Search PubMed⌕ Search

SEARCH · Search PubMed

Results for “ENCODE”

Search indexed PubMed citations on genomics, clinical trials, systematic reviews and public health. Explore titles, authors and supplied subject terms, then open the PubMed record.

Quote a phrase for an exact phrase match. Source license links do not imply unrestricted reuse.

At least 1,369 records · Page 76Linked to original sources
PubMed2003

Genetic and phylogenetic characterization of the type II cyclobutane pyrimidine dimer photolyases encoded by Leporipoxviruses.

Shope fibroma virus and myxoma virus encode proteins predicted to be Type II photolyases. These are enzymes that catalyze light-dependent repair of cyclobutane pyrimidine dimers (CPDs). When the Shope fibroma virus S127L gene was expressed in an Escherichia coli strain lacking functional CPD repair pathways, the expressed gene protected the bacteria from 70-75% of the ultraviolet (UV) light-induced cytotoxic DNA damage. This proportion suggests that Leporipoxvirus photolyases can only repair CPDs, which typically comprise approximately 70% of the damage caused by short wavelength UV light. To test whether these enzymes can protect virus genomes from UV, we exposed virus suspensions to UV-C light followed by graded exposure to filtered visible light. Viruses encoding a deletion of the putative photolyase gene were unable to photoreactivate UV damage while this treatment again eliminated 70-90% of the lethal photoproducts in wild-type viruses. Western blotting detected photolyase protein in extracts prepared from purified virions and it can be deduced that the poxvirion interior must be fluid enough to permit diffusion of this approximately 50-kDa DNA-binding protein to the sites where it catalyzes photoreactivation. Photolyase promoters are difficult to categorize using bioinformatics methods, as they do not obviously resemble any of the known poxvirus promoter motifs. By fusing the SFV promoter to DNA encoding a luciferase open reading frame, the photolyase promoter was found to exhibit very weak late promoter activity. These data show that the genomes of Leporipoxviruses, similar to that of fowlpox virus, encode catalytically active photolyases. Phylogenetic studies also confirmed the monophyletic origin of poxviruses and suggest an ancient origin for these genes and perhaps poxviruses.

Animals↗
PubMed1995

Characterization of proteins encoded by ORFs 2 to 7 of Lelystad virus.

The genome of Lelystad virus (LV), a positive-strand RNA virus, is 15 kb in length and contains 8 open reading frames (ORFs) that encode putative viral proteins. ORFs 2 to 7 were cloned in plasmids downstream of the Sp6 RNA polymerase promoter, and the translation of transcripts generated in vitro yielded proteins that could be immunoprecipitated with porcine anti-LV serum. Synthetic polypeptides of 15 to 17 amino acids were selected from the amino acid sequences of ORFs 2 to 7 and antipeptide sera were raised in rabbits. Antisera that immunoprecipitated the in vitro translation products of ORFs 2 to 5 and 7 were obtained. Sera containing antibodies directed against peptides from ORFs 3 to 7 reacted positively with LV-infected alveolar lung macrophages in the immunoperoxidase monolayer assay. Using these antipeptide sera and porcine anti-LV serum, we identified three structural proteins and assigned their corresponding genes. Virions were found to contain a nucleocapsid protein of 15 kDa (N), an unglycosylated membrane protein of 18 kDa (M), and a glycosylated membrane protein of 25 kDa (E). The N protein is encoded by ORF7, the M protein is encoded by ORF6, and the E protein is encoded by ORF5. The E protein in virus particles contains one or two N-glycans that are resistant to endo-beta-N-acetyl-D-glucosaminidase H. This finding indicates that the high-mannose glycans are processed into complex glycans in the Golgi compartment. The protein composition of the LV virions further confirms that LV is evolutionarily related to equine arteritis virus, simian hemorrhagic fever virus, and lactate dehydrogenase-elevating virus.

Amino Acid Sequence↗
PubMed1998

Cloning and sequencing of a gene encoding a 2S albumin seed storage protein precursor from English walnut (Juglans regia), a major food allergen.

