Distinct ribonucleoprotein reservoirs for microRNA and siRNA populations in C. elegans.

MicroRNAs (miRNAs) are regulatory molecules that share both biosynthetic derivation (cleavage from short hairpin precursor RNAs) and functional roles (downregulation of specific mRNAs through targeted degradation and/or translational inhibition). A distinct family of small RNAs, termed siRNAs, have some common characteristics but exhibit distinct modes of biosynthesis and function. In this study, we report procedures for purification of a predominant species of miRNA-containing ribonucleoprotein complexes from Caenorhabditis elegans and demonstrate that this population is distinct from the predominant pool of siRNA-containing ribonucleoprotein complexes.

Elongation factor G stabilizes the hybrid-state conformation of the 70S ribosome.

Following peptide bond formation, transfer RNAs (tRNAs) and messenger RNA (mRNA) are translocated through the ribosome, a process catalyzed by elongation factor EF-G. Here, we have used a combination of chemical footprinting, peptidyl transferase activity assays, and mRNA toeprinting to monitor the effects of EF-G on the positions of tRNA and mRNA relative to the A, P, and E sites of the ribosome in the presence of GTP, GDP, GDPNP, and fusidic acid. Chemical footprinting experiments show that binding of EF-G in the presence of the non-hydrolyzable GTP analog GDPNP or GDP.

Single-molecule detection sensitivity using planar integrated optics on a chip.

We present a fully planar integrated optical approach to single-molecule detection based on microfabricated planar networks of intersecting solid and liquid-core waveguides. We study fluorescence from dye molecules in liquid-core antiresonant reflecting optical waveguides, and demonstrate subpicoliter excitation volumes, parallel excitation through multiple pump waveguides, and single-molecule detection sensitivity. Integrated silicon photonics combined with single-molecule detection in solution create a compact, robust, and sensitive platform that has applications in numerous fields ranging from atomic physics to the life sciences.

A method for the determination of environmental contaminants in living marine mammals using microscale samples of blubber and blood.

As part of a study examining the possible effects of organochlorine compounds on juvenile northern elephant seals (Mirounga angustirostris), blubber and blood samples were taken from animals present on the Año Nuevo (California) rookery, and from animals admitted for rehabilitation at The Marine Mammal Center (Sausalito CA). Blubber samples were collected from immobilized seals. A pre-cleaned 6 mm K-medic biopsy punch was used to extract the blubber from a 1 cm incision near the hip, near the dorsal mid point.

Dominant lethal mutations in a conserved loop in 16S rRNA.

The 530 stem-loop region in 16S rRNA is among the most phylogenetically conserved structural elements in all rRNAs, yet its role in protein synthesis remains mysterious. G-530 is protected from kethoxal attack when tRNA, or its 15-nucleotide anticodon stem-loop fragment, is bound to the ribosomal A site. Based on presently available evidence, however, this region is believed to be too remote from the decoding site for this protection to be the result of direct contact.

Binding of tRNA to the ribosomal A and P sites protects two distinct sets of nucleotides in 16 S rRNA.

Transfer RNA protects a characteristic set of bases in 16 S rRNA from chemical probes when it binds to ribosomes. We used several criteria, based on construction of well-characterized in vitro ribosome-tRNA complexes, to assign these proteins to A or P-site binding. All of these approaches lead to similar conclusions.

Evolution of the vacuolar H+-ATPase: implications for the origin of eukaryotes.

Active transport across the vacuolar components of the eukaryotic endomembrane system is energized by a specific vacuolar H+-ATPase. The amino acid sequences of the 70- and 60-kDa subunits of the vacuolar H+-ATPase are approximately equal to 25% identical to the beta and alpha subunits, respectively, of the eubacterial-type F0F1-ATPases. We now report that the same vacuolar H+-ATPase subunits are approximately equal to 50% identical to the alpha and beta subunits, respectively, of the sulfur-metabolizing Sulfolobus acidocaldarius, an archaebacterium (Archaeobacterium).

The H+ ATPase regulatory subunit of Methanococcus thermolithotrophicus: amplification of an 800 bp fragment by polymerase chain reaction.

An 800 bp fragment of Methanococcus thermolithotrophicus genomic DNA was amplified by the polymerase chain reaction method using primers designed from conserved regions of the V-type H+ ATPase regulatory subunits from the archaebacterium Sulfolobus, and several eukaryotes. Although more than one product was obtained, only one of them had the expected size and was exclusively amplified in the presence of the left and right primers. The DNA and the deduced protein sequences of the putative Methanococcus H+ ATPase subunit revealed homology to the corresponding sequences in Sulfolobus and eukaryotes (about 60% identical residues) and a less evident homology to the eubacterial F1-ATPase alpha-subunit (22% identical residues with E.