A highly conserved nuclear gene for low-level phylogenetics: elongation factor-1 alpha recovers morphology-based tree for heliothine moths

Molecular systematists need increased access to nuclear genes. Highly conserved, low copy number protein-encoding nuclear genes have attractive features for phylogenetic inference but have heretofore been applied mostly to very ancient divergences. By virtue of their synonymous substitutions, such genes should contain a wealth of information about lower-level taxonomic relationships as well, with the advantage that amino acid conservatism makes both alignment and primer definition straightforward. We tested this postulate for the elongation factor-1 alpha (EF-1 alpha) gene in the noctuid moth subfamily Heliothinae, which has probably diversified since the middle Tertiary. We sequenced 1,240 bp in 18 taxa representing heliothine groupings strongly supported by previous morphological and allozyme studies. The single most parsimonious gene tree and the neighbor-joining tree for all nucleotides show almost complete concordance with the morphological tree. Homoplasy and pairwise divergence levels are low, transition/transversion ratios are high, and phylogenetic information is spread evenly across gene regions. The EF-1 alpha gene and presumably other highly conserved genes hold much promise for phylogenetics of Tertiary age eukaryote groups.      

Mesodern-inducing factors and mesodermal patterning

Identification of the signalling molecules involved in mesoderm formation in amphibian embryos still presents problems. None of the original candidates, such as activin, have been definitively ruled out, and the new factors, such as the nodal-related genes, have come on to the scene. Of the original candidates, activin has been definitively shown to act as a morphogen, whereas bone morphogenetic protein (BMP)-4 has emerged as a ventral inducer and an inhibitor of neural differentiation. The effects of BMP-4 are antagonized by chordin, a molecule related to the product of the Drosophila gene short gastrulation.     

Synergistic effect of purine derivatives on the toxicity of pyrazofurin and 6-azauridine towards cultured mammalian cells

A novel synergistic effect of several purine derivatives such as adenine, adenosine, hypoxanthine, and guanine on the toxicity of nucleoside analogs pyrazofurin and 6-azauridine towards cultured Chinese hamster ovary (CHO) cells has been observed. The presence of the above purine derivatives enhanced the toxicity of pyrazofurin and 6-azauridine, in a dose dependent manner. The growth inhibitory effects of these nucleoside analogs either alone or in combination with the purine derivatives were reversed by uridine and cytidine, providing evidence that the synergistic effect of the purine derivatives was exerted at the level of pyrimidine nucleotide biosynthesis. Studies with mutant cells lacking various purine phosphorylating enzymes show that phosphorylation of purine derivatives through reactions utilizing phosphoribosylpyrophosate (PRPP) is essential for observing the synergistic response. It is suggested that the above purine derivatives (including adenosine, via conversion to hypoxanthine) exert their synergistic effects by depleting the cellular pool of PRPP by two separate mechanisms (direct utilization and feedback inhibition of its synthesis), which as a result becomes rate limiting in the synthesis of orotidine monophosphate (OMP). The reduced levels of OMP, which is a competing substrate with pyrazofurin- and 6-azauridine-5'-monophosphates for binding to the target enzyme OMP decarboxylase, could then account for the inhibition of the enzyme at lower concentrations of these analogs.     

Characterization of crystals of a cytochrome oxidase (nitrite reductase) from Pseudomonas aeruginosa by x-ray diffraction and electron microscopy

Cytochrome oxidase from Pseudomonas aeruginosa has been crystallized from 2 m-ammonium sulfate. The crystals occur principally as thin diamond-shaped plates of space group P2₁2₁2 with unit cell dimensions of 92 Å × 115 Å × 76 Å. Determination of the density of glutaraldehyde-fixed, water-equilibrated crystals (1.167 g/cm³), coupled with the unit cell volume (804,000 ų), indicates that there is one subunit (~63,000 Mr) per asymmetric unit. X-ray diffraction data which were limited to 12 Å resolution due to small crystal size were obtained for the hk0 and 0kl zones using precession photography. Amplitude and phase data for the hk0, 0kl, and h0l zones were obtained from computer-based Fourier analysis of appropriate micrographs recorded from negatively stained microplates and thin sections of larger crystals using minimal beam electron microscopy. For crystals embedded in the presence of tannic acid it was possible to achieve 20 Å resolution which is comparable to the resolution achieved with negative staining of thin crystalline arrays. In addition, unstained electron diffraction on glutaraldehyde-fixed, glucose-stabilized plates was recorded to a resolution of 9 Å. The three-dimensional packing of the cytochrome oxidase dimer in the unit cell has been deduced from computer reconstructed images of the three principal projections along the crystallographic axes. The cytochrome oxidase dimer is located in the unit cell with the dimer axis coincident with a crystallographic 2-fold axis; thus within the resolution of the present data in projection (9 Å) the two subunits are identical, in agreement with biochemical evidence. The crystals have been prepared with the enzyme in the fully oxidized state and upon reduction a progressive cracking of the crystals is observed, possibly due to a conformational change dependent on the oxidation state of the heme iron.     

Auditory sensitivity of the saccule in the American toad (Bufo americanus)

Summary Single unit recordings in the posterior nerve branchlet from the saccule have shown that, in the American toad (Bufo americanus), approximately 30% of the fibers respond to airborne sounds in a way similar to fibers from the two known auditory organs, the amphibian and basilar papillae. In response to tones, saccule fibers have best excitatory frequencies which fall into two disjoint populations: units in the low-frequency-sensitive group (below 300 Hz) show tone-on-tone suppression while those in the high-frequency-sensitive group (700–1,200 Hz) show no evidence of peripheral inhibition. Saccule units have somewhat higher thresholds than those from the other auditory organs. It is suggested that the high-frequency-sensitive fibers might be useful for discriminating mating calls in an intense chorus while the low-frequency-sensitive units likely respond to other high intensity sounds in the environment.Research supported by the U.S. Public Health Service (NIH grant NS-09244).