Increased dopaminergic neurotransmission in therapy-naïve asymptomatic HIV patients is not associated with adaptive changes at the dopaminergic synapses.

Central dopaminergic (DA) systems are affected during human immunodeficiency virus (HIV) infection. So far, it is believed that they degenerate with progression of HIV disease because deterioration of DA systems is evident in advanced stages of infection. In this manuscript we found that (a) DA levels are increased and DA turnover is decreased in CSF of therapy-naïve HIV patients in asymptomatic infection, (b) DA increase does not modulate the availability of DA transporters and D2-receptors, (c) DA correlates inversely with CD4+ numbers in blood. These findings show activation of central DA systems without development of adaptive responses at DA synapses in asymptomatic HIV infection. It is probable that DA deterioration in advanced stages of HIV infection may derive from increased DA availability in early infection, resulting in DA neurotoxicity. Our findings provide a clue to the synergism between DA medication or drugs of abuse and HIV infection to exacerbate and accelerate HIV neuropsychiatric disease, a central issue in the neurobiology of HIV.

Iron regulates transcription of the Escherichia coli ferric citrate transport genes directly and through the transcription initiation proteins.

Ferric citrate induces transcription of the ferric citrate transport genes fecABCDE in Escherichia coli by binding to the outer-membrane receptor protein FecA without entering the cell. Replete iron concentrations inhibit transcription of the fec transport system via the iron-loaded Fur repressor. Here we show that the Fur repressor activated by Mn2+ (used instead of Fe2+) binds to the promoter of the regulatory genes fecIR and to the promoter of fecABCDE. DNase I footprint analysis revealed that Mn2+-Fur (50 nM) protected 30 nucleotides of the coding strand and 24 nucleotides of the noncoding strand of the fecIR promoter. Higher amounts of Mn2+-Fur (100 nM) covered 41 nucleotides of the coding strand of the fecIR promoter and 38 nucleotides of the coding strand of the fecA promoter. The corresponding region of the noncoding strand of the fecA promoter was hypersensitive to DNase I. The results of a deletion analysis of the fecA promoter supported the previously assigned -35 and -10 regions and nucleotide position +11 for FecI-RNA polymerase interaction. Induction of fecIR transcription by iron limitation increased fecB-lacZ transcription 3.5-fold, whereas under constitutive fecIR transcription, iron limitation increased fecB-lacZ transcription twofold. The two iron-regulated sites of fec transport gene transcription suggest a fast response to sufficient intracellular iron concentrations by repression of fecABCDE transcription and a slower adaptation as the result of fecIR transcription inhibition.

Regulation of citrate-dependent iron transport of Escherichia coli: fecR is required for transcription activation by FecI.

Citrate-dependent Fe3+ transport into Escherichia coli K-12 is induced by iron and citrate. The inducer is probably ferric dicitrate which does not have to be taken up into the cytoplasm to induce transcription of the fec transport genes. Two regulatory genes, fecI and fecR, located upstream of the fecABCDE transport genes, are required for induction. We report that in vivo the chromosomally encoded FecI protein activates transcription of the fecA and fecB transport genes in response to ferric citrate and the FecR protein. Cells expressing chromosomally and plasmid-encoded truncated FecR derivatives no longer responded to ferric citrate and expressed the fec transport genes constitutively. The smallest active FecR derivative contained 59 amino acid residues as compared to the 317 residues of wild-type FecR. Constitutive induction was lower than induction of the FecR wild-type strain by ferric citrate. It is concluded that the N-terminal portion of FecR activates FecI and that the C-terminal portion of FecR responds to ferric citrate. Transcription of the fec transport genes is positively regulated by FecI and FecR and negatively regulated by the Fe2(+)-Fur repressor. Transcription activation and repression may occur independently of each other.

Iron transport systems of Serratia marcescens.

