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S Yarus

Publications and source records attributed to S Yarus.

12 recordsLinked to original sources
PubMed1997

The carboxy-terminal domain of human surfactant protein B is not required for secretion in milk of transgenic mice.

Previous studies in which human pulmonary surfactant protein B (SP-B) was targeted to the mammary gland of transgenic mice using the rat whey acidic protein (WAP) regulatory sequences resulted in secretion of only the unprocessed proprotein (42 kDa) in milk. To test the feasibility of producing a partially processed SP-B protein in milk, a new construct was designed in which the coding region for the carboxy-terminal domain was deleted. Expression of rWAP/SP-BDELTA C mRNA was detected in all three transgenic lines generated, and the expected carboxy-terminal deleted SP-B molecule (28 kDa), identified by using domain-specific antibodies, was secreted in the milk. Histochemical examination of lactating mammary tissue from the transgenic line expressing the highest levels of WAP/SP-BDelta C mRNA revealed an inhibition of lobulo-alveolar development, and led to growth retardation in pups, apparently due to the decreased milk production. Mothers from this line tended to cannibalize litters in mid-lactation. This phenotype has been observed previously with several other WAP-based transgenes. This phenotype suggests that there may be an upper limit to the level of SP-BDeltAC which can be produced in milk

Animals↗
PubMed1997

Secretion of unprocessed human surfactant protein B in milk of transgenic mice.

Because of the apparent clinical importance of human pulmonary surfactant B (SP-B), the expression of SP-B was directed to the mammary gland of transgenic mice using previously characterized rat whey acidic protein (WAP) regulatory sequences. rWAP/SP-B mRNA was expressed specifically in the mammary gland, and ranged from 1 to 5% of the endogenous WAP mRNA levels. SP-B was detected immunologically in both tissue and milk. The transgene product had an apparent molecular weight of 40-45 kDa, corresponding to the predicted size of the SP-B proprotein. Incubation of an SP-B-enriched fraction of milk with cathepsin D in vitro produced 20-25 kDa species, consistent with cleavage of the amino terminal domain by cathepsin D. This was confirmed using antibodies specific to the carboxy-terminal domain of SP-B. However, the appearance of only the SP-B proprotein in milk suggests that cathepsin D is not involved in the in vivo processing of SP-B. The SP-B proprotein in milk suggests that cathepsin D is not involved in the in vivo processing of SP-B. The SP-B proprotein can be expressed in milk of transgenic mice without any observed effects on mammary gland morphology or lactation.

Animals↗
PubMed1996

Production of active bovine tracheal antimicrobial peptide in milk of transgenic mice.

Tracheal antimicrobial peptide (TAP) is a member of the beta-defensin family of antibiotic peptides found in the tracheal mucosa of the cow. TAP gene expression in the bovine airway is inducible by lipopolysaccharide and inflammatory mediators, suggesting that it functions to protect the upper airway from infection. Limited availability of bovine TAP (bTAP) has precluded investigation of its potential utility in agriculture and medicine. To overcome this problem, transgenic mice expressing bTAP using an expression vector driven by control sequences from the murine whey acidic protein (WAP) gene have been generated. The WAP/bTAP transcript was detected in RNA isolated from mammary tissue of transgenic females. bTAP was purified to homogeneity from milk via acid precipitation, reverse-phase HPLC, and ion-exchange chromatography. This milk-derived bTAP had antimicrobial activity against Escherichia coli. Amino-terminal peptide sequencing confirmed the identity of this material as a bTAP isoform. bTAP available from a mammary gland bioreactor will allow evaluation of bTAP for use as an antibiotic in agriculture and medicine.

Amino Acid Sequence↗
PubMed1995

Production of human surfactant protein C in milk of transgenic mice.

Respiratory distress syndrome (RDS), caused by lack of pulmonary surfactant, affects 65 000 infants annually in the USA. Surfactant replacement therapy reduces the morbidity and mortality associated with RDS. Human surfactant protein C (SP-C) is an important component of pulmonary surfactant. To produce human SP-C, a construct using the rat whey acidic protein (WAP) promoter and 3' untranslated regions to target expression of the human SP-C gene to the mammary gland of transgenic mice was created. WAP/SP-C mRNA expression was detected in all transgenic lines analysed. SP-C was expressed in a copy-number-dependent and integration-site-independent fashion, with levels of expression ranging from 0.01% to 36.0% of the endogenous mouse WAP mRNA, and WAP/SP-C mRNA expression levels were greater than those of of the endogenous mouse lung SP-C mRNA. Expression at the RNA level was specific to the mammary gland and paralleled the endogenous WAP expression pattern during mammary gland development. Expression and secretion of the SP-C protein in the lactating mammary gland was demonstrated by western blots performed on whole milk using an anti-SP-C polyclonal antibody. Immunoreactive proteins of MW 22 and 12-14 kDa appeared only in transgenic milk. The 22 kDa protein represents the proprotein, and the 12-14 kDa is a processed form of SP-C.

