDNAs studied and was detected by homology to X-linked processed OAT pseudogenes. The 5'-flanking region of the OAT gene has features of housekeeping genes (GC enrichment and three Sp1 binding consensus sequences) and tissue-specific, inducible genes (TATA box-like element and two CCAAT boxes). A 22-base pair region of partial dyad symmetry containing homology to estrogen responsive elements overlaps the OAT transcription site. Another 5' sequence, GTATCCTGCCCTC, is homologous to sequences in the promoter regions of the genes of three urea cycle enzymes.

Nucleotide sequence and transcriptional start site of the Methylobacterium organophilum XX methanol dehydrogenase structural gene

The nucleotide sequence of a cloned 2.5-kilobase-pair SmaI fragment containing the methanol dehydrogenase (MDH) structural gene from Methylobacterium organophilum XX was determined. A single open reading frame with a coding capacity of 626 amino acids (molecular weight, 66,000) was identified on one strand, and N-terminal sequencing of purified MDH revealed that 27 of these residues constituted a putative signal peptide. Primer extension mapping of in vivo transcripts indicated that the start of mRNA synthesis was 160 to 170 base pairs upstream of the ATG codon. Northern (RNA) blot analysis further demonstrated that the transcript was 2.1 kilobase pairs in length and therefore appeared to encode only MDH.

A yeast sigma composite element, TY3, has properties of a retrotransposon.

Sigma is a 340- or 341-base pair repetitive element which is located almost exclusively within 19 base pairs of the 5' ends of various tRNA genes in the Saccharomyces cerevisiae genome. Although most sigma elements characterized to date are isolated insertions, a few of the elements occur relatively closely spaced. One of these pairs is a direct repeat of the sigma element separated by an internal domain 4.7 kilobase pairs in length. Not only does this structure resemble a composite transposable element, but regions within the sigma elements and intervening domain are homologous to conserved regions in retroviruses and retrotransposons of yeast and other organisms. Two features suggest that the sigma elements and intervening DNA transposed in a concerted event: only one of the two sigma elements is associated with a tRNA gene, and only the outside ends of the two elements are flanked by the 5-base pair direct repeats that usually flank individual sigma insertions. Examination of genomic DNA from five laboratory strains indicates that the 4.7 kilobase pair internal domain is present in one to four copies per haploid genome and that the genomic location of this domain differs from strain to strain. In addition, Northern blot analysis showed the presence of a 5.2 kilobase poly(A) transcript which hybridizes to both sigma and internal domain-specific probes. The existence of this composite element may suggest new ways to consider the mechanisms by which retrotransposons select their targets.

Isolation and characterization of a temperate bacteriophage from Streptomyces galilaeus.

A new temperate actinophage from Streptomyces galilaeus ATCC 31133 was purified after that strain was crossed with S. peucetius ATCC 29050. Sensitive hosts became lysogenized and yielded turbid plaques of 2 to 3 mm in diameter. Host-range analysis indicated that 16 of 27 Streptomyces strains tested were sensitive to infection on solid medium. S. lividans and S. coelicolor A3(2) were among those not infected by this new actinophage. The new actinophage, designated phi SPK1, belongs to the Bradley group B morphological type, the pH optimum for infection is 6.75 to 7.0, it is not efficiently induced by mitomycin C or UV irradiation, it has a circular chromosome of 35.8 +/- 0.5 kilobase pairs in length containing overlapping (cohesive) ends, and the G+C content of its DNA was calculated from the buoyant density of 1.7240 to be 69 mol%. The DNA of phage phi SPK1 was cleaved by the restriction endonucleases ApaI, AluII, EcoRI, PvuII, and SalI, but, in all cases except that with EcoRI, treatment yielded greater than 20 restriction fragments. No sites were detected for BamHI, BclI, BglII, ClaI, HindIII, MluI, PstI, SmaI, SphI, SstI, XbaI, or XhoI.

Plasmid pCBI carries genes for anaerobic benzoate catabolism in Alcaligenes xylosoxidans subsp. denitrificans PN-1.

