The main immunogenic region (MIR) of the nicotinic acetylcholine receptor and the anti-MIR antibodies

Myasthenia gravis (MG) is caused by autoantibodies against the nicotinic acetylcholine receptor (AChR) of the neuromuscular junction. The anti-AChR antibodies are heterogeneous. However, a small region on the extracellular part of the AChR alpha subunit, called the main immunogenic region (MIR), seems to be the major target of the anti-AChR antibodies, but not of the specific T-cells, in experimental animals and possibly in MG patients. The major loop of the overlapping epitopes for all testable anti-MIR monoclonal antibodies (MAbs) was localized within residues 67-76 (WNPADYGGIK for Torpedo and WNPDDYGGVK for human AChR) of the alpha subunit. The N-terminal half of alpha 67-76 is the most critical, Asn68 and Asp71 being indispensable for binding. Yet anti-MIR antibodies are functionally and structurally quite heterogeneous. Anti-MIR MAbs do not affect channel gating, but they are very potent in mediating acceleration of AChR degradation (antigenic modulation) in cell cultures and in transferring experimental MG in animals. Fab fragments of anti-MIR MAbs bound to the AChR prevent the majority of the MG patients' antibodies from binding to and causing loss of the AChR. Whether this inhibition means that most MG antibodies bind on the same small region or is a result of broad steric/allosteric effects is under current investigation.     

Aspermatogenic effect of the bull seminal ribonuclease (BS RNase) in the presence of anti-BS RNase antibodies in mice

Injection of mouse scrotum with the bull seminal ribonuclease (BS RNase) isolated from bull seminal vesicle fluid inhibited spermatogenesis and caused a decrease in the weight of the testes. Long-term injection of BS RNase evoked the production of antibodies which reached the titre 524448. These antibodies did not prevent the aspermatogenic action of BS RNase in vivo when a twofold higher amount of this enzyme was injected into mouse scrotum. Aspermatogenesis was reversible in both the first and second part of the experiment. During the period of aspermatogenesis the males were sterile. Increasing the amount of BS RNase injections in the second part of experiments caused aspermatogenesis around 3 months. No malformations were observed among offspring of males recovered from the first stage of aspermatogenesis. The antigen—antibody complex prepared in vitro and injected into testes of mice evoked the same degree of aspermatogenesis as the enzyme itself.     

Evaluation of a crown-of-thorns starfish (Acanthaster planci) control program at Grub Reef (central Great Barrier Reef)

A crown-of-thorns starfish control program was conducted at Grub Reef (central Great Barrier Reef) in an area (0.64 km²) which encompassed 53 individual patch reefs. During a two week period, 15 divers injected 3175 starfish with copper sulphate. The program was considered unsuccessful. Although starfish abundance had declined significantly after the control efforts, biological surveys indicated that a relatively large number of starfish remained. The surveys also indicated a general decline in the number of starfish along the reef perimeter, outside the control area. The total cost of the control program was $35 per starfish. These results have important implications for the implementation of future control programs and highlight the need to undertake before and after biological surveys to assess the effectiveness of the control efforts.     

The effects of Heterosigma akashiwo on juvenile Oncorhynchus tshawytscha and its implications for fish culture

The effects of natural blooms of Heterosigma akashiwo on freshwater-and saltwater-acclimated juvenile chinook salmon were assessed. Rates of fish mortality in the blooms were independent of acclimation of fish to seawater and the ambient oxygen levels, but were dependent on concentration of algae and ambient water temperatures. No pathological abnormality to gills or other internal organs in the fish were evident. Aeration or oxygenation of fish cages did not enhance or inhibit fish survival in a H. akashiwo bloom. Cause of death was considered to be due to a labile ichthyotoxic agent.     

New gene assignments using a complete, characterized sheep-hamster somatic cell hybrid panel

The generation and characterization of new sheep-hamster cell hybrids is reported from the fusion of sheep white blood cells with six different hamster auxotrophs. Selection from these and previously generated cell hybrids has led to the production of a panel of 30 hybrids covering the complete sheep genome of 28 chromosomes. Over half of the cell hybrids in this panel contain single sheep chromosomes. By complementation, the following new assignments have been made using the panel: phosphoribosyl N-formylglycinamide amidotransferase (PRFGA) to sheep chromosome (chr) 11; adenylosuccinate synthetase (ADSS) to sheep chr 12; adenylosuccinate lyase (ADSL) to sheep chr 3q; 3-hydroxy-3-methylglutaryl-coenzyme A synthase (HMGCS) to sheep chr 16; dihydrofolate reductase (DHFR) to sheep chr 5; and adenine phosphoribosyltransferase (APRT) to sheep chr 14. The gene phosphoribosylaminoinidazole-carboxamide formyltransferase/Inosinicase (PRACFT) has now been regionally assigned to chr 2q. By isozyme analysis, phosphogluconate dehydrogenase (PGD) was assigned to sheep chr 12, anchoring the sheep syntenic group U1 to this chromosome, and mannose phosphate isomerase (MPI) was assigned to sheep chr 18. Furthermore, the chromosomal assignment of 110 microsatellites was confirmed using this cell panel.     

Determination of deuterium-labeled tryptophan in proteins by means of high-performance liquid chromatography and thermospray mass spectrometry

In order to develop direct methods for determining the extent of metabolic incorporation of isotopically labeled amino acids into a protein, the determination of deuterated tryptophan in [2H5]tryptophan-bacteriorhodopsin was investigated. The isotopically modified protein was subjected to alkaline hydrolysis. After phenyl isothiocyanate derivatization of the hydrolysate, the mixture was separated by reversed-phase liquid chromatography. Field desorption mass spectrometry and thermospray mass spectrometry were investigated for their ability to determine the ratio between [2H5]tryptophan and total tryptophan in the collected fractions. In order to check the procedure a set of known tryptophan/[2H5]tryptophan mixtures were passed through the same derivatization, HPLC separation, and lyophilization procedure as used for the biological samples.