Extended phenotypes on coral reefs: cryptic phenotypes modulate coral-vermetid interactions.

Phenotypic variation can lead to variation in the strength and outcome of species interactions. Variation in phenotypic traits can arise due to plastic responses to environmental stimuli, underlying genetic variation, or both, and may reflect differences in the focal organism or aspects of the extended phenotype (e.g., associated microbes). We used a reciprocal transplant experiment of Porites corals to evaluate the role of plasticity vs. heritable diversity on phenotypic traits and performance of corals that varied in their prior exposure to vermetid gastropods, an organism known to reduce coral growth and survival. We measured a suite of phenotypic traits associated with coral performance, many of which showed a plastic response to vermetid exposure. Vermetids decreased calcification of corals, increased microbial diversity, and shifted microbial composition. Most traits also showed a signature of previous exposure environment that persisted even when exposure was reversed: i.e., under the same conditions, corals naïve to vermetids had slower calcification rates, thicker tissues, higher Symbiodiniaceae densities, and different microbiomes than corals previously exposed to vermetids. We suggest the phenotypic differences are heritable, as reefs with and without vermetids were comprised of two different mitotypes, that revealed high, consistent genetic variation. Vermetids were only found on the fast-growing coral mitotype that was characterized by thin tissue, and that likely had a history of disturbance. As extended phenotypes can have community impacts, we suggest vermetid, in addition to microbes, are part of the extended community phenotype of these corals. Coral genotypes can establish different reef trajectories, with thin-tissue types more prone to disturbance and subsequent colonization by other species, like vermetids, which can further facilitate the degradation of coral reefs. The effects of the extended phenotype of species likely influence heterogeneity across landscapes as well as temporal differences in community composition.

Landscape edges shape dispersal and population structure of a migratory fish.

Many freshwater organisms have a life-history stage that can disperse through seawater. This has obvious benefits for colonization and connectivity of fragmented sub-populations, but requires a physiologically challenging migration across a salinity boundary. We consider the role of landscape boundaries between freshwater and seawater habitats, and evaluate their potential effects on traits and developmental histories of larvae and juveniles (i.e., dispersing life-history stages) of an amphidromous fish, Galaxias maculatus. We sampled juvenile fish on their return to 20 rivers in New Zealand: 10 rivers had abrupt transitions to the sea (i.e., emptying to an open coastline); these were paired with 10 nearby rivers that had gradual transitions to the sea (i.e., emptying into estuarine embayments). We reconstructed individual dispersal histories using otolith microstructure, otolith microchemistry, and stable isotope analysis. We found that fish recruiting to embayment rivers had distinct dispersal and foraging histories, were slower growing, smaller in size, and older than fish recruiting to nearby non-embayment rivers. Our results indicate that landscape edges can affect dispersal capabilities of aquatic organisms, potentially leading to divergent life-history strategies (i.e., limited- versus widespread-dispersal). Patterns also suggest that dispersal potential among landscape boundaries can create heterogeneity in the traits of individuals, with implications for metapopulation dynamics.

Born at the right time? A conceptual framework linking reproduction, development, and settlement in reef fish.

Parents are expected to make decisions about reproductive timing and investment that maximize their own fitness, even if this does not maximize the fitness of each individual offspring. When offspring survival is uncertain, selection typically favors iteroparity, which means that offspring born at some times can be disadvantaged, while others get lucky. The eventual fate of offspring may be further modified by their own decisions. Are fates of offspring set by birthdates (i.e., determined by parents), or can offspring improve upon the cards they've been dealt? If so, do we see adaptive plasticity in the developmental timing of offspring? We evaluate these questions for a coral reef fish (the sixbar wrasse, Thalassoma hardwicke) that is characterized by extreme iteroparity and flexible larval development. Specifically, we monitored larval settlement to 192 small reefs over 11 lunar months and found that most fish settled during new moons of a lunar cycle (consistent with preferential settlement on dark nights). Settlement was significantly lower than expected by chance during the full moon and last quarter of the lunar cycle (consistent with avoidance of bright nights). Survival after settlement was greatest for fish that settled during times of decreasing lunar illumination (from last quarter to new moon). Fish that settled on the last quarter of the lunar cycle were ~10% larger than fish that settled during other periods, suggesting larvae delay settlement to avoid the full moon. These results are consistent with a numerical model that predicts plasticity in larval development time that enables avoidance of settlement during bright periods. Collectively, our results suggest that fish with inauspicious birthdates may alter their developmental trajectories to settle at better times. We speculate that such interactions between parent and offspring strategies may reinforce the evolution of extreme iteroparity and drive population dynamics, by increasing the survival of offspring born at the "wrong" time by allowing them to avoid the riskiest times of settlement.