BACKGROUND: Walnuts rank third in per capita consumption of tree nuts in the United States and can be associated with systemic IgE-mediated reactions in some individuals. OBJECTIVE: The objectives of the study were to clone a gene encoding one of the major food allergens in the walnut kernel and to characterize the recombinant allergen. METHODS: A cDNA expression library in the lambda vector Uni-ZAP, which was prepared from walnut somatic embryos, was screened by using a patient's sera that reacted with multiple protein bands on immunoblotting. RESULTS: A cDNA clone containing an insert of 663 bp was identified and named Jug r 1. DNA sequence analysis of this clone revealed that it encoded a protein 142 amino acids in length. Comparison of the encoded protein sequences with protein databases revealed that this clone exhibits a 46.1% identity with the Brazil nut (Bertholletia excelsa) methionine-rich 2S albumin seed storage protein precursor, Ber e 1. Jug r 1 appears to be an important walnut food allergen; 12 of 16 sera from patients allergic to walnuts demonstrated IgE binding to the 2S albumin seed storage protein precursor fusion protein. An IgE-binding inhibition study suggests that the walnut 2S protein precursor undergoes posttranslational modification into a large and small subunit that is similar to castor seed, cottonseed, mustard seed, and Brazil nut 2S seed storage protein allergens. Interestingly, the gene encoding this allergenic protein in Brazil nuts has recently gained notoriety because of its experimental use as a transgene to enhance the nutritional quality of legumes. CONCLUSION: This is now the sixth definitive 2S albumin seed storage protein demonstrated to bind IgE, suggesting that this class of proteins is inherently allergenic.

2S Albumins, Plant↗
PubMed1999

Identification and cloning of a complementary DNA encoding a vicilin-like proprotein, jug r 2, from english walnut kernel (Juglans regia), a major food allergen.

BACKGROUND: Walnuts and other tree nuts are important food-allergen sources that have the potential to be associated with life-threatening, IgE-mediated systemic reactions in some individuals. OBJECTIVE: The purpose of this study was to characterize a complementary (c)DNA clone encoding one of the walnut food allergens. METHODS: A cDNA expression library prepared from walnut somatic embryo was screened for IgE reactivity with patient serum. A reactive clone of 2060 bp, which encoded a protein of 593 amino acids in length, was subcloned by excision into the pGEX expression vector. IgE-binding inhibition experiments were performed. RESULTS: A recombinant fusion protein was induced and shown to bind serum IgE from 9 of 15 patients tested, thus identifying a major allergen. This clone, named Jug r 2, exhibited significant homology with genes encoding the vicilin group of seed proteins. An IgE-binding inhibition experiment suggested that the encoded protein undergoes posttranslational modification into at least one major polypeptide (47 kd) and possibly several others, which is similar to the vicilin-like proteins characterized in cocoa bean (Theobroma cacao) and cottonseed (Gossypium hirsutum). N-terminal sequencing of the 47-kd band, Jug r 2, identified it as a mature protein obtained from the precursor. A second IgE-binding inhibition experiment showed that there is minimal or no cross-reactivity between Jug r 2 and pea vicilin, peanut proteins, or cacao proteins. CONCLUSION: Jug r 2 is the third vicilin food allergen identified in addition to vicilins from soy and peanut. The availability of recombinant food allergens should help advance studies on the immunopathogenesis and possible treatment of IgE-mediated food hypersensitivity.

Allergens↗
PubMedDate not supplied

The role of specificity in the lexical encoding of participants.

In addition to information about phonology, morphology and syntax, lexical entries contain semantic information about participants (e.g., Agent). However, the traditional criteria for determining how much participant information is lexically encoded have proved unreliable. We have proposed two semantic criteria (obligatoriness and selectivity) that jointly identify the participants that are lexically encoded in verbs. We tested whether one of these criteria, semantic selectivity, makes psychologically real distinctions between participant information that is lexically encoded and participant information that is not. We examined how readers integrated syntactically optional WH-constituents in filler-gap sentences when the participant information conveyed by the WH-filler was specific to a restricted class of verbs (i.e., source locations) and when it was not (i.e., event locations). Our results provide support for the role of specificity in the lexical encoding of participant information of syntactically optional constituents.

