Hidden ribozymes in eukaryotic genome sequence

The small self-cleaving ribozymes fold into complex tertiary structures to promote autocatalytic cleavage or ligation at a precise position within their sequence. Until recently, relatively few examples had been identified. Two papers now reveal that self-cleaving ribozymes are prevalent in eukaryotic genomes and, in some cases, might play a role in regulating gene expression

The endogenous mex-3 3 UTR is required for germline repression and contributes to optimal fecundity in C. elegans

RNA regulation is essential to successful reproduction. Messenger RNAs delivered from parent to progeny govern early embryonic development. RNA-binding proteins (RBPs) are the key effectors of this process, regulating the translation and stability of parental transcripts to control cell fate specification events prior to zygotic gene activation. The KH-domain RBP MEX-3 is conserved from nematode to human. It was first discovered in Caenorhabditis elegans, where it is essential for anterior cell fate and embryo viability. Here, we show that loss of the endogenous mex-3 3 UTR disrupts its germline expression pattern. An allelic series of 3 UTR deletion variants identify repressing regions of the UTR and demonstrate that repression is not precisely coupled to reproductive success. We also show that several RBPs regulate mex-3 mRNA through its 3 UTR to define its unique germline spatiotemporal expression pattern. Additionally, we find that both poly(A) tail length control and the translation initiation factor IFE-3 contribute to its expression pattern. Together, our results establish the importance of the mex-3 3 UTR to reproductive health and its expression in the germline. Our results suggest that additional mechanisms control MEX-3 function when 3 UTR regulation is compromised

POS-1 and GLD-1 repress glp-1 translation through a conserved binding-site cluster

RNA-binding proteins (RBPs) coordinate cell fate specification and differentiation in a variety of systems. RNA regulation is critical during oocyte development and early embryogenesis, in which RBPs control expression from maternal mRNAs encoding key cell fate determinants. The Caenorhabditis elegans Notch homologue glp-1 coordinates germline progenitor cell proliferation and anterior fate specification in embryos. A network of sequence-specific RBPs is required to pattern GLP-1 translation. Here, we map the cis-regulatory elements that guide glp-1 regulation by the CCCH-type tandem zinc finger protein POS-1 and the STAR-domain protein GLD-1. Our results demonstrate that both proteins recognize the glp-1 3\u27 untranslated region (UTR) through adjacent, overlapping binding sites and that POS-1 binding excludes GLD-1 binding. Both factors are required to repress glp-1 translation in the embryo, suggesting that they function in parallel regulatory pathways. It is intriguing that two equivalent POS-1-binding sites are present in the glp-1 3\u27 UTR, but only one, which overlaps with a translational derepression element, is functional in vivo. We propose that POS-1 regulates glp-1 mRNA translation by blocking access of other RBPs to a key regulatory sequence

hnRNP A1 and secondary structure coordinate alternative splicing of Mag

Myelin-associated glycoprotein (MAG) is a major component of myelin in the vertebrate central nervous system. MAG is present in the periaxonal region of the myelin structure, where it interacts with neuronal proteins to inhibit axon outgrowth and protect neurons from degeneration. Two alternatively spliced isoforms of Mag mRNA have been identified. The mRNA encoding the shorter isoform, known as S-MAG, contains a termination codon in exon 12, while the mRNA encoding the longer isoform, known as L-MAG, skips exon 12 and produces a protein with a longer C-terminal region. L-MAG is required in the central nervous system. How inclusion of Mag exon 12 is regulated is not clear. In a previous study, we showed that heteronuclear ribonucleoprotein A1 (hnRNP A1) contributes to Mag exon 12 skipping. Here, we show that hnRNP A1 interacts with an element that overlaps the 5\u27 splice site of Mag exon 12. The element has a reduced ability to interact with the U1 snRNP compared with a mutant that improves the splice site consensus. An evolutionarily conserved secondary structure is present surrounding the element. The structure modulates interaction with both hnRNP A1 and U1. Analysis of splice isoforms produced from a series of reporter constructs demonstrates that the hnRNP A1-binding site and the secondary structure both contribute to exclusion of Mag exon 12