Production of cis-11-eicosenoic acid by Mortierella fungi.

AIM: To find cis-11-eicosenoic acid (20:1ω9, EA)-producing micro-organisms. METHODS AND RESULTS: We found EA-producing fungi by screening about 300 fungal strains and identified a major fatty acid accumulated in the Mortierella fungi as EA by means of GC-MS analysis. In particular, Mortierella chlamydospora CBS 529.75 produced a high amount of EA (36.3 mg g(-1) of dried cells) on cultivation at 28°C for 4 days and then at 12°C for 3 days. In the result of lipid analysis, most of the EA was a component of triacylglycerols, not phospholipids. CONCLUSION: We found that M. chlamydospora CBS 529.75 was the best producer for the microbial production of EA. SIGNIFICANCE AND IMPACT OF THE STUDY: EA is beneficial as a raw material for medical supplies and a moisturizing component of cosmetic creams. This is the first report of microbial production of EA.

Expression of novel carotenoid biosynthesis genes from Enterococcus gilvus improves the multistress tolerance of Lactococcus lactis.

AIMS: To determine whether the carotenoid production improves stress tolerance of lactic acid bacteria, the cloned enterococcal carotenoid biosynthesis genes were expressed in Lactococcus lactis ssp. cremoris MG1363, and the survival rate of carotenoid-producing engineered MG1363 strain under stress condition was investigated. METHODS AND RESULTS: We cloned carotenoid biosynthesis genes from yellow-pigmented Enterococcus gilvus. The cloned genes consisted of crtN and crtM and its promoter region were inserted into the shuttle vector pRH100, and the resulting plasmid was named pRC. The cloned crtNM was expressed using pRC in noncarotenoid-producing L. lactis ssp. cremoris MG1363. The expression of crtNM led to the production of C30 carotenoid 4,4'-diaponeurosporene. After exposure to 32 mmol l(-1) H2 O2 , low pH (1.5, acidified with HCl), 20% bile acid and 12 mg ml(-1) lysozyme, the survival rates of the MG1363 strain harbouring pRC were 18.7-, 6.8-, 8.8- and 4.4-fold higher, respectively, than those of MG1363 strain harbouring the empty vector pRH100. CONCLUSIONS: The expression of carotenoid biosynthesis genes from Ent. gilvus improves the multistress tolerance of L. lactis. SIGNIFICANCE AND IMPACT OF THE STUDY: First report of the improvement of multistress tolerance of lactic acid bacteria by the introduction of genes for carotenoid production.

Non-muscle cofilin is a component of tubulobulbar complexes in the testis.

Tubulobulbar complexes are finger-like structures that form at the interface between maturing spermatids and Sertoli cells prior to sperm release and at the interface between two Sertoli cells near the base of the seminiferous epithelium. They originate in areas previously occupied by actin filament-associated intercellular adhesion plaques known as ectoplasmic specializations. Actin filaments also are associated with tubulobulbar complexes where they appear to form a network, rather than the tightly packed bundles found in ectoplasmic specializations. Cofilin, a calcium-independent actin-depolymerizing protein, previously has been identified in the testis, but has not been localized to specific structures in the seminiferous epithelium. To determine if cofilin is found in Sertoli cells and is concentrated at actin-rich structures, we reacted fixed frozen sections of rat testis, fixed fragmented tissue, and blots of seminiferous epithelium with pan-specific and non-muscle cofilin antibodies. In addition, GeneChip microarrays (Affymetrix, Santa Clara, CA) were utilized to determine the abundance of mRNA for all cofilin isoforms in Sertoli cells. Using the monoclonal pan-specific cofilin antibody, we found specific labeling exclusively at tubulobulbar complexes and not at ectoplasmic specializations. On one-dimensional (1D) Western blots this antibody reacted monospecifically with one band, and on 2D blots reacted with two dots, which we interpret as phosphorylated and nonphosphorylated forms of a single cofilin isotype. Messenger RNA for non-muscle cofilin in Sertoli cells is about 8.5-fold higher than for muscle-type cofilin. To confirm that the non-muscle isoform of cofilin is present at tubulobulbar complexes, we used antibodies specific to non-muscle cofilin for immunofluorescent localization. As with the pan-specific antibody, we found that the non-muscle cofilin antibody exclusively labeled tubulobulbar complexes. Results presented here indicate that non-muscle cofilin is concentrated at tubulobulbar complexes. Our results also indicate that cofilin is not concentrated at ectoplasmic specializations.