Cognition↗
PubMed1997

Identification and characterization of a human Vlambda5 (T1) germline gene that encodes structurally unique lambda light chains.

The human germline Vlambda repertoire consists of about 30 functional genes that have been classified into 10 families on the basis of homologies in nucleotide sequences that encode approximately the first 96 to 104 residues of lambda light chains. One family, termed Vlambda5, is of special interest because the lambda light chain products of these genes have unique structural features. We have now isolated from genomic DNA one member of this family, designated IGLV5-1, using as a molecular probe a partial Vlambda5-germline-gene fragment generated by polymerase chain reaction. IGLV5-1 contains all the requisite elements of a potentially functional gene, including a Vlambda exon with an open reading frame specifying 104 residues. A Vlambda5-related cDNA (ZW) was also cloned from a bone marrow-derived plasma-cell population obtained from a patient with light-chain-associated (AL) amyloidosis. Comparison of the predicted protein sequences encoded by the IGLV5-1-germline gene, cDNA ZW, and three other reported Vlambda5-related cDNAs with those of the deduced or expressed products of the other nine known human Vlambda-gene families revealed that Vlambda5 proteins contain distinctive primary structural features. These include the presence within the second complementarity determining region (CDR2) and the third framework region (FR3) of 11 and 34 amino acids, respectively, rather than the 7 and 32 that occur in the most commonly expressed Vlambda1-, Vlambda2- and Vlambda3-type light chains. Although certain of the Vlambda-gene families encode either an elongated CDR2 or FR3, Vlambda5 proteins are remarkable in that they have additional residues in both regions of the molecule. In this respect, these polypeptides are most similar to surrogate light-chain-associated human and mouse VpreB components that also have these unusual primary structural features. Further, the four additional CDR2 residues and the two-residue FR3 insertion have been found among lambda-type light chains of certain non-mammalian species. The evolutionarily conserved nature of human Vlambda5-related genes and, in particular, the presumably novel tertiary structural effects induced by the unique features of the lambda light chains encoded by these elements suggest that the Vlambda5-gene family has biological and functional importance.

Amino Acid Sequence↗
PubMed2001

Cloning and sequencing of a cDNA encoding the bovine FcR gamma chain.

Phagocytic cells of the immune system express specific receptors for the Fc region of immunoglobulins (FcRs). In humans, most FcRs for IgG (FcgammaR), IgA (FcalphaR) and IgE (FcvarepsilonR) consist of an immunoglobulin (Ig) -binding subunit associated with a specialized signaling molecule, the FcR gamma chain. The FcR gamma chain is crucial for the transmission of intracellular signals following receptor ligation. In cattle, however, although four distinct complimentary DNAs (cDNAs) encoding IgG-binding subunits have been described (corresponding to bovine FcgammaRI, FcgammaRII, FcgammaRIII, and Fcgamma2R), virtually, nothing is known about signal transduction via bovine FcRs. Therefore, in this study, a cDNA encoding the bovine FcR gamma chain was cloned. The cDNA is 258 base pairs long and encodes a protein of 85 amino-acids. The mature protein shows high homology with the FcR gamma chains from several other species. Interestingly, the cytoplasmic domain of the bovine FcR gamma chain is one amino-acid shorter than those previously described. Cloning of a cDNA encoding, the bovine FcR gamma chain will allow for a better understanding of signal transduction processes triggered by bovine FcRs.

Amino Acid Sequence↗
PubMed2001

Production of polyclonal and monoclonal antibodies against CD54 molecules by intrasplenic immunization of plasmid DNA encoding CD54 protein.

