Using fluorescence-based human androgen receptor gene assay to analyze the clonality of microdissected dendritic cell tumors

The AGT1 permease is a alpha-glucoside-H+ symporter responsible for the active transport of maltose, trehalose, maltotriose, alpha-methylglucoside, melezitose and sucrose. In wild-type as well as in MAL constitutive strains, alpha-methylglucoside seemed to be the best inducer of transport activity, while trehalose had no inducing effect. Based on the initial rates of transport it seems that the sugar preferentially transported by this permease is trehalose, followed by sucrose.     

cDNA fingerprinting of osteoprogenitor cells to isolate differentiation stage-specific genes

A cDNA fingerprinting strategy was developed to identify genes based on their differential expression pattern during osteoblast development. Preliminary biological and molecular staging of cDNA pools prepared by global amplification PCR allowed discrim-inating choices to be made in selection of expressed sequence tags (ESTs) to be isolated. Sequencing of selected ESTs confirmed that both known and novel genes can be isolated from any developmental stage of interest, e.g. from primitive progenitors, intermediate precursors or mature osteoblasts. EST expression provides insight into possible interrelated physiological functions and putative interacting molecules during differentiation. This method offers a functional genomics approach to isolate differentiation stage-specific genes in samples as small as a single cell.     

Synergism of constitutive activity in alpha 1-adrenergic receptor activation

Recently a number of mutations have been found in vitro which maintain alpha 1-adrenergic receptors (ARs) in a partially activated form. We have previously identified two amino acid residue positions in the alpha 1b-adrenergic receptor (AR), Cys128 and Ala204, one in each of the third and fifth transmembrane segments, that constitutively activate the receptor when substituted for a phenylalanine or valine, respectively [Perez et al. (1996) Mol. Pharmacol. 49, 112-122; Hwa et al. (1996) J. Biol. Chem. 271, 7956-7964]. Another mutation analyzed previously, Ala293Glu, located in the third intracellular loop, also constitutively activates the receptor [Kjelsborg et al. (1992) J. Biol. Chem. 267, 1430-1433]. All three mutations displayed similar manifestations of constitutive activity such as higher binding affinity and potency for agonists as well as higher basal signal transduction as predicted by the revised ternary complex model of receptor activation. We hypothesized that the individual mutations because of their critical location alter the conformation of the transmembrane helices such that mimicry occur that partially conforms to the activated state, R*. To explore whether these potential conformations are independent, we combined these three mutations in all possible permutations. The combined triple mutation displays 700-fold higher binding affinity for (-)-epinephrine and 20-fold higher basal IP3 release than the wild-type receptor. We also observed that each mutation contributed independently and synergistically to both receptor agonist binding and activation with the combined mutations basal activity exceeding that of the fully-stimulated wild-type receptor. There was also a direct correlation between epinephrine's binding affinity and the degree of constitutive activity. Because the mutations affect different transmembrane domains, these results are consistent with a mechanism that helical movement acts in a concerted fashion in agonist-induced activation, a synergism predicted if multiple helix movement is involved in receptor activation.     

Liver cell necrosis during administration of clozapine

A 38-year-old woman was treated for mutism with clozapine. After a week liver function disturbances developed, which disappeared when the treatment was discontinued. Histopathological investigation of a liver biopsy specimen revealed extensive liver cell necrosis. So far two patients have been described with cholestatic jaundice induced by clozapine, and one patient with toxic hepatitis due to clozapine.