Isolation and characterization of enterocin SE-K4 produced by thermophilic enterococci, Enterococcus faecalis K-4.

Enterococcus sp. K-4, with a bacteriocin-like activity against E. faecium, was isolated from grass silage in Thailand. Morphological, physiological, and phylogenetic studies clearly identified strain K-4 as a strain of E. faecalis. Strain K-4 produced a maximal amount of bacteriocin at 43-45 degrees C. We purified, for the first time, the bacteriocin produced at high temperature by E. faecalis to homogeneity, using adsorption on cells of the producer strain and reversed-phase liquid chromatography. The bacteriocin, designated enterocin SE-K4, is a peptide of about 5 kDa as measured by SDS-PAGE, and Mass spectrometry analysis found the molecular mass of 5356.2, which is in good agreement. The amino acid sequencing of the N-terminal end of enterocin SE-K4 showed apparent sequence similarity to class IIa bacteriocins. Enterocin SE-K4 was active against E. faecium, E. faecalis, Bacillus subtilis, Clostridium beijerinckii, and Listeria monocytogenes. Enterocin SE-K4 is very heat stable.

Stress tolerance in doughs of Saccharomyces cerevisiae trehalase mutants derived from commercial Baker's yeast.

Accumulation of trehalose is widely believed to be a critical determinant in improving the stress tolerance of the yeast Saccharomyces cerevisiae, which is commonly used in commercial bread dough. To retain the accumulation of trehalose in yeast cells, we constructed, for the first time, diploid homozygous neutral trehalase mutants (Deltanth1), acid trehalase mutants (Deltaath1), and double mutants (Deltanth1 ath1) by using commercial baker's yeast strains as the parent strains and the gene disruption method. During fermentation in a liquid fermentation medium, degradation of intracellular trehalose was inhibited with all of the trehalase mutants. The gassing power of frozen doughs made with these mutants was greater than the gassing power of doughs made with the parent strains. The Deltanth1 and Deltaath1 strains also exhibited higher levels of tolerance of dry conditions than the parent strains exhibited; however, the Deltanth1 ath1 strain exhibited lower tolerance of dry conditions than the parent strain exhibited. The improved freeze tolerance exhibited by all of the trehalase mutants may make these strains useful in frozen dough.

Variability in relative importance of determinants of reef fish recruitment.

Multiple processes can act together to determine abundance of organisms and structure of communities. Recently, appreciation of this fact has motivated development of conceptual and statistical frameworks that quantitatively assess the relative importance of multiple causal factors. However, little consideration has been given to variability in the "importance" of processes through space and time (i.e. robustness), which represents another facet of a process's importance. Here, I focused on populations of a coral reef fish (Thalassoma hardwicke) and used an existing analytical method to assess the relative importance of initial population inputs (larval supply) and subsequent juvenile mortality in determining the average abundances of juvenile fish populations in different locations and times. The relative importance of processes varied significantly both temporally and spatially across a range of scales, and indicate a need for future assessments of relative importance to incorporate this variability.

Novel detoxification of the trichothecene mycotoxin deoxynivalenol by a soil bacterium isolated by enrichment culture.

