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The tree shrew apolipoprotein C-I cDNA: sequence and its expression.

A rabbit anti-serum to tree shrew apolipoprotein C-I (apo C-I) was used to screen an expression cDNA library constructed by us from tree shrew (TS) liver tissue. Two apo C-I cDNA clones were obtained. The longer one consists of 380 nucleotides, including 21 bp and 95 bp at the 5' and 3' end of the non-translated regions respectively, and a 264-bp fragment in an open reading frame encoding 88 amino acids prepropeptide which contains 26 amino acids of signal peptide and a mature protein (62 amino acids). Comparing the amino-acid sequence deduced from this cDNA with those of the published mammalian apo C-Is reveals that it shared some structural similarity with rat, mouse and dog apo C-I, but it had 5 more amino acids than that of human and baboon. The expression of apo C-I mRNA in 8 different tissues were also assayed with Northern blot. The results demonstrated that liver had the highest expression, intestine had much less expression and no expression in other tissues, which is much different from human and other species. This study has laid down a good foundation for further studying on the function and the stucture of tree shrew apo C-I gene.

Amino Acid Sequence↗

Cloning and cDNA sequence analysis of myelin/oligodendrocyte glycoprotein: a novel member of the immunoglobulin gene superfamily.

Myelin/oligodendrocyte glycoprotein (MOG) is a CNS-specific protein that has been identified on the external myelin sheath and oligodendrocyte processes. MOG is the primary target autoantigen for demyelinating antibodies in experimental autoimmune encephalomyelitis, a widely used animal model for autoimmune demyelinating diseases such as multiple sclerosis. We have isolated a number of rat MOG cDNA clones as tools to begin studies to ascertain MOG function. A full-length cDNA clone (1.6 kb) was sequenced and the amino acid sequence for MOG deduced. This cDNA clone encodes a signal peptide of 27 amino acids, followed by 218 residues for mature MOG (24962 MW). A single site for N-glycosylation is found at Asn-31. The N-terminal half of mature MOG shares 52% identity with bovine butyrophilin, a possible lipid receptor, and 39% identity with chicken B-G antigen, a major histocompatibility complex antigen. This homology with Ig-like chicken MHC B-G antigens raises the issue of MOG involvement in autoimmune demyelination. Both MOG and butyrophilin meet criteria for inclusion in the immunoglobulin gene superfamily. Moreover, MOG appears to represent a novel member of this superfamily in that it possesses two potential transmembrane domains, in contrast to a single membrane-spanning domain or glycophospholipid anchor found in all other members of this superfamily. MOG mRNA expression is restricted to CNS tissue, and peak expression occurs during active myelination.

Age Factors↗

A juvenile hormone-repressible transferrin-like protein from the bean bug, Riptortus clavatus: cDNA sequence analysis and protein identification during diapause and vitellogenesis.

We found several juvenile hormone-responsive cDNAs in the bean bug, Riptortus clavatus, by using mRNA differential display (Hirai et al., 1998). One of them, a juvenile hormone-repressible cDNA, JR-3, was cloned, sequenced, characterized and identified as a transferrin (RcTf). RcTf cDNA encoded 652 amino acids with a calculated molecular weight of 71,453 Da. The deduced amino acid sequence showed significant homology with the transferin genes of several insects, Manduca sexta (43% identity), Blaberus discoidalis (43%), Aedes aegypti (43%), Drosophila melanogaster (36%), Sarcophaga peregrina (36%) and the human (25%). Antiserum was prepared by using recombinant RcTf protein expressed in Escherichia coli as an antigen. The antiserum reacted specifically with both the recombinant protein and the native protein from the bugs, with sizes of 70 and 75 kDa, respectively. The 75 kDa protein was partially purified from hemolymph of diapausing female bugs and the first ten amino acids were found to be identical to that of RcTf cDNA, indicating that the 75 kDa protein is RcTf. The tissue distribution of RcTf in the bug was examined by Western blot analysis. In diapausing animals, RcTf was detected in the fat body, hemolymph and ovary but not in the gut. In the post-diapause stage, RcTf was also detected in eggs, in addition to the fat body and ovary. These results indicate that RcTf is incorporated into the oocytes during vitellogenesis, and suggest that it may provide iron for the developing embryos.