A compendium of Caenorhabditis elegans RNA binding proteins predicts extensive regulation at multiple levels

Gene expression is regulated at multiple levels, including transcription and translation, as well as mRNA and protein stability. Although systems-level functions of transcription factors and microRNAs are rapidly being characterized, few studies have focused on the posttranscriptional gene regulation by RNA binding proteins (RBPs). RBPs are important to many aspects of gene regulation. Thus, it is essential to know which genes encode RBPs, which RBPs regulate which gene(s), and how RBP genes are themselves regulated. Here we provide a comprehensive compendium of RBPs from the nematode Caenorhabditis elegans (wRBP1.0). We predict that as many as 887 (4.4%) of C. elegans genes may encode RBPs ~250 of which likely function in a gene-specific manner. In addition, we find that RBPs, and most notably gene-specific RBPs, are themselves enriched for binding and modification by regulatory proteins, indicating the potential for extensive regulation of RBPs at many different levels. wRBP1.0 will provide a significant contribution toward the comprehensive delineation of posttranscriptional regulatory networks and will provide a resource for further studies regulation by RBPs

Allosteric inhibition of a stem cell RNA-binding protein by an intermediary metabolite

Gene expression and metabolism are coupled at numerous levels. Cells must sense and respond to nutrients in their environment, and specialized cells must synthesize metabolic products required for their function. Pluripotent stem cells have the ability to differentiate into a wide variety of specialized cells. How metabolic state contributes to stem cell differentiation is not understood. In this study, we show that RNA-binding by the stem cell translation regulator Musashi-1 (MSI1) is allosterically inhibited by 18-22 carbon omega-9 monounsaturated fatty acids. The fatty acid binds to the N-terminal RNA Recognition Motif (RRM) and induces a conformational change that prevents RNA association. Musashi proteins are critical for development of the brain, blood, and epithelium. We identify stearoyl-CoA desaturase-1 as a MSI1 target, revealing a feedback loop between omega-9 fatty acid biosynthesis and MSI1 activity. We propose that other RRM proteins could act as metabolite sensors to couple gene expression changes to physiological state

WHOI Hawaii Ocean Timeseries Station (WHOTS) : WHOTS-6 2009 mooring turnaround cruise report

The Woods Hole Oceanographic Institution (WHOI) Hawaii Ocean Timeseries Site (WHOTS), 100 km north of Oahu, Hawaii, is intended to provide long-term, high-quality air-sea fluxes as a part of the NOAA Climate Observation Program. The WHOTS mooring also serves as a coordinated part of the Hawaiian Ocean Timeseries (HOT) program, contributing to the goals of observing heat, fresh water and chemical fluxes at a site representative of the oligotrophic North Pacific Ocean. The approach is to maintain a surface mooring outfitted for meteorological and oceanographic measurements at a site near 22.75°N, 158°W by successive mooring turnarounds. These observations will be used to investigate air–sea interaction processes related to climate variability. The first WHOTS mooring (WHOTS-1) was deployed in August 2004. Turnaround cruises for successive moorings (WHOTS-2 through WHOTS-5) have typically been in either June or July. This report documents recovery of the WHOTS-5 mooring and deployment of the sixth mooring (WHOTS-6). The moorings utilize Surlyn foam buoys as the surface element and are outfitted with two Air–Sea Interaction Meteorology (ASIMET) systems. Each ASIMET system measures, records, and transmits via Argos satellite the surface meteorological variables necessary to compute air–sea fluxes of heat, moisture and momentum. The upper 155 m of the mooring is outfitted with oceanographic sensors for the measurement of temperature, conductivity and velocity in a cooperative effort with R. Lukas of the University of Hawaii (UH). A pCO2 system is installed on the buoy in a cooperative effort with Chris Sabine at the Pacific Marine Environmental Laboratory. Dr. Frank Bradley, CSIRO, Australia, assisted with meteorological sensor comparisons. A NOAA “Teacher at Sea” and a NOAA “Teacher in the Lab” participated in the cruise. The WHOTS mooring turnaround was done on the University of Hawaii research vessel Kilo Moana, Cruise KM-09-16, by the Upper Ocean Processes Group of the Woods Hole Oceanographic Institution in cooperation with UH and NOAA’s Earth System Research Laboratory, Physical Sciences Division (ESRL/PSD). The cruise took place between 9 and 17 July 2009. Operations began with deployment of the WHOTS-6 mooring on 10 July at approximately 22°40.0'N, 157°57.0'W in 4758 m of water. This was followed by meteorological intercomparisons and CTDs at the WHOTS-6 and WHOTS-5 sites. The WHOTS-5 mooring was recovered on 15 July 2009. The Kilo Moana then moved to the HOT central site (22°45.0'N, 158°00.0'W) for CTD casts. This report describes the cruise operations in more detail, as well as some of the in-port operations and pre-cruise buoy preparations.Funding was provided by the National Oceanic and Atmospheric Administration under Grant No. NA17RJ1223 for the Cooperative Institute for Climate and Ocean Research (CICOR)

