ABSTRACT
The synaptic actions of the neurotransmitter serotonin are terminated by a selective re-uptake system located in the axonal membrane. To gain information about the quaternary structure of this membrane protein, we transiently expressed the recombinant rat serotonin transporter, SERT1, in human embryonic kidney 293 cells. Treatment with sulfhydryl oxidizing agents and the homobifunctional cross-linker dimethyl suberimidate (DMS) generated adducts of 130-180 kDa and 220-270 kDa, respectively. These data indicate an oligomeric structure of SERT1.
Subject(s)
Carrier Proteins/chemistry , Membrane Glycoproteins/chemistry , Membrane Transport Proteins , Nerve Tissue Proteins , Protein Conformation , Animals , Carrier Proteins/metabolism , Cell Line , Cell Membrane/chemistry , Cell Membrane/metabolism , Cross-Linking Reagents/pharmacology , Diamide/pharmacology , Dimethyl Suberimidate/pharmacology , Electrophoresis, Polyacrylamide Gel , Humans , Immunoblotting , Membrane Glycoproteins/metabolism , Molecular Weight , Phenanthrolines/pharmacology , Rats , Recombinant Proteins/chemistry , Recombinant Proteins/isolation & purification , Serotonin/metabolism , Serotonin Plasma Membrane Transport Proteins , Sulfhydryl Reagents/pharmacologyABSTRACT
During a systematic search for open reading frames in chromosome band Xp11.2, a novel gene (ZNF157) that encodes a putative 506-amino-acid protein with the sequence characteristics of a zinc-finger-containing transcription factor was isolated. ZNF157 is encoded by four exons distributed over > 20 kb of genomic DNA. The second and third exons contain sequences similar to those of the previously described KRAB-A and KRAB-B domains, motifs that have been shown to mediate transcriptional repression in other members of the protein family. A fourth exon contains 12 zinc finger DNA binding motifs and finger linking regions characteristic of ZNF proteins of the Krüppel family. ZNF157 maps to the telomeric end of a cluster of ZNF genes that includes ZNF21, ZNF41, and ZNF81.
Subject(s)
X Chromosome , Zinc Fingers/genetics , Amino Acid Sequence , Base Sequence , Cloning, Molecular , DNA, Complementary , Exons , Humans , Introns , Molecular Sequence Data , Sequence Homology, Amino AcidABSTRACT
The efflux of reduced glutathione (GSH) into bile appears to be a major contribution to the hepatic glutathione homeostasis. In bile-fistula rats pretreatment with phenobarbital strongly enhanced the biliary export of GSH, whereas hepatic damage due to carbon tetrachloride markedly decreased the transport of GSH into bile. An oral load of ethanol, which was not hepatotoxic in rats, had the same effect in this respect. Depletion of hepatic GSH in response to the treatment with diethylmaleate, vinylidene chloride or paracetamol was followed by a decrease in the efflux of GSH into bile. Phorone which depleted the hepatic GSH nearly totally, in contrast, even enhanced its biliary output indicating a rapid decomposition of the phorone-GSH-adduct. Our experiments support recent findings of others that the canalicular transport of reduced GSH requires an active energy-dependent carrier mechanism.
