ProSAAS and prohormone convertase 1 are broadly expressed during mouse development

ProSAAS (encoded by mouse gene Pcsk1n) is a recently described neuroendocrine secretory pathway protein that is cleaved into smaller peptides that may function in cell-cell signalling. ProSAAS and its processing intermediates are also potent inhibitors of prohormone convertase 1 (PC1), which is encoded by mouse gene Pcsk1. In order to gain insight into the function of proSAAS, we have examined the distribution of several proSAAS-derived peptides and PC1 by immunohistochemistry throughout mouse development. The distribution patterns of both SAAS and PC1 are broad from E9 to E11, with some enrichment in neural tube-derived tissues. By E15, the expression of SAAS is largely restricted to neuroendocrine tissues known to produce bioactive peptides. In general, the expression pattern of PC1 overlaps with that of SAAS and other proSAAS-derived peptides, consistent with the hypothesis that proSAAS functions as an endogenous PC1 inhibitor.     

Fluorescein-methotrexate transport in dogfish shark (Squalus acanthias) choroid plexus

The vertebrate choroid plexus removes potentially toxic metabolites and xenobiotics from cerebrospinal fluid (CSF) to blood for subsequent excretion in urine and bile. We used confocal microscopy and quantitative image analysis to characterize the mechanisms driving transport of the large organic anion, fluorescein-methotrexate (FL-MTX), from bath (CSF-side) to blood vessels in intact lateral choroid plexus from dogfish shark, Squalus acanthias, an evolutionarily ancient vertebrate. With 2 microM FL-MTX in the bath, steady-state fluorescence in the subepithelium/vascular space exceeded bath levels by 5- to 10-fold, and fluorescence in the epithelial cells was slightly below bath levels. FL-MTX accumulation in both tissue compartments was reduced by NaCN, Na removal, and ouabain, but not by a 10-fold increase in medium K. Certain organic anions, e.g., probenecid, MTX, and taurocholate, reduced FL-MTX accumulation in both tissue compartments; p-aminohippurate and estrone sulfate reduced subepithelial/vascular accumulation, but not cellular accumulation. At low concentrations, digoxin, leukotriene C4, and MK-571 reduced fluorescence in the subepithelium/vascular space while increasing cellular fluorescence, indicating preferential inhibition of efflux over uptake. In the presence of 10 microM digoxin (reduced efflux, enhanced cellular accumulation), cellular FL-MTX accumulation was specific, concentrative, and Na dependent. Thus transepithelial FL-MTX transport involved the following two carrier-mediated steps: electroneutral, Na-dependent uptake at the apical membrane and electroneutral efflux at the basolateral membrane. Finally, FL-MTX accumulation in both tissue compartments was reduced by phorbol ester and increased by forskolin, indicating antagonistic modulation by protein kinase C and protein kinase A.     

Cereulide synthetase gene cluster from emetic Bacillus cereus: Structure and location on a mega virulence plasmid related to Bacillus anthracis toxin plasmid pXO1

Background  

Cereulide, a depsipeptide structurally related to valinomycin, is responsible for the emetic type of gastrointestinal disease caused by Bacillus cereus. Recently, it has been shown that this toxin is produced by a nonribosomal peptide synthetase (NRPS), but its exact genetic organization and biochemical synthesis is unknown.     

Carboxypeptidase A6 in zebrafish development and implications for VIth cranial nerve pathfinding

Carboxypeptidase A6 (CPA6) is an extracellular protease that cleaves carboxy-terminal hydrophobic amino acids and has been implicated in the defective innervation of the lateral rectus muscle by the VIth cranial nerve in Duane syndrome. In order to investigate the role of CPA6 in development, in particular its potential role in axon guidance, the zebrafish ortholog was identified and cloned. Zebrafish CPA6 was secreted and interacted with the extracellular matrix where it had a neutral pH optimum and specificity for C-terminal hydrophobic amino acids. Transient mRNA expression was found in newly formed somites, pectoral fin buds, the stomodeum and a conspicuous condensation posterior to the eye. Markers showed this tissue was not myogenic in nature. Rather, the CPA6 localization overlapped with a chondrogenic site which subsequently forms the walls of a myodome surrounding the lateral rectus muscle. No other zebrafish CPA gene exhibited a similar expression profile. Morpholino-mediated knockdown of CPA6 combined with retrograde labeling and horizontal eye movement analyses demonstrated that deficiency of CPA6 alone did not affect either VIth nerve development or function in the zebrafish. We suggest that mutations in other genes and/or enhancer elements, together with defective CPA6 expression, may be required for altered VIth nerve pathfinding. If mutations in CPA6 contribute to Duane syndrome, our results also suggest that Duane syndrome can be a chondrogenic rather than a myogenic or neurogenic developmental disorder.