Rat pineal S-antigen: sequence analysis reveals presence of alpha-transducin homologous sequence

S-antigen (S-Ag) is a soluble, highly antigenic protein, the administration of which induces autoimmune uveitis. This protein is found in the retina and pineal. Retinal S-Ag from three species has been sequenced. In this study rat pineal S-Ag was sequenced. Clones were isolated from a rat pineal lambda gt11 cDNA library by probing with a 300 bp fragment of mouse retinal S-Ag cDNA containing the 5'-coding region. The largest clone isolated (RPS-118; 1364 bp) contained the entire coding sequence. Comparison of the rat pineal and mouse retinal S-Ag nucleotide sequences indicated a high homology (95%). The deduced amino acid sequence was found to contain 403 residues (congruent to 44 992 Da). Comparison of the rat pineal and mouse retinal S-Ag amino acid sequences also revealed high homology (97%). The similarity of both the nucleotide and amino acid sequences of rat pineal and mouse retinal S-Ag indicates that expression of the S-Ag gene in both tissues is similar. Further analysis of the rat pineal S-Ag sequence indicated that it contained essentially the same major uveitopathogenic region of S-Ag present in bovine retina; minor uveitopathogenic sites were somewhat different. As is true of retinal S-Ag, rat pineal S-Ag contains the same consensus phosphoryl-binding site present in many GTP/GDP-binding proteins and a homologous sequence found in the C-terminus of alpha-transducin. These sequences may play a role in the action of pineal S-Ag in transmembrane signal transduction.     

Ovariectomized steroid-treated mares as embryo transfer recipients and as a model to study the role of progestins in pregnancy maintenance

Embryo transfer into ovariectomized steroid-treated mares was used as a model to evaluate various progestin/estradiol treatments and to determine the level of progesterone necessary for the maintenance of pregnancy in mares. Once a donor mare was in estrus and had a >/=35 mm follicle, an ovariectomized recipient was selected and assigned to one of three groups: 1) 1 mg estradiol (E(2)) was injected subcutaneously daily until the donor mare ovulated; on the day of the donor mare's ovulation, daily intramuscular injections of 300 mg progesterone (P4) were commenced and continued until the end of the experiment (Day 35); 2) E(2) and P4 treatments were identical except E(2) was continued daily until Day 20; and 3) The same E(2) treatment as Group 1, 0.044 mg altrenogest per kilogram body weight were administered daily until Day 35. Embryos were recovered 7 d after the donor mare's ovulation and were transferred via surgical flank incision. Twenty additional embryos (controls) were transferred into intact recipients that ovulated 1 d before to 3 d after the donor. Pregnancy rates did not differ (P>0.05) among groups at Days 14 or 35. Pregnancy rates at Day 35 for mares administered injectable P4 (70%) were identical to those given altrenogest. Overall, pregnancy rates for ovariectomized-progestin treated recipients (28 of 40, 70%) were similar (>0.05) to that of intact mares (16 of 20, 80%). Dose of P4 was decreased in Groups 1 and 2 to 200 mg (Days 35 to 39), 100 mg (Days 40 to 44), 50 mg (Days 45 to 49) and 0 mg (>/=Day 50). Blood samples were collected once on Days 34, 35, 39, 40, 44, 45, 49 and 50 and assayed for P4. Dose of altrenogest was decreased to 0.022, 0.011, 0.0055 and 0 mg per kilogram body weight at Days 35 to 39, 40 to 44, 45 to 49 and >/=50. Number of mares in Groups 1 and 2 that lost their pregnancy while given 200, 100, 50 or 0 mg P4 was 0, 2, 8 and 4, respectively. Doses of 0.022, 0.011, 0.0055 and 0 mg altrenogest per kilogram body weight resulted in 0, 6, 4 and 3 mares aborting. Fetal death did not occur until concentrations of P4 decreased below 2.56 ng/ml 24 h after injection.     

Phylogenetic distribution in the genus Mus of t-complex-specific DNA and protein markers: inferences on the origin of t-haplotypes

We have examined the phylogenetic distribution of two t-specific markers among representatives of various taxa belonging to the genus Mus. The centromeric TCP-1a marker (a testicular protein variant specific for all t-haplotypes so far studied) has also been apparently detected in several non-t representatives of the Mus IVA, Mus IVB, and probably M. cervicolor species. By contrast, a t-specific restriction- fragment-length polymorphism allele (RFLP) of the telomeric alpha- globin pseudogene DNA marker alpha-psi-4 was found only in animals belonging to the M. musculus-complex species either bearing genuine t- haplotypes or, like the M. m. bactrianus specimen studied here, likely to do so. This t-specific alpha-psi-4 RFLP allele was found to be as divergent from the RFLP alleles of the latter, non-t, taxonomical groups as it is from Mus 4A, Mus 4B, or M. spretus ones. These results suggest the presence of t-haplotypes and of t-specific markers in populations other than those belonging to the M. m. domesticus and M. m. musculus subspecies, implying a possible origin for t-haplotypes prior to the radiation of the most recent offshoot of the Mus genus (i.e., the spretus/domesticus divergence), some 1-3 Myr ago.      