A mixed microbial culture capable of metabolizing deoxynivalenol was obtained from soil samples by an enrichment culture procedure. A bacterium (strain E3-39) isolated from the enrichment culture completely removed exogenously supplied deoxynivalenol from culture medium after incubation for 1 day. On the basis of morphological, physiological, and phylogenetic studies, strain E3-39 was classified as a bacterium belonging to the Agrobacterium-Rhizobium group. Thin-layer chromatographic analysis indicated the presence of one major and two minor metabolites of deoxynivalenol in ethyl acetate extracts of the E3-39 culture filtrates. The main metabolite was identified as 3-keto-4-deoxynivalenol by mass spectroscopy and 1H and 13C nuclear magnetic resonance analysis. The immunosuppressive toxicity of 3-keto-4-deoxynivalenol was evaluated by means of a bioassay based on the mitogen-induced and mitogen-free proliferations of mouse spleen lymphocytes. This compound exhibited a remarkably decreased (to less than one tenth) immunosuppressive toxicity relative to deoxynivalenol, indicating that the 3-OH group in deoxynivalenol is likely to be involved in exerting its immunosuppressive toxicity. Strain E3-39 was also capable of transforming 3-acetyldeoxynivalenol but not nivalenol and fusarenon-X.

Induction of actinorhodin production by rpsL (encoding ribosomal protein S12) mutations that confer streptomycin resistance in Streptomyces lividans and Streptomyces coelicolor A3(2).

A strain of Streptomyces lividans, TK24, was found to produce a pigmented antibiotic, actinorhodin, although S. lividans normally does not produce this antibiotic. Genetic analyses revealed that a streptomycin-resistant mutation str-6 in strain TK24 is responsible for induction of antibiotic synthesis. DNA sequencing showed that str-6 is a point mutation in the rpsL gene encoding ribosomal protein S12, changing Lys-88 to Glu. Gene replacement experiments with the Lys88-->Glu str allele demonstrated unambiguously that the str mutation is alone responsible for the activation of actinorhodin production observed. In contrast, the strA1 mutation, a genetic marker frequently used for crosses, did not restore actinorhodin production and was found to result in an amino acid alteration of Lys-43 to Asn. Induction of actinorhodin production was also detected in strain TK21, which does not harbor the str-6 mutation, when cells were incubated with sufficient streptomycin or tetracycline to reduce the cell's growth rate, and 40 and 3% of streptomycin- or tetracycline-resistant mutants, respectively, derived from strain TK21 produced actinorhodin. Streptomycin-resistant mutations also blocked the inhibitory effects of relA and brgA mutations on antibiotic production, aerial mycelium formation or both. These str mutations changed Lys-88 to Glu or Arg and Arg-86 to His in ribosomal protein S12. The decrease in streptomycin production in relC mutants in Streptomyces griseus could also be abolished completely by introducing streptomycin-resistant mutations, although the impairment in antibiotic production due to bldA (in Streptomyces coelicolor) or afs mutations (in S. griseus) was not eliminated. These results indicate that the onset and extent of secondary metabolism in Streptomyces spp. is significantly controlled by the translational machinery.

ADP-ribosylation of proteins in Bacillus subtilis and its possible importance in sporulation.

Endogenous ADP-ribosylation was detected in Bacillus subtilis, as determined in vitro with crude cellular extracts. The ADP-ribosylated protein profile changed during growth in sporulation medium, displaying a temporary appearance of two ADP-ribosylated proteins (36 and 58 kDa) shortly after the end of exponential growth. Mutants resistant to 3-methoxybenzamide, a known inhibitor of ADP-ribosyltransferase, were obtained, and a significant proportion (15%) were found to be defective in both sporulation and antibiotic production. These mutants failed to ADP-ribosylate the 36- and 58-kDa proteins. The parent strain also lost the ability to ADP-ribosylate these proteins when grown in the presence of 3-methoxybenzamide at a concentration at which sporulation but not cell growth was severely inhibited. Results from genetic transformations showed that the mutation conferring resistance to 3-methoxybenzamide, named brgA, was cotransformed with the altered phenotypes, i.e., defects in ADP-ribosylation and sporulation. spoOA and spoOF mutants displayed an ADP-ribosylation profile similar to that of the parent strain, but a spoOH mutant failed to ADP-ribosylate any proteins, including the 36- and 58-kDa proteins. The significance of protein ADP-ribosylation in sporulation was further indicated by the observation that ADP-ribosylation of the 36-kDa protein could be induced by treatment with decoyinine, an inhibitor of GMP-synthetase, and by amino acid limitation, both of which resulted in an immediate decrease in GTP pool size eventually leading to massive sporulation. We propose that a new sporulation gene, which presumably controls sporulation via ADP-ribosylation of certain functional proteins, exists.

Changes in patterns of ADP-ribosylated proteins during differentiation of Streptomyces coelicolor A3(2) and its development mutants.