Amino Acid Sequence↗

The isolation of cloned cDNA sequences which are differentially expressed in human lymphocytes and fibroblasts.

Poly(A)+ RNA populations derived from normal lymphocytes and fibroblasts have been compared by hybridising each RNA to cDNA derived from the other RNA population. This indicated that approximately 75% of the sequences were common to both, and that these were present at different concentrations in the two cell types. The two RNA populations were further compared by hybridising them to a cDNA recombinant library derived from lymphocyte poly(A)+ RNA. This allowed the identification of clones containing sequences which are abundant in lymphocyte poly(A)+ RNA but absent or rare in fibroblast poly(A)+ RNA. A direct estimation of the abundance of five of these sequences in lymphocyte cDNA demonstrated that clones can be detected by such a procedure if they represent 0.2% or greater of the original cDNA population.

Animals↗

A 11.5-kb 5'-terminal cDNA sequence of chicken breast muscle connectin/titin reveals its Z line binding region.

A partial 5'-region cDNA (11.5 kb) of chicken breast muscle connectin/titin encoding 3752 amino acids was sequenced. The predicted amino acid sequence contains 31 immunoglobulin C2 motifs and 10 interdomains. The sequence suggests a skeletal muscle type of connectin isoforms. Immunoelectron microscopic studies using antisera raised against several products of the cDNA fragments expressed in E. coli revealed that a region of some 800 amino acids from the N terminus of connectin is involved in its binding to the Z line in a sarcomere.

Animals↗

Human microtubule-associated protein 1a (MAP1A) gene: genomic organization, cDNA sequence, and developmental- and tissue-specific expression.

Microtubule-associated proteins (MAPs) regulate microtubule stability and play critical roles in neuronal development and the balance between neuronal plasticity and rigidity. MAP1a (HGMW-approved symbol MAP1A) stabilizes microtubules in postnatal axons. We describe human MAP1a's genomic organization and deduced cDNA and amino acid sequences. MAP1a is a single-copy gene spanning 10.5 kb. MAP1a coding sequence is contained in five exons. Translation begins in exon 3. Human MAP1a contains 2805 amino acids (predicted molecular weight 306.5 kDa) and is slightly larger than rat MAP1a (2774 amino acids). Like rat and bovine MAP1a, human MAP1a contains conserved tubulin binding motifs in the amino-terminal region. The carboxy-terminal portion contains a conserved pentadecapeptide that is present in the light chain portion of rat and bovine MAP1a/LC2 polyprotein. We show that human MAP1a gene expression occurs almost exclusively in the brain and that there is approximately 10-fold greater gene expression in adult brain compared to fetal brain. Strong, interspecies conservation between human and rat MAP1a cDNA and amino acid sequences indicates important relationships between MAP1a's function and its primary amino acid sequence.

Adult↗

Murine and human T11 (CD2) cDNA sequences suggest a common signal transduction mechanism.

The murine equivalent of the cDNA encoding the human T11 (CD2) sheep erythrocyte-binding protein has been cloned. It codes for a putative transmembrane protein which is homologous to human T11. In contrast to immunoglobulins whose domains consist of anti-parallel beta sheets, we predict that mouse and human T11 external domains probably belong to the alpha/beta protein folding class. The cytoplasmic region of T11 is a lengthy, proline-rich segment; secondary structural analysis predicts it to have a nonglobular conformation. This elongated tail could allow for interaction with multiple other intracellular proteins and may contain a cation-binding site involved in T lineage activation.

Amino Acid Sequence↗

Triticum aestivum puroindolines, two basic cystine-rich seed proteins: cDNA sequence analysis and developmental gene expression.