DNA immunization, in theory, is of great interest as a source of specific antibodies against different antigens. In an attempt to produce polyclonal and monoclonal antibodies against cell surface molecules by using the DNA immunization strategy, intramuscular and intrasplenic routes of DNA injection were compared. Two to five, but not a single, intramuscular DNA immunizations induced anti-CD54 and anti-CD147 antibody production. In contrast, a single intrasplenic immunization of CD54-encoding DNA could induce anti-CD54 antibody production. To produce monoclonal antibody (mAb), spleen cells obtained from an intrasplenic CD54-encoding DNA immunized mouse were fused with myeloma cells using the standard hybridoma technique. A hybridoma secreting specific mAb to CD54 was established. The generated mAb reacted to CD54 protein expressed on transfected COS cells and various cell types, the same as using standard CD54 mAb MEM-111. Our results demonstrated that direct immunization of antigen-encoding DNA into spleen is an effective route for production of both polyclonal and monoclonal antibodies to cell surface molecules. This finding is very useful for the production of antibodies to cell surface molecules where the protein antigen is not available or difficult to prepare, but cDNA encoding the corresponding protein is available.

Animals↗
PubMed2001

Sequence and phylogenetic analysis of the sigmaA-encoding gene of avian reovirus.

The full-length sigmaA-encoding gene nucleotide sequences (1251 bp) of ten avian reovirus (ARV) field-isolates and three vaccine strains were determined and analyzed to study the degree of genetic divergence and evolution. Strains were isolated over a 23-year period from different hosts, pathotypes, and geographic locations. A phylogenetic tree constructed from variation in the sigmaA nucleotide sequences among ARV isolates showed that Taiwanese isolates from different dates of isolation were grouped into two distant groups, indicating that they have evolved in nature. In paired identity analysis, there was over 97.3% nucleotide sequence identity in the sigmaA-encoding genes between group I Taiwanese isolates (T6, 750505, 919, and 918) and Japanese isolate OS161 as well as three US vaccine strains, suggesting that they might have descended from a common ancestor. However, the nucleotide sequences of these sigmaA-encoding genes varied extensively from those of group II Taiwanese isolates (601SI, R2/TW, 1017-1, 916, and 601G), displaying only 86% identity. These results revealed that the genetic diversity in the sigmaA-encoding gene of ARV correlated with the date of isolation and geographic locations.

Amino Acid Sequence↗
PubMed1999

Identification of the sigma C-encoded gene of avian reovirus by nested PCR and restriction endonuclease analysis.

A nested reverse transcription (RT)-polymerase chain reaction with subsequent restriction endonuclease analysis was developed for identification of the sigma C-encoded gene of avian reoviruses (ARV). PCR products derived from the sigma C-encoded gene of all tested ARVs resulted in a specific DNA band of 1023 bp, indicating that there were no apparent insertions or deletions in this region. Amplification with the nested primer pairs S1M-S1N and S1P-S1N generated 330 and 239 bp, respectively. PCR products amplified from the sigma C-encoded of all tested ARVs isolates were further confirmed by Southern blot hybridization and restriction endonuclease analysis. PCR amplified cDNA fragment (1023 bp) cleaved with Pst I generated two fragments of 565 and 458 bp. The amplified sigma C-encoded gene of ARV was subcloned into PQE 32 vector for further study of its antigenicity and immunogenicity. The sensitivity of RT-PCR was examined on nucleic acids from the ARV infected cell cultures. The detection limit was 10(0) to 10(-1) TCID50 of ARV in a ethidium bromide stained gel and could be increased further to 10(-1) to 10(-2) TCID50 of ARV by Southern blot hybridization using a digoxigenin-labeled cDNA probe. The sensitivity increased approximately 10(3) to 10(4) folds when the cDNA was reamplified with two sets of nested primers.

Animals↗
PubMed2002

Orofacial mechanoreceptors in humans: encoding characteristics and responses during natural orofacial behaviors.

