A DEAD-box RNA helicase promotes thermodynamic equilibration of kinetically trapped RNA structures in vivo.

RNAs must assemble into specific structures in order to carry out their biological functions, but in vitro RNA folding reactions produce multiple misfolded structures that fail to exchange with functional structures on biological time scales. We used carefully designed self-cleaving mRNAs that assemble through well-defined folding pathways to identify factors that differentiate intracellular and in vitro folding reactions. Our previous work showed that simple base-paired RNA helices form and dissociate with the same rate and equilibrium constants in vivo and in vitro.

The ydaO motif is an ATP-sensing riboswitch in Bacillus subtilis.

We report what is to our knowledge the first natural RNA that regulates gene expression in response to intracellular ATP. Using a biochemical screen, we found that several putative riboswitches bind ATP in vitro. The ydaO motif specifically bound ATP and regulated expression of endogenous and reporter genes in response to ATP concentrations in Bacillus subtilis.

The glmS riboswitch integrates signals from activating and inhibitory metabolites in vivo.

The glmS riboswitch belongs to the family of regulatory RNAs that provide feedback regulation of metabolic genes. It is also a ribozyme that self-cleaves upon binding glucosamine-6-phosphate, the product of the enzyme encoded by glmS. The ligand concentration dependence of intracellular self-cleavage kinetics was measured for the first time in a yeast model system and unexpectedly revealed that this riboswitch is subject to inhibition as well as activation by hexose metabolites.

mRNA secondary structures fold sequentially but exchange rapidly in vivo.

RNAs adopt defined structures to perform biological activities, and conformational transitions among alternative structures are critical to virtually all RNA-mediated processes ranging from metabolite-activation of bacterial riboswitches to pre-mRNA splicing and viral replication in eukaryotes. Mechanistic analysis of an RNA folding reaction in a biological context is challenging because many steps usually intervene between assembly of a functional RNA structure and execution of a biological function. We developed a system to probe mechanisms of secondary structure folding and exchange directly in vivo using self-cleavage to monitor competition between mutually exclusive structures that promote or inhibit ribozyme assembly.

Determination of intracellular RNA folding rates using self-cleaving RNAs.

We have developed a system that relies on RNA self-cleavage to report quantitatively on assembly of RNA structures in vivo. Self-cleaving RNA sequences are inserted into mRNAs or snoRNAs and expressed in yeast under the control of a regulated promoter. Chimeric RNAs that contain self-cleaving ribozymes turn over faster than chimeric RNAs that contain a mutationally inactivated ribozyme by an amount that reflects the rate at which the ribozyme folds and self-cleaves.

Resident and faculty perceptions of conflict of interest in medical education.

Objective: To determine resident and faculty perceptions of the pharmaceutical industry's influence on medical education.

Design, Setting, And Participants: Anonymous survey of categorical residents and faculty in the department of medicine at a large, Midwestern, urban, independent academic medical center.

Main Results: Eighty-one residents (69.