The effect of udder infection on the bacterial flora of the bulk milk of ten dairy herds

The significance of udder infection as a factor increasing the bacterial count of herd bulk milk was measured monthly for one year in ten dairy herds in Southern England. Staphylococcus aureus or mastitis streptococci were detected in 86% of samples, usually in numbers between 1000 and 10 000 c.f.u./ml of milk. However, in 8 and 2% of samples respectively greater than 20 000 or 100 000 c.f.u. of mastitis pathogens/ml of milk were detected. This occurred most commonly in the herds with a high incidence of Streptococcus uberis mastitis. The total bacterial counts of the herds' milks varied between 13 960 and 46 230 c.f.u./ml in the winter and between 6510 and 63 000 c.f.u./ml in the summer. No correlation was found between bacteriological quality of herd milk and the cleanliness of the milking machine and pipeline as assessed by plant rinses.     

Solubility of vesicular stomatitis virus M protein in the cytosol of infected cells or isolated from virions.

The peripheral membrane M protein of vesicular stomatitis virus purified by detergent extraction of virions and ion-exchange chromatography was determined to be a monomer in the absence of detergent at high salt concentrations. Reduction of the ionic strength below 0.2 M resulted in a rapid aggregation of M protein. This self-association was reversible by the detergent Triton X-100 even in low salt. However, aggregation was not reversible by high salt concentration alone. M protein is initially synthesized as a soluble protein in the cytosol of infected cells, thus raising the question of how the solubility of M protein is maintained at physiological ionic strength. Addition of radiolabeled M protein purified from virions to unlabeled cytosol from either infected or uninfected cells inhibited the self-association reaction. Cytosolic fractions from infected or uninfected cells were equally effective at preventing the self-association of M protein. Self-association could also be prevented by an irrelevant protein such as bovine serum albumin. Sedimentation velocity analysis indicated that most of the newly synthesized M protein is monomeric, suggesting that the solubility of M protein in the cytosol is maintained by either low-affinity interaction with macromolecules in the cytosol or interaction of a small population of M-protein molecules with cytosolic components.     

Mangrove zooplankton of North Queensland,Australia

Food consumption, growth, fish length distributions,population sizes and habitat use of the salmonids intwo lakes in the Høylandet area were studied in1986–89. The allopatric brown trout (Salmotrutta L.) in the tarn Røyrtjønna (27 ha) fed mainlyon organisms at the lake surface , crustaceanplankton, Trichoptera and Chironomidae. Only 5% ofthe trout reached an age of 6 years and a length of25 cm. Sexual maturation started at age 3 and a lengthof 14 cm. Through mark – recapture technique thenumber of trout >10 cm was estimated to 115 ha^-1.Growth, fish length frequencies and sexualmaturation of the sympatric brown trout and Arcticcharr (Salvelinus alpinus (L.)) in LakeStorgrønningen (530 ha) were not much different. TheStorgrønningen charr fed chiefly on zooplankton whichby volume represented 33% for the trout. The foodconsumption of Storgrønningen trout was at maximum inJuly with 2.06 mg food (d.w.) per g live fish and forcharr in September with 1.26 mg food. The maximumsize-independent growth rate of trout was 5.2%day^-1 in late June, and for charr 4.1%day^-1 in late July. Seventy percent of theirseasonal growth took place before 15 August. The charrstayed mainly deeper than 3-4 m, at water temperatures<15 °C. Brown trout stayed mainly the littoralzone and in near surface water of the pelagic. Thenumber of pelagic charr was estimated hydroacusticallyto 50 ind. ha^-1. The charr spawn in thelake. Mean numbers of juvenile trout in the twolargest tributaries were 26 and 48 per 100 m².Their annual length increment was 2.8–3.4 cm. Noindication of acidification or other human inducedimpacts were found. The lakes and their tributariesrepresent complex aquatic systems, representative forpristine oligotrophic Norwegian lowland lakes.John W. Jensen died shortly after easter in 1996     

Characterizing and correcting for the dynamic response of a bag-in-box system

A bag-in-box system (BBS) whose volume is monitored by a mechanical spirometer tends to have a slow response if the volume of the box is large, and this may significantly affect its measurement of gas flow. We describe a device for creating reproducible gas flows with which the impulse response of a BBS may be conveniently determined. Two computational techniques for correcting a BBS flow measurement for the effects of the impulse response were investigated: 1) an exponential model method that assumes a second-order model of the BBS dynamics and 2) a Fourier transform-based method of deconvolution known as Wiener filtering. Both correction methods produced a significant increase in the accuracy of BBS flow estimations, with the Wiener filter giving superior results.     

