Membrane targeting of RecA during genetic transformation

Recombination in prokaryotes and eukaryotes is mediated by the RecA family of proteins. Although the interactions between RecA and DNA are well studied, the cellular location of these interactions is not known. Using genetic transformation of Streptococcus pneumoniae as a model system, there was increased expression of a protein, colligrin, and RecA, products of the rec locus during genetic transfer. These proteins formed a complex and were found associated with the membranes of genetically competent cells. With immunoelectron microscopy and subcellular fractionation, we showed that the induction of competence led to the translocation of RecA and colligrin to the membrane and to the formation of clusters of RecA in a colligrin-dependent step. Based on the behaviour of colligrin and RecA during genetic exchange and the numerous proteins in prokaryotes and eukaryotes with domains similar to colligrin, we suggest that there may exist a family of proteins, which gathers macromolecules at specific sites in biological membranes.     

Synthesis of a 3-methyluridine phosphoramidite to investigate the role of methylation in a ribosomal RNA hairpin.

The synthesis of a 5'-O-BzH-2'-O-ACE-protected-3-methyluridine phosphoramidite is reported [BzH, benzhydryloxy-bis(trimethylsilyloxy)silyl; ACE, bis(2-acetoxyethoxy)methyl]. The phosphoramidite was employed in solid-phase RNA synthesis to generate a series of RNA hairpins containing single or multiple modifications, including the common nucleoside pseudouridine. Three 19-nucleotide hairpin RNAs that represent the 1920-loop region (G(1906)-C(1924)) of Escherichia coli 23S ribosomal RNA were generated. Modifications were present at positions 1911, 1915, and 1917. The stabilities and structures of the three RNAs were examined by using thermal melting, circular dichroism, and NMR spectroscopy     

Presence of haemocyte-like cells in coccinellid reflex blood

Abstract.  Contrary to current assumptions, the reflex blood of two-spot ladybirds, Adalia bipunctata , and seven-spot ladybirds, Coccinella septempunctata , contains haemocyte-like cells. Furthermore, DNA can be extracted and amplified from coccinellid reflex blood, confirming the presence of haemocyte-like cells and demonstrating a nondestructive method of DNA extraction.     

In-vivo effects of ivermection onRhipicephalus appendiculatus: the influence of tick feeding patterns and drug pharmacokinetics

Sublethal effects seen amongstRhipicephalus appendiculatus feeding on ivermectin-treated rabbits were diverse and dependent both on drug dose, pharmacokinetics and tick feeding patterns: changes in drug formulation, the time of infestation relative to treatment, and the tick instar used, profoundly influenced acaricidal activity. Death was a sequel to paralysis only if tick feeding was interrupted for sufficient time to produce irreversible dehydration. Concurrent pharmacokinetic investigations revealed that, for the larvae ofR. appendiculatus, the mean critical lethal dose of ivermectin imbibed over a 5-day engorgement period was 3500 g/kg. This quantity of ivermectin was achieved in the blood-meals of larvae feeding on rabbits treated subcutaneously with a single dose of Ivomec injection (MSD)^*800 g/kg, provided infestation took place within 24 h of treatment. At lower drug doses, or if larval infestations were delayed for>24 h post-treatment, the quantity of circulating ivermectin (and thus imbibed by the tick larvae) fell below 3500 g/kg and an increasing percentage of larvae successfully engorged and detached. More than 90% of such larvae moulted to the nymphal stage. Nymphae and larvae exhibited similar susceptibility to ivermectin on treated rabbits which could be explained by similar feeding patterns. However, adult female and male ticks were markedly less susceptible and interpretation of ivermectin-induced effects was more complex.     

Whole-embryo culture of Drosophila : development of embryonic tissues in vitro

Summary We have devised techniques to culture whole, dissected embryos of Drosophila melanogaster. We examine multiple aspects of the morphological and physiological development of the epidermis, musculature, nervous system, and internal organs in this cultured preparation, and show that in vitro development closely parallels normal embryogenesis. These techniques permit a wide range of experimental manipulations during embryogenesis and allow us to extend observations through late embryonic stages, after cuticle deposition. Applications of this technique are presented.     

Survey for mitochondrial-desmosome complexes in differentiating epithelia

Summary Mitochondria are frequently found to be closely associated with the plaques of desmosomes in a variety of columnar or cuboidal epithelia of fetal or early postnatal mammals (mouse, rat, human being). The organs in which mitochondrial-desmosome complexes were found include stomach, small intestine, pancreas, kidney, epididymis, seminal vesicle, coagulating gland, thyroid gland. The association has not been observed in simple squamous epithelium (vascular endothelium). Mitochondria lie quite close to desmosomes in the stratum spinosum of stratified squamous mucous epithelium of fetal animals and also to axo-dendritic synapses in still poorly differentiated central nervous system. Mitochondria have also been detected close to attachment sites in ectoderm of the early frog gastrulae. Here there is as yet no visible plaque material.We suggest that the mitochondria may provide energy or some chemical for the formation of the plaque. This hypothesis does not explain why the complexes are not found in poorly differentiated epithelia from older animals.Dedicated to Professor Berta V. Scharrer on her 60th birthday, with affection and admiration. — This study was supported by U.S.P.H.S. research grants NB-05219 and GM-10757 from the National Institutes of Health.