Animals↗
PubMed1993

Isolation and characterization of target sequences of the chicken CdxA homeobox gene.

The DNA binding specificity of the chicken homeodomain protein CDXA was studied. Using a CDXA-glutathione-S-transferase fusion protein, DNA fragments containing the binding site for this protein were isolated. The sources of DNA were oligonucleotides with random sequence and chicken genomic DNA. The DNA fragments isolated were sequenced and tested in DNA binding assays. Sequencing revealed that most DNA fragments are AT rich which is a common feature of homeodomain binding sites. By electrophoretic mobility shift assays it was shown that the different target sequences isolated bind to the CDXA protein with different affinities. The specific sequences bound by the CDXA protein in the genomic fragments isolated, were determined by DNase I footprinting. From the footprinted sequences, the CDXA consensus binding site was determined. The CDXA protein binds the consensus sequence A, A/T, T, A/T, A, T, A/G. The CAUDAL binding site in the ftz promoter is also included in this consensus sequence. When tested, some of the genomic target sequences were capable of enhancing the transcriptional activity of reporter plasmids when introduced into CDXA expressing cells. This study determined the DNA sequence specificity of the CDXA protein and it also shows that this protein can further activate transcription in cells in culture.

Amino Acid Sequence↗
PubMed1991

Genomic organization and expression during embryogenesis of the chicken CR1 repeat.

CR1 is one of the middle repetitive sequence elements present in the chicken genome. One such repetitive element (GG1-CR1) was found upstream of the chicken CHox E homeobox gene. Sequencing of GG1-CR1 demonstrated that it is one of the longest CR1 elements analyzed. Detailed comparison of all the CR1 sequences published has revealed three subfamilies of CR1 repeats containing various parts of the consensus sequence. We prepared DNA fragments from GG1-CR1 and used them to probe Southern blots and genomic and cDNA library lifts. The results confirm the division of CR1 into three subelements, two of which occur independently in many places in the genome. Northern blot analysis of the CR1 to chicken embryo RNA showed that the CR1 repeat can be part of poly(A)+ transcripts. These results suggest that the CR1 can be transcribed by readthrough from the promoter of the neighboring gene without detrimental effects on the expression of the gene itself. The level of CR1 containing transcripts rises during the first 5 days of embryonic development and then decreases.

Animals↗
PubMed1990

Passive immunization of chickens against Eimeria maxima infection with a monoclonal antibody developed against a gametocyte antigen.

Eimeria maxima gametocytes contain two major antigens with molecular masses of 56 and 82 kilodaltons (kDa) which are recognized by convalescent sera from immune chickens. Preparations enriched in these two antigens were used to immunize mice, and several monoclonal antibodies which specifically reacted with the 56-kDa antigen were produced. One of these monoclonal antibodies of the immunoglobulin M subclass, along with immune chicken sera raised against affinity-purified 56- and 82-kDa antigens, was used to passively immunize chicks. On the basis of the parameter of total oocyst output, it was found that these antibodies provided partial protection (40 to 50% inhibition) against E. maxima challenge infections.

Animals↗
PubMed1989

Eimeria maxima: identification of gametocyte protein antigens.

The antigenicity of Eimeria maxima gametocyte proteins during the course of an infection and when injected into mice and rabbits was demonstrated using the Western blotting technique. Serum taken from chickens at various times postinfection reacted to a few gametocyte proteins, with the strongest reactivity seen with serum taken 14-days postinfection. Two major antigens having molecular weights of 56,000 and 82,000 were consistently detected by these sera. Using immune rabbit or mouse sera to whole gametocyte detergent extracts, the 56,000 and 82,000 molecular weight proteins were again the immunodominant antigens, despite their representing only a small proportion of the extract which was used to immunize the animals. These results, together with those obtained by Rose (1971) using recovered chicken serum to passively immunize chickens, indicate that these two gametocyte antigens may play a role in protective immunity to E. maxima.

Animals↗