The contractile basis of amoeboid movement: V. The control of gelation, solation, and contraction in extracts from dictyostelium discoideum

Motile extracts have been prepared from Dictyostelium discoideum by homogenization and differential centrifugation at 4 degrees C in a stabilization solution (60). These extracts gelled on warming to 25 degrees Celsius and contracted in response to micromolar Ca++ or a pH in excess of 7.0. Optimal gelation occurred in a solution containing 2.5 mM ethylene glycol-bis (β-aminoethyl ether)N,N,N',N'-tetraacetate (EGTA), 2.5 mM piperazine-N-N'-bis [2-ethane sulfonic acid] (PIPES), 1 mM MgC1(2), 1 mM ATP, and 20 mM KCI at ph 7.0 (relaxation solution), while micromolar levels of Ca++ inhibited gelation. Conditions that solated the gel elicited contraction of extracts containing myosin. This was true regardless of whether chemical (micromolar Ca++, pH >7.0, cytochalasin B, elevated concentrations of KCI, MgC1(2), and sucrose) or physical (pressure, mechanical stress, and cold) means were used to induce solation. Myosin was definitely required for contraction. During Ca++-or pH-elicited contraction: (a) actin, myosin, and a 95,000-dalton polypeptide were concentrated in the contracted extract; (b) the gelation activity was recovered in the material sqeezed out the contracting extract;(c) electron microscopy demonstrated that the number of free, recognizable F-actin filaments increased; (d) the actomyosin MgATPase activity was stimulated by 4- to 10-fold. In the absense of myosin the Dictyostelium extract did not contract, while gelation proceeded normally. During solation of the gel in the absense of myosin: (a) electron microscopy demonstrated that the number of free, recognizable F- actin filaments increased; (b) solation-dependent contraction of the extract and the Ca++-stimulated MgATPase activity were reconstituted by adding puried Dictyostelium myosin. Actin purified from the Dictyostelium extract did not gel (at 2 mg/ml), while low concentrations of actin (0.7-2 mg/ml) that contained several contaminating components underwent rapid Ca++ regulated gelation. These results indicated : (a) gelation in Dictyostelium extracts involves a specific Ca++-sensitive interaction between actin and several other components; (b) myosin is an absolute requirement for contraction of the extract; (c) actin-myosin interactions capable of producing force for movement are prevented in the gel, while solation of the gel by either physical or chemical means results in the release of F-actin capable of interaction with myosin and subsequent contraction. The effectiveness of physical agents in producting contraction suggests that the regulation of contraction by the gel is structural in nature.     

Protein dynamics in sperm membranes: implications for sperm function during gamete interaction.

A number of mammalian sperm plasma membrane antigens have been implicated as playing a functional role in sperm-egg interaction, by virtue of the fact that antibodies against these antigens interfere with fertilization. Two such mouse sperm plasma membrane antigens are M42, a 200/220 kD glycoprotein doublet, and M5, a 150-160 kD glycoprotein. We show that both of these antigens are concentrated on the posterior region of caudal epididymal and capacitated mouse sperm heads and are relatively diffusible, as determined by fluorescence recovery after photobleaching measurements (D = 3-8 x 10(-9) cm2/s with approximately 23% diffusing). Crosslinking of these antigens with bivalent antibodies causes them to redistribute into the anterior region (acrosomal crescent) of the sperm head. In contrast, we describe a third antigen, P220, which is also localized to the posterior region of the sperm head on caudal epididymal sperm but which exhibits very little diffusion and does not redistribute upon crosslinking. Bivalent anti-M42 blocks the ZP3-induced acrosome reaction. We have found that monovalent Fab fragments of anti-M42 do not block the ZP3-induced acrosome reaction, but that inhibition is restored by addition of a second antibody which crosslinks the Fabs. Thus, crosslinking is required for both inhibition of the acrosome reaction and redistribution. This suggests that redistribution of antigen away from the posterior region of the head may be part of the mechanism of inhibition of the ZP3-induced acrosome reaction.