rRNA complementarity within mRNAs: A possible basis for mRNA-ribosome interactions and translational control

Our recent demonstration that many eukaryotic mRNAs contain sequences complementary to rRNA led to the hypothesis that these sequences might mediate specific interactions between mRNAs and ribosomes and thereby affect translation. In the present experiments, the ability of complementary sequences to bind to rRNA was investigated by using photochemical cross-linking. RNA probes with perfect complementarity to 18S or 28S rRNA were shown to cross-link specifically to the corresponding rRNA within intact ribosomal subunits. Similar results were obtained by using probes based on natural mRNA sequences with varying degrees of complementarity to the 18S rRNA. RNase H cleavage localized four such probes to complementary regions of the 18S rRNA. The effects of complementarity on translation were assessed by using the mRNA encoding ribosomal protein S15. This mRNA contains a sequence within its coding region that is complementary to the 18S rRNA at 20 of 22 nucleotides. RNA from an S15-luciferase fusion construct was translated in a cell-free lysate and compared with the translation of four related constructs that were mutated to decrease complementarity to the 18S rRNA. These mutations did not alter the amino acid sequence or the codon bias. A correlation between complementarity and translation was observed; constructs with less complementarity increased the amount of translation up to 54%. These findings raised the possibility that direct base-pairing of particular mRNAs to rRNAs within ribosomes may function as a mechanism of translational control.

Parkinson disease: A new link between monoamine oxidase and mitochondrial electron flow

Two factors that contribute to the progression of Parkinson disease are a brain defect in mitochondrial respiration and the generation of hydrogen peroxide (H2O2) by monoamine oxidase (MAO). Here we show that the two are linked. Metabolism of the neurotransmitter dopamine, or other monoamines (benzylamine, tyramine), by intact rat brain mitochondria suppresses pyruvate- and succinate-dependent electron transport. MAO inhibitors prevent this action. Mitochondrial damage is also reversed during electron flow. A probable explanation is that MAO-generated H2O2 oxidizes glutathione to glutathione disulfide (GSSG), which undergoes thiol-disulfide interchange to form protein mixed disulfides, thereby interfering reversibly with thiol-dependent enzymatic function. In agreement with this premise, direct addition of GSSG to mitochondria resulted in similar reversible inhibition of electron transport. In addition, the monoamines induced an elevation in protein mixed disulfides within mitochondria. These observations imply that (i) heightened activity and metabolism of neurotransmitter by monoamine neurons may affect neuronal function, and (ii) apparent defects in mitochondrial respiration associated with Parkinson disease may reflect, in part, an established increase in dopamine turnover. The experimental results also target mitochondrial repair mechanisms for further investigation and may, in time, lead to newer forms of therapy.

Why are patients prescribed proton pump inhibitors? Retrospective analysis of link between morbidity and prescribing in the General Practice Research Database

Objectives: To establish the relation between new prescriptions for proton pump inhibitors and recorded upper gastrointestinal morbidity within a large computerised general practitioner database.

GIPC and GAIP Form a Complex with TrkA: A Putative Link between G Protein and Receptor Tyrosine Kinase Pathways

NGF initiates the majority of its neurotrophic effects by promoting the activation of the tyrosine kinase receptor TrkA. Here we describe a novel interaction between TrkA and GIPC, a PDZ domain protein. GIPC binds to the juxtamembrane region of TrkA through its PDZ domain. The PDZ domain of GIPC also interacts with GAIP, an RGS (regulators of G protein signaling) protein. GIPC and GAIP are components of a G protein-coupled signaling complex thought to be involved in vesicular trafficking. In transfected HEK 293T cells GIPC, GAIP, and TrkA form a coprecipitable protein complex. Both TrkA and GAIP bind to the PDZ domain of GIPC, but their binding sites within the PDZ domain are different. The association of endogenous GIPC with the TrkA receptor was confirmed by coimmunoprecipitation in PC12 (615) cells stably expressing TrkA. By immunofluorescence GIPC colocalizes with phosphorylated TrkA receptors in retrograde transport vesicles located in the neurites and cell bodies of differentiated PC12 (615) cells. These results suggest that GIPC, like other PDZ domain proteins, serves to cluster transmembrane receptors with signaling molecules. When GIPC is overexpressed in PC12 (615) cells, NGF-induced phosphorylation of mitogen-activated protein (MAP) kinase (Erk1/2) decreases; however, there is no effect on phosphorylation of Akt, phospholipase C-γ1, or Shc. The association of TrkA receptors with GIPC and GAIP plus the inhibition of MAP kinase by GIPC suggests that GIPC may provide a link between TrkA and G protein signaling pathways.

Cancer-predisposing mutations within the RING domain of BRCA1: Loss of ubiquitin protein ligase activity and protection from radiation hypersensitivity

BRCA1 is a breast and ovarian cancer-specific tumor suppressor that seems to be involved in transcription and DNA repair. Here we report that BRCA1 exhibits a bona fide ubiquitin (Ub) protein ligase (E3) activity, and that cancer-predisposing mutations within the BRCA1 RING domain abolish its Ub ligase activity. Furthermore, these mutants are unable to reverse γ-radiation hypersensitivity of BRCA1-null human breast cancer cells, HCC1937. Additionally, these mutations within the BRCA1 RING domain are not capable of restoring a G2 + M checkpoint in HCC1937 cells. These results establish a link between Ub protein ligase activity and γ-radiation protection function of BRCA1, and provide an explanation for why mutations within the BRCA1 RING domain predispose to cancer. Furthermore, we propose that the analysis of the Ub ligase activity of RING-domain mutations identified in patients may constitute an assay to predict predisposition to cancer.