Akt induces down regulation of MUC5AC production in NCI-H292 human airway epithelial cells cultured on extracellular matrix.

MUC5AC mucin overproduction is a key feature of asthma as contributes to airway obstruction. The production of MUC5AC is regulated in part by signals from extracellular matrix via integrin pathways, but it remains largely unclear. We investigated the role of Akt, a typical signal transducer in the integrin pathway, in the regulation of MUC5AC production. When NCI-H292 human airway epithelial cells were cultured on laminin or Matrigel, we found that the activity of Akt was suppressed, as compared to control cells with upregulated MUC5AC production. In contrast, Akt was activated in cells cultured on type IV collagen with downregulated MUC5AC production. The Akt inhibitor induced upregulation of MUC5AC. In contrast, overexpression of active Akt induced downregulation of MUC5AC production. These results suggest that a signal from laminin or Matrigel induces upregulation of MUC5AC by suppressing Akt activity, whereas a signal from type IV collagen induces downregulation of MUC5AC, mediated by Akt activation.

Production of bioactive bovine fibroblast growth factor 4 in E. coli based on the common nucleotide sequence of its structural gene in three breeds.

Fibroblast growth factor 4 (FGF4) is considered a crucial gene in the proper development of bovine embryos. We recently determined the FGF4 gene sequence in eight cattle derived from three breeds and revealed a common nucleotide sequence of the structural gene encoding FGF4, which leads to the deletion and mutation of amino acid sequences in the mature FGF4 (Pro(32) -Leu(206) ) compared with the sequence previously reported. In the present study, HisbFGF4, a 6× histidine-tagged bovine FGF4 (Pro(32) -Leu(206) ), was produced in Escherichia coli based on the validated nucleotide sequence and purified by heparin column chromatography. In primary bovine fibroblasts, HisbFGF4 showed significant mitogenic activity, whereas, intriguingly, the activity of a commercially available recombinant human FGF4 (Gly(25) -Leu(206) ) produced in E. coli was weaker than that of HisbFGF4. In conclusion, the present study provides a simple method for the production of a bioactive bovine FGF4 derivative in E. coli utilizing its structural gene elucidated by us.

Common nucleotide sequence of structural gene encoding fibroblast growth factor 4 in eight cattle derived from three breeds.

Fibroblast growth factor 4 (FGF4) is considered as a crucial gene for the proper development of bovine embryos. However, the complete nucleotide sequences of the structural genes encoding FGF4 in identified breeds are still unknown. In the present study, direct sequencing of PCR products derived from genomic DNA samples obtained from three Japanese Black, two Japanese Shorthorn and three Holstein cattle, revealed that the nucleotide sequences of the structural gene encoding FGF4 matched completely among these eight cattle. On the other hand, differences in the nucleotide sequences, leading to substitutions, insertions or deletions of amino acid residues were detected when compared with the already reported sequence from unidentified breeds. We cannot rule out a possibility that the structural gene elucidated in the present study is widely distributed in cattle. To the best of our knowledge, this is the first determination of the complete nucleotide sequence of the structural gene encoding bovine FGF4 in identified breeds.