UVSSA and USP7, a new couple in transcription-coupled DNA repair

Transcription-coupled nucleotide excision repair (TC-NER) specifically removes transcription-blocking lesions from our genome. Defects in this pathway are associated with two human disorders: Cockayne syndrome (CS) and UV-sensitive syndrome (UVSS). Despite a similar cellular defect in the UV DNA damage response, patients with these syndromes exhibit strikingly distinct symptoms; CS patients display severe developmental, neurological, and premature aging features, whereas the phenotype of UVSS patients is mostly restricted to UV hypersensitivity. The exact molecular mechanism behind these clinical differences is still unknown; however, they might be explained by additional functions of CS proteins beyond TC-NER. A short overview of the current hypotheses addressing possible molecular mechanisms and the proteins involved are presented in this review. In addition, we will focus on two new players involved in TC-NER which were recently identified: UV-stimulated scaffold protein A (UVSSA) and ubiquitin-specific protease 7 (USP7). UVSSA has been found to be the causative gene for UVSS and, together with USP7, is implicated in regulating TC-NER activity. We will discuss the function of UVSSA and USP7 and how the discovery of these proteins contributes to a better understanding of the molecular mechanisms underlying the clinical differences between UVSS and the more severe CS

The use of in vivo brain dialysis of dopamine, acetylcholine, aminoacids and lactic acid in studies on the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)

The use of intracerebral brain dialysis in freely moving rats in neurochemical and neurotoxicological research is discussed and exemplified by studies on the neurotoxin MPTP. Intrastriatal administration of its toxic metabolite MPP+, via the dialysis tube, induced massive changes in the release of neurotransmitters and metabolites. Release enhancing effects could not be repeated by a second MPP+ perfusion and decreases in neurotransmitter or metabolite output were persistent. This indicates that MPP+ has irreversible, toxic effects on various neuortransmitter systems. The MPP+-induced release of DA has been characterized by studying the effect of pretreatment with various drugs, as well by comparison of the time courses of MPP+-induced DA release with those of amphetamine-induced DA release and of MPP+-induced lactate overflow

Affinity for dopamine D-2, D-3, and D-4 receptors of 2-aminotetralins. Relevance of D-2 agonist binding for determination of receptor subtype selectivity

A series of 2-aminotetralins, substituted with a methoxy or a hydroxy group on the 5- or 7-position, and with varying N-alkyl or N-arylalkyl substituents, were prepared and evaluated in binding assays for human dopamine (DA) D-2, D-3, and D-4 receptors. Some members of this series were prepared in former studies, but were never tested in vitro with single receptor subtypes, and these were examined again. None of the tested 2-aminotetralins showed high affinity for the dopamine D-4 receptor. However, a number of the 2-aminotetralins showed high affinity for both the D-2 and the D-3 DA receptors, as exemplified by compounds 11-15 and 21-26, while some had a reasonable selectivity for the DA D-3 receptors. The affinities of the 2-aminotetralins for the D-2L receptor depended on the type of radioligand (agonist or antagonist) used. The agonist affinity data, obtained by using the agonist ligand [H-3]N-0437, are thought to be more relevant for calculating DA receptor subtype selectivity

Mechanism of Translesion Transcription by RNA Polymerase II and Its Role in Cellular Resistance to DNA Damage

UV-induced cyclobutane pyrimidine dimers (CPDs) in the template DNA strand stall transcription elongation by RNA polymerase II (Pol II). If the nucleotide excision repair machinery does not promptly remove the CPDs, stalled Pol II creates a roadblock for DNA replication and subsequent rounds of transcription. Here we present evidence that Pol II has an intrinsic capacity for translesion synthesis (TLS) that enables bypass of the CPD with or without repair. Translesion synthesis depends on the trigger loop and bridge helix, the two flexible regions of the Pol II subunit Rpb1 that participate in substrate binding, catalysis, and translocation. Substitutions in Rpb1 that promote lesion bypass in vitro increase UV resistance in vivo and substitutions that inhibit lesion bypass decrease cell survival after UV irradiation. Thus, translesion transcription becomes essential for cell survival upon accumulation of the unrepaired CPD lesions in genomic DNA