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The influence of DctA and DctB on the regulation of the dctA, dctB and dctD promoters was analyzed. LacZ fusion analysis was used to determine transcription in dctB and dctD. The result showed that both have only one promoter. Nitrogen limitation which induces the dctA promoter requires DctB-DctD and NtrC. DctA controls signal detection by DctB, thus the ligand specificity of DctB is wider when there is a lack of DctA. This suggests that DctA has an important role in the signal specificity of DctB-DctD.

Amino acid uptake by the general amino acid permease (Aap) of Rhizobium leguminosarum strain 3841 was greatly reduced by the presence of aspartate in the growth medium when glucose was the carbon source. This was due to the prevention of uptake and not by transcriptional repression. However, the Ntr-regulated gene glnII was repressed by aspartate. The negative regulatory effect on both the Aap and glnII was prevented by mutation of any component of the dicarboxylate transport system or by the inclusion of a C4-dicarboxylate in the growth medium, including the non-metabolizable analogue 2-methylsuccinate.

Enzyme assays suggest the involvement of 2-keto-myo-inositol and D-2,3-diketo-4-deoxy epi-inositol in the myo-inositol catabolism of Rhizobium leguminosarum bv. viciae. Growth of R. leguminosarum on myo-inositol increases the enzyme's myo-inositol dehydrogenase and 2-keto-myo-inositol dehydratase. Examination of the growth in glucose as carbon source and resuspension studies in myo-inositol show that O2 consumption is an inducible route. Mutation studies reveal the necessity of intra-cellular dicarboxylation for the repression of these enzymes. Isolation and studies of three transposon mutants that do not induce both enzymes suggest that myo-inositol is not vital as an energy source in bacteriods.

A Tn5 mutant (RU301) of R. leguminosarum that lacks the O-antigen, and which has rough colony morphology and is unable to swarm in TY agar, produces only lipopolysaccharide layer (LPS II). RU301 fails to grow on organic acids but high concentration of calcium or magnesium restores the growth. Southern blotting shows the location at transposon insertion in RU301 in a 3 kb EcoRI fragment. There is a separation of 10 kb between dctD and the region mutated in strain RU301. Additional calcium is not necessary for all LPS rough mutants, and gene mutation in RU301 is essential for nitrogen fixation.

Mucus within the cervical canal represents a hormonally regulated barrier that reconciles the need to exclude the vaginal microflora from the uterus during progesterone dominance, while permitting sperm transport at estrus. Its characteristics change during the estrous cycle to facilitate these competing functional requirements. Hydrated mucin glycoproteins synthesized by the endocervical epithelium form the molecular scaffold of this mucus. This study uses the bovine cervix as a model to examine functional groups of genes related to mucin biosynthesis and mucus production over the periestrous period when functional changes in cervical barrier function are most prominent. Cervical tissue samples were collected from 30 estrus synchronized beef heifers. Animals were slaughtered in groups starting 12 h after the withdrawal of intravaginal progesterone releasing devices (controlled internal drug releases) until 7 days postonset of estrus (luteal phase). Subsequent groupings represented proestrus, early estrus, late estrus, metestrus, and finally the early luteal phase. Tissues were submitted to next generation RNA-seq transcriptome analysis. We identified 114 genes associated with biosynthesis and intracellular transport of mucins, and postsecretory modifications of cervical; 53 of these genes showed at least a twofold change in one or more experimental group in relation to onset of estrus, and the differences between groups were significant (P < 0.05). The majority of these genes showed the greatest alteration in their expression in the 48 h postestrus and luteal phase groups. [ABSTRACT FROM AUTHOR]

The biochemical and biophysical properties of mucinsare largely determined by extensive O-glycosylation of serine- andthreonine-rich tandem repeat (TR) domains. In a number of humandiseases aberrant O-glycosylation is associated with variationsin the properties of the cell surface–associated and secretedmucins. To evaluate in vivo the O-glycosylationof mucin TR domains, we generated recombinant chimeric mucins withTR sequences from MUC2, MUC4, MUC5AC, or MUC5B, which were substituted forthe native TRs of epitope-tagged MUC1 protein (MUC1F). These hybridmucins were extensively O-glycosylated and showed the expected associationwith the cell surface and release into culture media. The presenceof different TR domains within the chimeric mucins appears to have limitedinfluence on their posttranslational processing. Alterations inglycosylation were detailed by fast atom bombardment mass spectrometryand reactivity with antibodies against particular blood-group andtumor-associated carbohydrate antigens. Future applications of thesechimeras will include investigations of mucin posttranslationalmodification in the context of disease. [ABSTRACT FROM PUBLISHER]

The cause of the mucus clearance problems associated with cystic fibrosis remains poorly understood though it has been suggested that mucin hypersecretion, dehydration of mucins, and biochemical abnormalities in the glycosylation of mucins may be responsible. Since the biochemical and biophysical properties of a mucin are dependent on O‐glycosylation, our aim was to evaluate the O‐glycosylation of a single mucin gene product in matched pairs of cells that differed with respect to CFTR expression. An epitope‐tagged MUC1 mucin cDNA (MUC1F) was used to detect variation in mucin glycosylation in stably transfected colon carcinoma cell lines HT29 and Caco2. The glycosylation of MUC1F mucin was evaluated in matched pairs of Caco2 cell lines that either express wild‐type CFTR or have spontaneously lost CFTR expression. The general glycosylation pattern of MUC1F was evaluated by determining its reactivity with a series of monoclonal antibodies against known blood group and tumor‐associated carbohydrate antigens. Metabolic labeling experiments were used to estimate the gross levels of glycosylation and sulfation of MUC1F mucin in these matched pairs of cell lines. Expression of CFTR in this experimental system did not affect the gross levels of glycosylation or sulfation of the MUC1F mucin nor the types of carbohydrates structures attached to the MUC1F protein. [ABSTRACT FROM PUBLISHER]