Serratia marcescens W225 expresses an unconventional iron(III) transport system. Uptake of Fe3+ occurs in the absence of an iron(III)-solubilizing siderophore, of an outer membrane receptor protein, and of the TonB and ExbBD proteins involved in outer membrane transport. The three SfuABC proteins found to catalyze iron(III) transport exhibit the typical features of periplasmic binding-protein-dependent systems for transport across the cytoplasmic membrane. In support of these conclusions, the periplasmic SfuA protein bound iron chloride and iron citrate but not ferrichrome, as shown by protection experiments against degradation by added V8 protease. The cloned sfuABC genes conferred upon an Escherichia coli aroB mutant unable to synthesize its own enterochelin siderophore the ability to grow under iron-limiting conditions (in the presence of 0.2 mM 2.2'-dipyridyl). Under extreme iron deficiency (0.4 mM 2.2'-dipyridyl), however, the entry rate of iron across the outer membrane was no longer sufficient for growth. Citrate had to be added in order for iron(III) to be translocated as an iron citrate complex in a FecA- and TonB-dependent manner through the outer membrane and via SfuABC across the cytoplasmic membrane. FecA- and TonB-dependent iron transport across the outer membrane could be clearly correlated with a very low concentration of iron in the medium. Expression of the sfuABC genes in E. coli was controlled by the Fur iron repressor gene. S. marcescens W225 was able to synthesize enterochelin and take up iron(III) enterochelin. It contained an iron(III) aerobactin transport system but lacked aerobactin synthesis. This strain was able to utilize the hydroxamate siderophores ferrichrome, coprogen, ferrioxamine B, rhodotorulic acid, and schizokinen as sole iron sources and grew on iron citrate as well. In contrast to E. coli K-12, S. marcescens could utilize heme. DNA fragments of the E. coli fhuA, iut, exbB, and fur genes hybridized with chromosomal S. marcescens DNA fragments, whereas no hybridization was obtained between S. marcescens chromosomal DNA and E. coli fecA, fhuE, and tonB gene fragments. The presence of multiple iron transport systems was also indicated by the increased synthesis of at least five outer membrane proteins (in the molecular weight range of 72,000 to 87,000) after growth in low-iron media. Serratia liquefaciens and Serratia ficaria produced aerobactin, showing that this siderophore also occurs in the genus Serratia.

Nucleotide sequences of the sfuA, sfuB, and sfuC genes of Serratia marcescens suggest a periplasmic-binding-protein-dependent iron transport mechanism.

The cloned sfu region of the Serratia marcescens chromosome confers the ability to grow on iron-limited media to an Escherichia coli K-12 strain that is unable to synthesize a siderophore. This DNA fragment was sequenced and found to contain three genes termed sfuA, sfuB, and sfuC, arranged and transcribed in that order. The sfuA gene encoded a periplasmic polypeptide with calculated molecular weights of 36,154 for the precursor and 33,490 for the mature protein. The sfuB gene product was a very hydrophobic protein with a molecular weight of 56,589. The sfuC gene was found to encode a rather polar but membrane-bound protein with a molecular weight of 36,671 which exhibited strong homology to consensus sequences of nucleotide-binding proteins. The number, structural characteristics, and locations of the SfuABC proteins were typical of a periplasmic-binding-protein-dependent transport mechanism. How Fe3+ is solubilized and taken up across the outer membrane remains an enigma.

Mechanistically novel iron(III) transport system in Serratia marcescens.

A novel iron(III) transport system of Serratia marcescens, named SFU, was cloned and characterized in Escherichia coli. Iron acquisition by this system differed from that by E. coli and related organisms. No siderophore production and no receptor protein related to the SFU system could be detected. In addition, iron uptake was independent of the TonB and ExbB functions. On the cloned 4.8-kilobase sfu fragment, two loci encoding a 36-kilodalton (kDa) protein and three proteins with molecular masses of 40, 38, and 34 kDa were identified; the 40-kDa protein represents a precursor form. Furthermore, chromosomally encoded functions of E. coli were required for the uptake of iron by this system.