We used microneurography to characterize stimulus-encoding properties of low-threshold mechanoreceptive afferents in human orofacial tissues. Signals were recorded from single afferents in the infraorbital, lingual and inferior alveolar nerves while localized, controlled, mechanical stimuli were delivered to the facial skin, lips, oral mucosa and teeth. We likewise analyzed activity in these afferents during orofacial behaviors such as speech, chewing and biting. The afferents in the soft tissues functionally resemble four types described in the human hand: hair follicle afferents, slowly adapting (SA) type I and type II afferents and fast adapting (FA) type I afferents. Afferents in the facial skin, lips and buccal mucosa respond not only to contact with environmental objects, but also to contact between the lips, changes in air pressure generated for speech sounds, and to facial skin and mucosa deformations that accompany lip and jaw movements associated with chewing and swallowing. Hence, in addition to exteroceptive information, these afferents provide proprioceptive information. In contrast, afferents terminating superficially in the tongue do not signal proprioceptive information about tongue movements in this manner. They only respond when the receptive field is brought into contact with other intraoral structures or objects, e.g. the teeth or food. All human periodontal afferents adapt slowly to maintained tooth loads. Populations of periodontal afferents encode information about both which teeth are loaded and the direction of forces applied to individual teeth. Most afferents exhibit a markedly curved relationship between discharge rate and force amplitude, featuring the highest sensitivity to changes in tooth load at low forces (below 1 N). Accordingly, periodontal afferents efficiently encode tooth load when subjects first contact, hold, and gently manipulate food by the teeth. In contrast, only a minority of the afferents encodes the rapid and strong force increase generated when biting through food. We conclude, that humans use periodontal afferent signals to control jaw actions associated with intraoral manipulation of food rather than exertion of jaw power actions.

Behavior↗
PubMed1997

Cloning and expression of a cDNA encoding a nonintegrin laminin-binding protein from Echinococcus granulosus with localization of the laminin-binding domain.

We have isolated a cDNA from the hydatid tapeworm, Echinococcus granulosus, encoding a protein that binds laminin. This is the first report of a helminth parasite laminin-binding protein and the first description of a cDNA encoding a laminin-binding protein from a parasite. The cDNA clone (egmo3) was isolated from an E. granulosus protoscolex cDNA expression library, and identified on the basis of sequence homology to the nonintegrin mammalian metastasis-associated 67-kDa laminin receptor (67-LR). The amino acid sequence predicted from the cDNA sequence is 268 residues long with a calculated molecular mass of 29.9 kDa. Southern blot analysis suggested that many copies of the gene may occur in the E. granulosus genome. A Northern blot revealed that the gene is expressed as a single transcript of approximately 1 kb consistent with the size of the cDNA insert. Antibodies raised to the purified protein interacted with a 30 kDa protein in whole E. granulosus protoscoleces. A Western blot of the purified and refolded recombinant protein specifically bound 125I-labelled laminin, as did a synthetic peptide derived from the inferred amino acid sequence of egmo3 which is similar in homology to peptide G, the active ligand-binding site of 67-LR. We also isolated the 3' end of the cDNA encoding the homologous protein from the closely related species, E. multilocularis. The polypeptide encoded by egmo3 also shares substantial identity with the acidic class of ribosomal proteins which are involved in protein synthesis. As such, the egmo3 protein may be multifunctional in E. granulosus, acting as a laminin-binding molecule but also playing a role in cell division and growth.

Amino Acid Sequence↗
PubMed2000

Long-term differential modulation of genes encoding orexigenic and anorexigenic peptides by leptin delivered by rAAV vector in ob/ob mice. Relationship with body weight change.