From a mid-maturation seed cDNA library we have isolated cDNA clones encoding two Triticum aestivum puroindolines. Puroindoline-a and puroindoline-b, which are 55% similar, are basic, cystine-rich and tryptophan-rich proteins. Puroindolines are synthesized as preproproteins which include N- and C-terminal propeptides which could be involved in their vacuolar localization. The mature proteins have a molecular mass of 13 kDa and a calculated isoelectric point greater than 10. A notable feature of the primary structure of puroindolines is the presence of a tryptophan-rich domain which also contains basic residues. A similar tryptophan-rich domain was found within an oat seed protein and a mammalian antimicrobial peptide. The ten cysteine residues of puroindolines are organized in a cysteine skeleton which shows similarity to the cysteine skeleton of other wheat seed cystine-rich proteins. Northern blot analysis showed that puroindoline genes are specifically expressed in T. aestivum developing seeds. No puroindoline transcripts as well as no related genes were detected in Triticum durum. The identity of puroindolines to wheat starch-granule associated proteins is discussed as well as the potential role of puroindolines in the plant defence mechanism.

Amino Acid Sequence↗

Molecular cloning and analysis of cDNA sequences derived from poly A+ RNA from barley endosperm: identification of B hordein related clones.

A collection of over 130 cDNA clones has been constructed in the bacterial plasmids pPH207 and pBR322 using as template the poly A+ RNA from membrane-bound polysomes of barley endosperm (cv. Sundance). Fifty four B hordein cDNA clones have been identified by cross-hybridization analysis and in vitro translation of plasmid-selected mRNAs. Hybridization of 11 of the B hordein cDNA clones to Northern blots of size-fractionated RNA indicated that the B hordein mRNA is ca. 1300 nucleotides long. One cDNA clone, pHvE-c16, has been partially sequenced and shown by comparison with C-terminal and other peptide sequences to be related to B1 hordein polypeptides. The results obtained from the analysis of the B hordein cDNA clones support the idea that the Hor 2 locus, which specifies the B hordeins, is complex and codes for a family of related mRNA species.

Base Sequence↗

Identification of the probable site of choleragen-catalyzed ADP-ribosylation in a Go alpha-like protein based on cDNA sequence.

Go alpha, a 39-kDa guanyl nucleotide-binding protein, is functionally and structurally similar to the alpha subunits of the stimulatory and inhibitory guanyl nucleotide-binding proteins (Gs alpha, Gi alpha) of adenylate cyclase and to the alpha subunit of transducin (T alpha), the guanyl nucleotide-binding protein of the retinal photon reception system. A cDNA clone was isolated from a bovine retinal lambda gt10 library by using oligonucleotide probes complementary to sequences in two putative T alpha clones. Partial sequence analysis revealed a deduced amino acid sequence identical to sequences of four tryptic peptides from bovine brain Go alpha. Gs alpha and T alpha are known to serve as substrates for ADP-ribosylation by choleragen. Other workers have established the sequence of the tetrapeptide in T alpha containing the arginine that is ADP-ribosylated and its location in the amino acid sequence deduced from T alpha cDNA. The Go alpha cDNA described here includes a region encoding an amino acid sequence very similar to that surrounding the ADP-ribosylation site in T alpha, consistent with observations that Go alpha can also be a substrate for choleragen. A corresponding sequence in the recently identified Gs alpha cDNA is less homologous to that in T alpha or Go alpha. The reported differences in conditions that promote choleragen-catalyzed ADP-ribosylation of Gs alpha vs. Go alpha could be related to differences in amino acid sequence in the region of the acceptor arginine.

Adenosine Diphosphate Ribose↗

The cDNA sequence encoding human protein kinase C-zeta.

