Proceedings of the National Academy of Sciences of the United States of America
-
Proc. Natl. Acad. Sci. U.S.A. · Nov 2013
Correction of mutations within the cystic fibrosis transmembrane conductance regulator by site-directed RNA editing.
Adenosine deaminases that act on RNA are a conserved family of enzymes that catalyze a natural process of site-directed mutagenesis. Biochemically, they convert adenosine to inosine, a nucleotide that is read as guanosine during translation; thus when editing occurs in mRNAs, codons can be recoded and the changes can alter protein function. By removing the endogenous targeting domains from human adenosine deaminase that acts on RNA 2 and replacing them with an antisense RNA oligonucleotide, we have engineered a recombinant enzyme that can be directed to edit anywhere along the RNA registry. ⋯ In Xenopus oocytes, we show that a genetically encoded version of our editase can correct cystic fibrosis transmembrane conductance regulator mRNA, restore full-length protein, and reestablish functional chloride currents across the plasma membrane. Finally, in a human cell line, we show that a genetically encoded version of our editase and guide RNA can correct a nonfunctional version of enhanced green fluorescent protein, which contains a premature termination codon. This technology should spearhead powerful approaches to correcting a wide variety of genetic mutations and fine-tuning protein function through targeted nucleotide deamination.
-
Proc. Natl. Acad. Sci. U.S.A. · Nov 2013
Dynamics of dendritic spines in the mouse auditory cortex during memory formation and memory recall.
Long-lasting changes in synaptic connections induced by relevant experiences are believed to represent the physical correlate of memories. Here, we combined chronic in vivo two-photon imaging of dendritic spines with auditory-cued classical conditioning to test if the formation of a fear memory is associated with structural changes of synapses in the mouse auditory cortex. ⋯ Memory recall triggered by the reexposure of mice to the sound cue did not lead to changes in spine dynamics. Our findings provide a synaptic mechanism for plasticity in sound responses of auditory cortex neurons induced by auditory-cued fear conditioning; they also show that retrieval of an auditory fear memory does not lead to a recapitulation of structural plasticity in the auditory cortex as observed during initial memory consolidation.
-
Proc. Natl. Acad. Sci. U.S.A. · Oct 2013
Clinical TrialPlacebo improves pleasure and pain through opposite modulation of sensory processing.
Placebo analgesia is often conceptualized as a reward mechanism. However, by targeting only negative experiences, such as pain, placebo research may tell only half the story. We compared placebo improvement of painful touch (analgesia) with placebo improvement of pleasant touch (hyperhedonia) using functional MRI and a crossover design. ⋯ Both placebo responses were associated with similar patterns of activation increase in circuitry involved in emotion appraisal, including the pregenual anterior cingulate, medial orbitofrontal cortex, amygdala, accumbens, and midbrain structures. Importantly, placebo-induced coupling between the ventromedial prefrontal cortex and periaqueductal gray correlated with somatosensory decreases to painful touch and somatosensory increases to pleasant touch. These findings suggest that placebo analgesia and hyperhedonia are mediated by activation of shared emotion appraisal neurocircuitry, which down- or up-regulates early sensory processing, depending on whether the expectation is reduced pain or increased pleasure.