Activity-dependent neuroprotective protein modulates its own gene expression.

We investigated whether activity-dependent neuroprotective protein (ADNP) could autoregulate its own expression. Both the endogenous ADNP gene and reporter gene constructs were analysed in response to overexpression of ADNP, supplied either as wild-type ADNP or a mutant form lacking the NAP motif, a motif which has neuroprotective properties. Overexpression of these two forms of ADNP resulted in both decreased endogenous ADNP expression and repressed ADNP promoter-directed reporter gene activity.

The SLC6A4 VNTR genotype determines transcription factor binding and epigenetic variation of this gene in response to cocaine in vitro.

We demonstrated that the genotype of the variable number tandem repeats (VNTRs) in the linked polymorphic region (LPR) of the 5' promoter and in the intron 2 (Stin2) transcriptional regulatory domains of the serotonin transporter SLC6A4 gene determined its promoter interactions with transcription factors and co-activators in response to cocaine in the JAr cell line. The LPR variants contain 14 (short, s) or 16 (long, l) copies of a 22-23 bp repeat element, whereas the Stin2 VNTR exists as three variants containing 9, 10 or 12 copies of a 16-17 bp repeat. We observed a differential effect of cocaine on the association of the promoter with the transcription factor CTCF, which bound to both LPR alleles prior to cocaine exposure but only to the l-allele following exposure.

Lithium chloride regulation of the substance P encoding preprotachykinin a, Tac1 gene in rat hippocampal primary cells.

In rat hippocampal cultures, the preprotachykinin A (PPTA/Tac1) gene, which encodes the neuropeptide substance P, is regulated by the action of lithium. We used reporter gene and expression constructs to demonstrate that this mechanism of action of lithium is mediated via a previously characterised cis-regulatory Ebox element in the proximal promoter, which binds members of the basic Helix-Loop-Helix family of transcription factors. Consistent with this, in hippocampal cells, both the expression of the endogenous gene and the function of this promoter element are differentially regulated by the basic Helix-Loop-Helix factors, upstream stimulatory factor 1 and 2 (USF1/2).

Distinct gene expression profiles directed by the isoforms of the transcription factor neuron-restrictive silencer factor in human SK-N-AS neuroblastoma cells.

Neuron-restrictive silencer factor (NRSF) and its isoforms are differentially regulated in rodent models of self-sustaining status epilepticus (SSSE). NRSF isoforms regulate genes associated with SSSE, including the proconvulsant tachykinins, brain-derived neurotrophic factor and multiple ion channels. NRSF isoforms may direct distinct gene expression patterns during SSSE, and the ratio of each isoform may be a causative factor in traumatic damage to the central nervous system.

Involvement of preprotachykinin A gene-encoded peptides and the neurokinin 1 receptor in endotoxin-induced murine airway inflammation.

Tachykinins encoded by the preprotachykinin A (TAC1) gene such as substance P (SP) and neurokinin A (NKA) are involved in neurogenic inflammatory processes via predominantly neurokinins 1 and 2 (NK1 and NK2) receptor activation, respectively. Endokinins and hemokinins encoded by the TAC4 gene also have remarkable selectivity and potency for the NK1 receptors and might participate in inflammatory cell functions. The aim of the present study was to investigate endotoxin-induced airway inflammation and consequent bronchial hyper-reactivity in TAC1(-/-), NK1(-/-) and also in double knockout (TAC1(-/-)/NK1(-/-)) mice.

Combinatorial interaction between two human serotonin transporter gene variable number tandem repeats and their regulation by CTCF.

Two distinct variable number tandem repeats (VNTRs) within the human serotonin transporter gene (SLC6A4) have been implicated as predisposing factors for CNS disorders. The linked polymorphic region in the 5'-promoter exists as short (s) and long (l) alleles of a 22 or 23 bp elements. The second within intron 2 (Stin2) exists as three variants containing 9, 10 or 12 copies of a 16 or 17 bp element.

Induction of tachykinin production in airway epithelia in response to viral infection.

Background: The tachykinins are implicated in neurogenic inflammation and the neuropeptide substance P in particular has been shown to be a proinflammatory mediator. A role for the tachykinins in host response to lung challenge has been previously demonstrated but has been focused predominantly on the release of the tachykinins from nerves innervating the lung. We have previously demonstrated the most dramatic phenotype described for the substance P encoding gene preprotachykinin-A (PPT-A) to date in controlling the host immune response to the murine gammaherpesvirus 68, in the lung.

Analysis of yeast artificial chromosome DNA by restriction digestion, southern blotting nucleic acid hybridization, and polymerase chain reaction.

Restriction enzyme analysis of yeast artificial chromosome (YAC) clones in combination with Southern blotting and polymerase chain reaction (PCR) is necessary to establish the integrity of the DNA contained within the YAC clone, as well as obtain information on the integrity of the cloned DNA. Restriction analysis is also a useful tool that allows a direct comparison between endogenous genomic DNA and the DNA fragment present within a YAC clone. This chapter summarizes the techniques of enzyme digestion of YAC DNA and the separation of the DNA fragments by pulse-field gel electrophoresis.