A 1779-bp complementary DNA (cDNA) that encodes protein kinase C-zeta (PKC-zeta) has been isolated from a human frontal cortex library using traditional plaque-screening methods and PCR screening. The deduced 592-amino-acid sequence of the human PKC-zeta clone has a 95-96% identity to those deduced from the previously described rat and mouse PKC-zeta clones.

Amino Acid Sequence↗

Characterization of a cDNA sequence encoding the peroxisome proliferator activated receptor alpha in the chicken.

Peroxisome proliferator-activated receptors (PPAR) belong to the superfamily of the nuclear hormone receptors that play an important role in lipid homeostasis. A partial complementary deoxyribonucleic acid clone encoding a PPAR alpha from chicken liver was isolated and sequenced. Comparison with human, mouse, rat, and Xenopus PPAR alpha cDNA indicates a high degree of homology, especially at the level of the inferred peptide sequence (greater than 90%). The tissue pattern of expression indicates that PPAR alpha expression in the chicken is similar to that reported in other species, i.e., high expression levels in the liver, heart, and kidney, but also occurs in the lipogenic uropygial gland specific to birds.

Amino Acid Sequence↗

cDNA sequences of three kinds of beta-tubulins from rice.

Complete nucleotide sequences of three kinds of rice beta-tubulin cDNA clones (pTUB22, R1623 and R2242) were determined. Southern hybridization indicated that these beta-tubulins consist of one gene family. Using RFLP mapping, these three beta-tubulin cDNAs were mapped to different chromosomes indicating at least three loci for the beta-tubulin gene. The deduced amino acid sequences of these cDNAs showed a high similarity to other plant beta-tubulins. The asparagine residue located at the 100th amino acid from the N-terminus of plant beta-tubulins was also conserved with these three beta-tubulins. This asparagine is thought to be responsible for the sensitivity against rhizoxin, the toxin of the pathogen of rice seedling blight, Rhizopus sp. a soil-borne microorganism. Expression of the three beta-tubulin genes was analyzed by Northern blotting and all three clones were expressed in root, the possible target tissue of rhizoxin. These results suggest that these clones are candidates of beta-tubulins targeted by rhizoxin.

Amino Acid Sequence↗

Nuclear-encoded chloroplast ribosomal protein L27 of Nicotiana tabacum: cDNA sequence and analysis of mRNA and genes.

A tobacco (Nicotiana tabacum cv. Petite Havana) leaf cDNA library was constructed in the expression vector lambda gt11. Immunological and nucleic acid hybridization screening yielded several cDNAs encoding an M(r) 19,641 precursor to an M(r) 14,420 mature protein which is homologous to Escherichia coli ribosomal protein L27. One cDNA (L27-1; 882 nucleotides long) contains 104 bp of 5'-noncoding sequence, 51 codons for a transit peptide, 128 codons for the predicted mature L27 polypeptide, and 241 bp of 3'-noncoding sequence, including the poly(A)29 tail. A beta-galactosidase-L27 fusion protein was bound to nitrocellulose filters, expressed, and used as an affinity matrix to purify monospecific antibody to L27 protein from an antiserum of rabbits immunized with 50S chloroplast ribosomal proteins. Using this monospecific antibody, protein L27 was identified among HPLC-purified tobacco chloroplast ribosome 50S subunit proteins. The predicted amino terminus of the mature L27 protein was confirmed by partial sequencing of the HPLC-purified L27 protein. The mature L27 protein has 66%, 61%, 56%, and 48% amino acid sequence identity with the L27-type ribosomal proteins of Bacillus subtilis, E. coli, Bacillus stearo-thermophilus, and yeast mitochondria (MRP7), respectively, in the homologous overlapping regions. The transit peptide of tobacco chloroplast ribosomal protein L27 has 41% amino acid sequence similarity with the MRP7 mitochondrial targeting sequence. Tobacco chloroplast L27 protein also has a 40 amino acid long carboxyl-terminal extension (compared to its bacterial counterparts) which is similar to the corresponding portion of yeast MRP7.(ABSTRACT TRUNCATED AT 250 WORDS)

Amino Acid Sequence↗

cDNA sequence of the horse (Equus caballus) LAMA3 gene and characterization of two intronic SNP markers.