Variability in trough total and unbound teicoplanin concentrations and achievement of therapeutic drug monitoring targets in adult patients with hematological malignancy

The objective of this study was to explore the following aspects of teicoplanin use in patients with hematological malignancy: early attainment of target trough concentrations with current high-dose teicoplanin regimens, variability in unbound teicoplanin fractions, factors associated with observed total and unbound trough concentrations, efficacy and toxicity, and renal function estimation. This was a single-center, prospective study. Samples for determination of trough concentrations were taken on days 3, 4, 7, and 10. Total and unbound teicoplanin concentrations were determined using validated high-performance liquid chromatography methods. Regression analyses were used to identify the factors associated with the trough concentration. Thirty teicoplanin-treated adults with hematological malignancy were recruited. Despite the use of dosages higher than the conventional dosages, the proportions of patients with a trough concentration of >= 20 mg/liter at 48 h and at 72 h were 16.7% and 37.9%, respectively. Renal function was significantly negatively associated with total trough concentrations at 48 h and 72 h (P < 0.05). For an average hematological malignancy patient (creatinine clearance = 70 ml/min), sequential loading doses of at least 12 mg/kg of body weight may be needed to achieve early adequate exposure. In the absence of measured creatinine clearance, estimates obtained using the Cockcroft-Gault (total body weight) equation could prove to be an acceptable surrogate. The unbound fractions of teicoplanin were highly variable (3.4 to 18.8%). Higher unbound fractions were observed in patients with low serum albumin concentrations. Teicoplanin was well tolerated. Teicoplanin loading doses higher than those in current use appear to be necessary. Increased dosing is needed in patients with increased renal function. The high variability in protein binding supports the contention for therapeutic drug monitoring of unbound teicoplanin concentrations. (This study has been registered with EudraCT under registration no. 2013-004535-72.

Quaking Regulates Hnrnpa1 Expression through Its 3′ UTR in Oligodendrocyte Precursor Cells

In mice, Quaking (Qk) is required for myelin formation; in humans, it has been associated with psychiatric disease. QK regulates the stability, subcellular localization, and alternative splicing of several myelin-related transcripts, yet little is known about how QK governs these activities. Here, we show that QK enhances Hnrnpa1 mRNA stability by binding a conserved 3′ UTR sequence with high affinity and specificity. A single nucleotide mutation in the binding site eliminates QK-dependent regulation, as does reduction of QK by RNAi. Analysis of exon expression across the transcriptome reveals that QK and hnRNP A1 regulate an overlapping subset of transcripts. Thus, a simple interpretation is that QK regulates a large set of oligodendrocyte precursor genes indirectly by increasing the intracellular concentration of hnRNP A1. Together, the data show that hnRNP A1 is an important QK target that contributes to its control of myelin gene expression