Pseudomonas sp. strain PN-1 is reclassified as Alcaligenes xylosoxidans subsp. denitrificans PN-1. Strain PN-1 is a gram-negative, rod-shaped organism, is motile by means of lateral flagella, is oxidase positive, and does not ferment sugars. Plasmid pCBI, carrying genes for the anaerobic degradation of benzoate in strain PN-1, is 17.4 kilobase pairs in length and is transmissible to a number of denitrifying Pseudomonas aeruginosa and Pseudomonas stutzeri strains. A restriction endonuclease map was constructed.

Initial catabolism of aromatic biogenic amines by Pseudomonas aeruginosa PAO: pathway description, mapping of mutations, and cloning of essential genes.

Pseudomonas aeruginosa PAO1 was able to utilize several aromatic biogenic amines as sole sources of carbon or nitrogen. These included the phenethylamines tyramine and dopamine and the phenethanolamines octopamine, synephrine, and norepinephrine. Initial catabolism of the phenethylamines was mediated by a membrane-bound tyramine dehydrogenase which produced 4-hydroxyphenylacetaldehyde (4HPAL) with tyramine as the substrate. The enzyme was induced by growth with both classes of amines. Initial catabolism of octopamine (except when present as the sole source of carbon and nitrogen) was mediated by a soluble enzyme with activity against the phenethanolamines but not against tyramine or dopamine. The product of the reaction with octopamine as substrate was also 4HPAL. Addition of NAD to reaction mixtures yielded 4-hydroxyphenylacetic acid and NADH. These activities, octopamine hydrolyase and 4-HPAL dehydrogenase (measured as a combined activity, OCAH-4HPALDH), were only induced by growth with phenethanolamines. However, the combined activities were not observed in extracts from cells grown with octopamine as the sole source of carbon and nitrogen, suggesting that an alternate pathway is used under this growth condition. Two independently isolated mutant strains were unable to utilize tyramine as a sole source of carbon or nitrogen. These mutants were also unable to utilize dopamine but grew at wild-type rates on the phenethanolamines. The mutations were mapped at about 70 min on the PAO1 chromosome with the chromosome-mobilizing plasmid R68.45, and both were linked to the catA1, mtu-9002, tyu-9009, and puuE mutations. DNA complementing both of the mutations was cloned on a single BamHI fragment approximately 13.8 kilobase pairs in length. Analysis of a subcloned fragment showed that the two mutations were in different genes.

Structural organization and chromosomal assignment of the parvalbumin gene.

The structure of the rat parvalbumin gene has been elucidated from analysis of six overlapping clones isolated from a rat lambda Charon 4A genomic library. Two of the clones were mapped in detail, and all exons were localized by Southern hybridization using fragments of a full-length parvalbumin cDNA (Epstein, P., Means, A. R., and Berchtold, M. W. (1986) J. Biol. Chem. 261, 5886-5891). The rat parvalbumin transcription unit is 15.5 kilobase pairs in length and contains four introns. The first intron divides the 5'-nontranslated region, whereas the other three interrupt coding DNA. All intron/extron boundaries were sequenced as was 377 base pairs immediately 5' from the putative transcription initiation site. The promoter region contains eukaryotic regulatory homologies to the "TATA" box at -24 and "CAAT" box at -47 and -156. In addition, two doublets consisting of 11-base pair direct repeats exist in the promoter region. Parvalbumin binds two Ca2+, whereas many other members of the same superfamily bind four. Comparison of the genes that encode these proteins provides a strong confirmation of the hypothesis that parvalbumin evolved from an ancestral gene specifying a four-domain Ca2+-binding protein. The rat parvalbumin gene was also utilized to assign its human counterpart to chromosome 22 from data obtained by hybridization to DNA from a somatic cell hybrid panel. It was also used to isolate a 7.5-kilobase pair EcoRI fragment from a human chromosome 22 DNA library.