We investigated the long-term effects of physiological levels of leptin produced by gene therapy on body weight (BW) and expression of genes that encode orexigenic and anorexigenic peptides in the hypothalamus. Recombinant adeno-associated viral vector (rAAV), a non-pathogenic and non-immunogenic vector, encoding leptin (betaOb) was generated and administered iv to ob/ob mice lacking endogenous leptin. Whereas the lowest dose of rAAV-betaOb (6x10(9) particles) was ineffective, the middle dose (6x10(10) particles) curbed BW gain without affecting food consumption for 75 days of observation. A ten-fold higher dose (6x10(11) particles) resulted in increased blood leptin levels and suppressed both BW gain and food consumption throughout the duration of the experiment. rAAV-betaOb doses that either curbed BW without affecting food consumption or evoked BW loss and reduced food intake, decreased the expression of genes encoding the orexigenic peptides, neuropeptide Y and agouti-related peptide in the ARC, and the two doses were equally effective. Concomitantly, the expression of genes encoding the anorexigenic peptide, alpha-melanocyte stimulating hormone and cocaine-and-amphetamine regulatory transcript, was augmented with the latter gene displaying a dose-dependant response. These results document the efficacy of delivering biologically active leptin for extended periods by an iv injection of rAAV-betaOb and show that physiological leptin concentrations simultaneously exert a tonic inhibitory effect on orexigenic and a stimulatory effect on anorexigenic signaling in the hypothalamus. This intricate dynamic interplay induced by leptin regulates BW with or without an effect on food intake in leptin-deficient ob/ob mice. Further, these results suggest that gene therapy is an effective mode of delivery to the hypothalamus of those therapeutic proteins that cross the blood-brain barrier to ameliorate neuroendocrine disorders.

Agouti-Related Protein↗
PubMed2001

Characterization and regulation of Schizosaccharomyces pombe gene encoding thioredoxin.

A cDNA coding thioredoxin (TRX) was isolated from a cDNA library of Schizosaccharomyces pombe by colony hybridization. The 438 bp EcoRI fragment, which was detected by Southern hybridization, reveals an open reading frame which encodes a protein of 103 amino acids. The genomic DNA encoding TRX was also isolated from S. pombe chromosomal DNA using PCR. The cloned sequence contains 1795 bp and encodes a protein of 103 amino acids. However, the C-terminal region obtained from the cDNA clone is -Val-Arg-Leu-Asn-Arg-Ser-Leu, whereas the C-terminal region deduced from the genomic DNA appears to contain -Ala-Ser-Ile-Lys-Ala-Asn-Leu. This indicates that S. pombe cells contain two kinds of TRX genes which have dissimilar amino acid sequences only at the C-terminal regions. The heterologous TRX 1C produced from the cDNA clone could be used as a subunit of T7 DNA polymerase, while the TRX 1G from the genomic DNA could not. The upstream sequence and the region encoding the N-terminal 18 amino acids of the genomic DNA were fused into the promoterless beta-galactosidase gene of the shuttle vector YEp357 to generate the fusion plasmid pYKT24. Synthesis of beta-galactosidase from the fusion plasmid was found to be enhanced by hydrogen peroxide, menadione and aluminum chloride. It indicates that the expression of the cloned TRX gene is induced by oxidative stress.

Aluminum Chloride↗
PubMed1998

Characterization and functional expression of the cDNA encoding human brain quinolinate phosphoribosyltransferase.

Mammalian quinolinate phosphoribosyltransferase (QPRTase) (EC 2.4.2.19) is a key enzyme in catabolism of quinolinate, an intermediate in the tryptophan-nicotinamide adenine dinucleotide (NAD) pathway. Quinolinate acts as a most potent endogenous exitotoxin to neurons. Elevation of quinolinate levels in the brain has been linked to the pathogenesis of neurodegenerative disorders. As the first step to elucidate molecular basis underlying the quinolinate metabolism, the cDNA encoding human brain QPRTase was cloned and characterized. Utilizing partial amino acid sequences obtained from highly purified porcine kidney QPRTase, a human isolog was obtained from a human brain cDNA library. The cDNA encodes a open reading frame of 297 amino acids, and shares 30 to 40% identity with those of bacterial QPRTases. To confirm that the cDNA clone encodes human QPRTase, its functional expression was studied in a bacterial host. Introduction of the human cDNA into a QPRTase defective (nadC) E. coli strain brought about an abrupt increase in QPRTase activity and allowed the cells to grow in the absence of nicotinic acid. It is concluded that the cloned cDNA encodes human QPRTase which is functional beyond the phylogenic boundary.

Amino Acid Sequence↗
PubMed1999

RT-PCR cloning, characterization and mRNA expression analysis of a cDNA encoding a type II asparagine synthetase in common bean.