Laminins are large heterotrimeric basement membrane glycoproteins composed of alpha, beta and gamma chains. The Laminin 5 isoform has an alpha3beta3gamma2 composition and is essential for the adhesion of basal keratinocytes to the underlying epithelial basement membrane where it is mainly located. Mutations in the genes coding for the 3 chains have been associated with a severe skin blistering disease, Herlitz's junctional epidermolysis bullosa (JEB), observed in different species as man, dog, cat and horse. In this study, we report the sequence of the 5.2 kb horse laminin alpha 3 cDNA (LAMA3) as well as the detection of two intronic SNPs. These data will be useful to further identify causal mutations for the disease in this gene.

Amino Acid Sequence↗

Chromosome localization and cDNA sequence of murine and human genes for ras p21 GTPase activating protein (GAP).

The cDNA coding for mouse and human ras p21 GTPase-activating protein (GAP) was isolated; the deduced amino acid sequences share over 96% homology with that previously determined for bovine brain GAP. Both the mouse and human GAP cDNAs were used as probes for the chromosomal localization of this gene. The locus designations for the gene encoding GAP in human and mouse are RASA and Rasa (for ras-activating protein), respectively. By somatic cell hybrid analysis and in situ chromosomal hybridization, we have assigned the RASA gene to human chromosome band 5q13.3. In addition, with somatic cell genetics and linkage analysis in recombinant inbred mouse strains, the murine Rasa gene was localized to the distal end of mouse chromosome 13. These assignments place the gene encoding GAP in a known conserved syntenic region.

Amino Acid Sequence↗

Prokaryotic features of a nucleus-encoded enzyme. cDNA sequences for chloroplast and cytosolic glyceraldehyde-3-phosphate dehydrogenases from mustard (Sinapis alba).

Two cDNA clones, encoding cytosolic and chloroplast glyceraldehyde-3-phosphate dehydrogenases (GAPDH) from mustard (Sinapis alba), have been identified and sequenced. Comparison of the deduced amino acid sequences with one another and with the GAPDH sequences from animals, yeast and bacteria demonstrates that nucleus-encoded subunit A of chloroplast GAPDH is distinct from its cytosolic counterpart and the other eukaryotic sequences and relatively similar to the GAPDHs of thermophilic bacteria. These results are compatible with the hypothesis that the nuclear gene for subunit A of chloroplast GAPDH is of prokaryotic origin. They are in puzzling contrast with a previous publication demonstrating that Escherichia coli GAPDH is relatively similar to the eukaryotic enzymes [Eur. J. Biochem. 150, 61-66 (1985)].

Amino Acid Sequence↗

Evolution of tissue-specific keratins as deduced from novel cDNA sequences of the lungfish Protopterus aethiopicus.

Lungfishes are possibly the closest extant relatives of the land vertebrates (tetrapods). We report here the cDNA and predicted amino acid sequences of 13 different keratins (ten type I and three type II) of the lungfish Protopterus aethiopicus. These keratins include the orthologs of human K8 and K18. The lungfish keratins were also identified in tissue extracts using two-dimensional polyacrylamide gel electrophoresis, keratin blot binding assays and immunoblotting. The identified keratin spots were analyzed by peptide mass fingerprinting which assigned seven sequences (inclusively Protopterus K8 and K18) to their respective protein spot. The peptide mass fingerprints also revealed the fact that the major epidermal type I and type II keratins of this lungfish have not yet been sequenced. Nevertheless, phylogenetic trees constructed from multiple sequence alignments of keratins from lungfish and distantly related vertebrates such as lamprey, shark, trout, frog, and human reveal new insights into the evolution of K8 and K18, and unravel a variety of independent keratin radiation events.

Amino Acid Sequence↗