Primary structure of the fifth component of murine complement

A cDNA library was constructed by the method of Okayama and Berg, [Okayama, H., & Berg, P. (1983) Mol. Cell. Biol. 3, 280-289] employing size-selected (greater than 28 S) poly(A+) liver RNA from the mouse strain B10.WR. A total of 150,000 recombinants were screened with a partial human C5 cDNA probe; 16 C5-positive clones were identified, 1 of which contained an insert greater than 5.2 kilobase pairs in length. This cDNA insert was fully sequenced by the dideoxy method. The DNA sequence of this insert had an open reading frame of 4920 base pairs specifying a sequence of 1640 amino acid residues. The region corresponding to positions 372-812 exhibited high homology with the previously determined partial structure for human C5 of 438 amino acid residues. A four-residue basic sequence (Arg-Ser-Lys-Arg) was identified upstream of the amino-terminal Asn of C5a, thereby specifying a beta alpha-chain orientation for the promolecule form of murine C5. The 3' end of this clone contained 351 base pairs of untranslated sequence. The presumed polyadenylation recognition site CATAAA was located 17 base pairs upstream of the poly(A) tail. Comparison of the derived murine C5 sequence with previously determined structures for murine C3 and C4 revealed regions of high sequence similarity, including the thiol ester region present in C3 and C4. The cysteine and proximal glutamine which give rise to the intramolecular thiol ester bond in C3 and C4 were absent in C5, having been replaced by serine and alanine, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Cloning of multiple genes involved with cobalamin (Vitamin B12) biosynthesis in Bacillus megaterium.

An effective shotgun cloning procedure was developed for Bacillus megaterium by amplifying gene libraries in Bacillus subtilis. This technique was useful in isolating at least 11 genes from B. megaterium which are involved with cobalamin (vitamin B12) biosynthesis. Amplified plasmid banks were transformed into protoplasts of both a series of Cob mutants blocked before the biosynthesis of cobinamide and Cbl mutants blocked in the conversion of cobinamide into cobalamin. Amplification of gene libraries overcame the cloning barriers inherent in the relatively low protoplast transformation frequency of B. megaterium. A family of plasmids was isolated by complementation of seven different Cob and Cbl mutants. Each plasmid capable of complementing a Cob or Cbl mutant was transformed into each one of the series of Cob and Cbl mutants; many of the plasmids isolated by complementation of one mutation carried genetic activity for complementation of other mutations. By these criteria, four different complementation groups were resolved. At least six genes involved in the biosynthesis of cobinamide are carried on a fragment of DNA approximately 2.7 kilobase pairs in length; other genes involved in the biosynthesis of cobinamide were located in two other complementation groups. The physical and genetic data permitted an ordering of genes within several of the complementation groups. The presence of complementing plasmids in mutants blocked in cobalamin synthesis resulted in restoration of cobalamin biosynthesis.

Cloning and expression in [formula omitted] of cDNA for arginase of rat liver

A cDNA expression library constructed in a plasmid pUC8 from poly(A) + RNA of rat liver was screened immunologically, using an antibody against arginase of rat liver. A cDNA clone was isolated and identified by hybrid-selected translation. The clone contained an insert approximately 1.35 kilobase pairs in length. In the bacterial clone, we detected a specific protein of M r = about 43000 that is slightly larger than the purified arginase ( M r = about 40000) and a high activity of arginase was expressed. The arginase mRNA species of about 1600 bases long was detected in the liver, but not in the small intestine, kidney, spleen and heart of the rats.

Chicken U2 and U1 RNA genes are found in very different genomic environments but have similar promoter structures.