Following a RT-PCR strategy based on the design of degenerate oligonucleotides resembling conserved domains of asparagine synthetase (AS; EC 6.3.5.4), we isolated a 2 kb cDNA clone (PVAS2) from root tissue of the common bean (Phaseolus vulgaris). PVAS2 encodes a protein of 584 amino acids with a predicted relative molecular mass of 65810 Da, an isoelectric point of 6.4, and a net charge of -7.2 at pH 7.0. The amino acid sequence of the protein encoded by PVAS2 is very similar to that encoded by the soybean SAS2 asparagine synthetase gene. The amino-terminal residues of the predicted PVAS2 protein are identical to the amino acids that constitute the glutamine-binding (GAT) domain of AS from other plant species, which suggests that the PVAS2 cDNA encodes a type II glutamine-dependent form of asparagine synthetase. Southern blot analysis indicates that the common bean AS is part of a small family composed of at least two genes. Expression analysis by Northern blot revealed that the PVAS2 transcript accumulates to a high level in roots and, to a lesser extent, in nodules and developing pods. Accumulation of the PVAS2 transcript in the root seems to be negatively regulated by light and sucrose, and positively regulated by nitrate.

Amino Acid Sequence↗
PubMed1998

Regulated expression during development and following sciatic nerve injury of mRNAs encoding the receptor tyrosine phosphatase HPTPzeta/RPTPbeta.

Three major isoforms of the receptor protein tyrosine phosphatase HPTPzeta/RPTPbeta (RPTPzeta/beta) have been previously identified, two with identical transmembrane and intracellular catalytic domains that differ by virtue of a long cysteine-free extracellular region, and a soluble proteoglycan called phosphacan that lacks the transmembrane and carboxy-terminal catalytic domains. To determine whether these RPTPzeta/beta variants are produced by alternative mRNA splicing of a common primary transcript, we performed genomic Southern analysis and characterized several rat cDNA and genomic RPTPzeta/beta clones. These studies indicated that the three major transcripts which encode phosphacan and the two RPTPzeta/beta phosphatase variants are encoded by a single gene, and further that additional alternative mRNA splicing is likely to result in the deletion of a 7 amino acid insert from the intracellular juxtamembrane region of both long and short phosphatase isoforms. Simultaneous quantitation of the three major isoforms by RNase protection analysis indicated that the mRNA encoding phosphacan had the highest relative abundance in the CNS while that encoding the short phosphatase isoform was most abundant relative to the other RPTPzeta/beta variants in the PNS. Following peripheral nerve crush, all RPTPzeta/beta mRNAs, including phosphacan and the phosphatase variants with and without the 21 base insert, were significantly induced in the distal segments of the sciatic nerve with a time course that correlated well with the response of Schwann cells to this injury.

Alternative Splicing↗
PubMed2001

Immunization with plasmid DNA encoding the integral membrane protein, Sm23, elicits a protective immune response against schistosome infection in mice.

Schistosomes are helminth parasites infecting at least 200 million people worldwide. In this study, we evaluated the feasibility of using a nucleic acid vaccine to induce protective immune responses to the Schistosoma mansoni integral membrane protein Sm23. C57BL/6 mice were immunized by intramuscular injection in three separate vaccination trials. ELISA and Western Blot analyses indicated that mice immunized with a DNA plasmid construct encoding Sm23 (Sm23-pcDNA) generated specific IgG for Sm23, while sera from mice immunized with the control pcDNA plasmid did not. The vaccine elicited IgG(2a), and IgG(1) antibody isotypes. We also tested the adjuvant activity of IL-12 and IL-4 on humoral responses to Sm23. Co-immunization with plasmid encoding IL-12 did not affect the level of anti-Sm23 IgG(2a), but did reduce the IgG(1) level. In contrast, co-injection with a plasmid encoding IL-4 significantly reduced the level of anti-Sm23 IgG(2a), while the level of IgG(1) was largely unchanged. Importantly, the Sm23-pcDNA vaccine provided statistically significant levels of protection against challenge infection (21-44%, P<0.001-0.02). Co-administration of plasmids encoding either IL-12 or IL-4 did not significantly enhance this protective effect.

Animals↗