RNA helicase SACY-1 is required for longevity caused by various genetic perturbations in <i>Caenorhabditis elegans</i>

<p>RNA helicases, which unwind RNAs, are essential for RNA metabolism and homeostasis. However, the roles of RNA helicases in specific physiological processes remain poorly understood. We recently reported that an RNA helicase, HEL-1, promotes long lifespan conferred by reduced insulin/insulin-like growth factor-1 (IGF-1) signaling (IIS) in <i>Caenorhabditis elegans</i>. We also showed that HEL-1 induces the expression of longevity genes by physically interacting with Forkhead box O (FOXO) transcription factor. Thus, the HEL-1 RNA helicase appears to regulate lifespan by specifically activating FOXO in IIS. In the current study, we report another longevity-promoting RNA helicase, Suppressor of ACY-4 sterility 1 (SACY-1). SACY-1 contributed to the longevity of <i>daf-2</i>/insulin/IGF-1 receptor mutants. Unlike HEL-1, SACY-1 was also required for the longevity due to mutations in genes involved in non-IIS pathways. Thus, SACY-1 appears to function as a general longevity factor for various signaling pathways, which is different from the specific function of HEL-1.</p

High-Resolution Live-Cell Imaging and Analysis by Laser Desorption/Ionization Droplet Delivery Mass Spectrometry

We have developed a new ambient-ionization mass spectrometric technique named laser desorption/ionization droplet delivery mass spectrometry (LDIDD-MS). LDIDD-MS permits high-resolution, high-sensitivity imaging of tissue samples as well as measurements of both single-cell apoptosis and live-cell exocytosis. A pulsed (15 Hz) UV laser beam (266 nm) is focused on a surface covered with target analytes to trigger their desorption and ionization. A spray of liquid droplets is simultaneously directed onto the laser-focused surface region to capture the ionized analytes and deliver them to a mass spectrometer. The approach of rapid and effective capturing of molecules after laser desorption/ionization allows the limit of detection for the amino acid lysine to be as low as 2 amol under ambient ionization conditions. Two-dimensional maps of the desorbed/ionized species are recorded by moving the sample on an XY translational stage. The spatial resolution for imaging with LDIDD-MS was determined to be 2.4 μm for an ink-printed pattern and 3 μm for mouse brain tissue. We applied LDIDD-MS to single-cell analysis of apoptotic HEK cells. Differences were observed in the profiles of fatty acids and lipids between healthy HEK cells and those undergoing apoptosis. We observed upregulation of phosphatidylcholine (PC) with a relatively shorter carbon chain length and downregulation of PC with a relatively longer carbon chain length. We also applied LDIDD-MS for a real-time direct measurements of live-cell exocytosis. The catecholamine dopamine and trace amines (phenethylamine and tyramine) were detected from live PC12 cells without damaging them

Precise Expression Profiling by Stuffer-Free Multiplex Ligation-Dependent Probe Amplification

In systems biological studies, precise expression profiling of functionally important gene sets is crucial. Real-time polymerase chain reaction is generally used for this purpose. Despite its widespread acceptance, however, this method is not suitable for multiplex analysis, resulting in an inefficient assay process. One alternative technology in the spotlight is multiplex ligation-dependent probe amplification (MLPA). But MLPA depends on length-based discrimination of amplified products, which complicates probe design and compromises analysis results. Here, we devised a variation of MLPA that utilizes conformation-sensitive capillary electrophoresis, and demonstrated the simplicity of the probe-design process and improved precision of the assay in analyses of 33 <i>Escherichia coli</i> metabolic genes and 16 <i>Caenorhabditis elegans</i> longevity-related genes. The results showed that relative expression could be quantitatively measured over a relevant dynamic range by using similar-sized probes. Importantly, the improved precision compared to conventional MLPA promises a wider application of this method for various biological systems

Selective Fluorescent Detection of RNA in Living Cells by Using Imidazolium-Based Cyclophane

A water-soluble imidazolium-based fluorescent chemosensor senses RNA selectively through fluorescence enhancement over other biologically relevant biomolecules in aqueous solution at physiological pH 7.4. Fluorescence image detection of RNA in living cells such as onion cells, HeLa cells, and animal model cells was successfully demonstrated which displays a chelation-enhanced fluorescence effect. These affinities can be attributed to the strong electrostatic (C–H)⁺···A^– ionic H-bonding and the aromatic moiety driven π-stacking of imidazolium-based cyclophane with the size-complementary major groove of RNA