We have cloned and analyzed a gene that codes for chicken U2 small nuclear RNA (snRNA). In the haploid chicken genome, there are approximately 35-40 copies of the U2 RNA gene arranged in tandemly repeated units 5.35 kilobase pairs in length. This U2 gene organization contrasts with that of chicken U1 RNA genes, which are found in heterogeneous genomic environments. Although U snRNA genes are transcribed by RNA polymerase II, they lack the usual TATA and CAAT homologies found in the 5' control regions of most RNA polymerase II transcription units. Nevertheless, a comparison of chicken U2 and U1 RNA gene 5'-flanking DNA sequences reveals two upstream blocks of homology which are also evolutionarily conserved in U2 and U1 RNA genes of other vertebrate species. The first block of conserved sequence is centered around position -55 relative to the RNA cap site, and the other is located near position -200. Interestingly, stretches of sequence with the potential to form Z DNA are located either within or immediately adjacent to both of these two conserved upstream sequence elements, suggesting a possible role for Z DNA in U1/U2 gene expression. Moreover, the chicken U2 and U1 gene promoter regions also contain specific short sequences (i.e., the hexamer GGGCGG and the octamer ATGCAAAT) that have been shown to be required for the expression of a number of mRNA-encoding genes. These findings suggest that the transcription of snRNA genes is controlled by a complex set of factors, some shared with other RNA polymerase II transcription units and others which may be unique to the snRNA genes.

Cloning of DNA fragments carrying hydrogenase genes of Rhodopseudomonas capsulata

A cosmid library of Rhodopseudomonas capsulata DNA was constructed in Escherichia coli HB101 using the broad-host-range cosmid vector pLAFR1. More than ninety per cent of the clones in the bank contained cosmids with DNA inserts averaging 20 kilobase pairs in length. Mutants deficient in uptake hydrogenase (Hup-) were obtained from R. capsulata strain B10 by ethylmethylsulfonate (EMS) mutagenesis. The content of hydrogenase protein in Hup- mutant cells was tested by rocket immunoelectrophoresis. Hup- mutants (Rifr) were complemented with the clone bank by conjugation and, from the transconjugants selected by rifampicin and tetracycline resistance, Hup+ transconjugants were screened for the ability to grow photoautotrophically and to reduce methylene blue in a colony assay. The recombinant plasmid pAC57 restored hydrogenase activity in the Hup- mutants RCC8, RCC10, RCC12 and ST410 whereas pAG202 restored that of IR4. The cloned R. capsulata DNA insert of pAC57 gave 5 restriction fragments by cleavage with EcoRI endonuclease. Fragment 1 (7 kb) restored hydrogenase activity in Hup- mutant strains RCC12 and ST410 and fragment 5 (1.3 kb) in strains RCC8 and RCC10. Since the 2 cosmids pAC57 and pAG202 are different cosmids, as indicated by restriction analyses and absence of cross hybridization, it is concluded that at least two hup genes are required for the expression of hydrogenase activity in R. capsulata.

Molecular cloning of rat mitochondrial glutamic oxaloacetic transaminase mRNA and regulation of its expression in regenerating liver

cDNA clones for rat mitochondrial glutamic oxaloacetic transaminase (mGOT) have been isolated from a rat liver cDNA library. One of the clones, designated p501, contained a cDNA insert of 1.4 kilobase pairs in length and hybridized to a mRNA of 2.4 kilobases from rat liver. We measured mGOT mRNA content in a regenerating rat liver. In a regenerating rat liver, mGOT activity was increased and reached maximum (170% of control activity) at about 48 h following the operation. Using the cDNA of mGOT, it was revealed that the increase of mGOT in the regenerating rat liver depended on its mRNA content.

The human gene encoding GM-CSF is at 5q21-q32, the chromosome region deleted in the 5q- anomaly.

Human granulocyte-macrophage colony-stimulating factor (GM-CSF) is a 22,000-dalton glycoprotein that stimulates the growth of myeloid progenitor cells and acts directly on mature neutrophils. A full-length complementary DNA clone encoding human GM-CSF was used as a probe to screen a human genomic library and isolate the gene encoding human GM-CSF. The human GM-CSF gene is approximately 2.5 kilobase pairs in length with at least three intervening sequences. The GM-CSF gene was localized by somatic cell hybrid analysis and in situ hybridization to human chromosome region 5q21-5q32, which is involved in interstitial deletions in the 5q- syndrome and acute myelogenous leukemia. An established, human promyelocytic leukemia cell line, HL60, contains a rearranged, partially deleted GM-CSF allele and a candidate 5q- marker chromosome, indicating that the truncated GM-CSF allele may reside at the rejoining point for the interstitial deletion on the HL60 marker chromosome.