Mutants resistant to 3-aminobenzamide, a known inhibitor of ADP-ribosyltransferase, were obtained from Streptomyces coelicolor A3(2). One (strain 27) was analyzed in detail. Mutant 27 had a reduced ADP-ribosyl-transferase activity, exhibited substantial changes from the wild type in ADP-ribosylated protein profile during cell aging, and was defective in producing aerial mycelium and antibiotics. A 92-kDa ADP-ribosylated protein disappeared at the onset of differentiation in the parent strain but was present in mutant 27. Four ADP-ribosylated proteins (39, 41, 43, and 46 kDa) appeared at the onset of differentiation in the parent strain but were missing in mutant 27. Failure to ADP-ribosylate these four proteins was detected when the parent strain was grown in the presence of subinhibitory amounts of 3-aminobenzamide. Genetic analysis showed that the mutation, named brgA, conferring resistance to 3-aminobenzamide, cosegregated with the altered phenotypes (i.e., defects in ADP-ribosylation and aerial mycelium formation) and was mapped to a new locus near uraA. The brgA mutants were nonconditionally deficient in producing aerial mycelium and antibiotics, as determined by using various media, and had a morphological and physiological phenotype quite different from that of a bldG mutant carrying a mutation which was previously mapped near uraA. Among the known bld mutants, bldA, bldD, and bldG mutants exhibited a ADP-ribosylated protein profile similar to that of the wild type, while like mutant 27, bldB, bldC, and bldH mutants failed to ADP-ribosylate certain proteins.

New polyenic antibiotics active against gram-positive and gram-negative bacteria. VIII. Construction of synthetic medium for production of mono-chloro-congeners of enacyloxins.

New antibiotics enacyloxins (ENXs) are a family of non-lactonic polyene antibiotics produced by Frateuria sp. W-315. For the production of antibiotics, we had to employ two-step fermentations, the first is the production of spent medium of Neurospora crassa and the second is the production of antibiotics by Frateuria. To simplify the production of antibiotics, systematic analyses have been done on the spent medium, and factors which affect the production of antibiotics characterized. From the above results, we constructed a new medium for antibiotic production. Moreover, we could get a new antibiotic named enacyloxin IIIa (1), C33H48O11NCl (m/z 669). 1 was deduced to be one of the congeners of enacyloxins because it was similar to ENX IIa or ENX IVa both in biological and physico-chemical properties. Chlorine of 1 could be replaced by bromine, biosynthetically, and the resultant bromine-containing antibiotic also showed an antibacterial activity comparable to 1.

New polyenic antibiotics active against gram-positive and gram-negative bacteria. IV. Structural elucidation of enacyloxin IIa.

The chemical structure of a unique polyenic antibiotic enacyloxin IIa (former name: fr. 2) produced by Frateuria (formerly Gluconobacter) sp. W-315 has been determined by extensive spectroscopic studies, in particular by NMR spectral analysis. It has a novel non-lactonic structure involving 3,4-dihydroxycyclohexanecarboxylic acid with a chlorine-containing polyenic and polyhydroxy acyl side chain attached as an ester to the 3-hydroxyl substituent of the acid.

Immune dysfunction expressed selectively on L3T4+ T cells in the tumor-bearing state.

Spleen cells from normal C3H/He or BALB/c mice generate cytotoxic T-lymphocyte (CTL) responses to both trinitrophenyl (TNP)-modified syngeneic cells (TNP-self) and allogeneic cells. In contrast, cells from these strains of mice bearing a syngeneic tumor failed to induce anti-TNP-self-CTL responses, although portions of the same responding cells generated comparable anti-allo-CTL responses to those induced by normal responding cells. Although anti-allo-CTL responses were inducible from only Lyt-2+ T-cell subset of responding cells from normal or tumor-bearing mice, induction of TNP-CTL responses required the participation of L3T4+ T-cell subset as well as Lyt-2+ CTL precursors. In addition, the fact that the addition of concanavalin A-stimulated culture supernatant to cultures of responding cells from tumor-bearing mice resulted in the induction of appreciable anti-TNP-self CTL responses demonstrated the defect of L3T4+ T-cell function in the tumor-bearing state. Such a functional defect was ascribed neither to the loss of L3T4+ T cells nor to the generation of suppressor cells in spleen cells of tumor-bearing mice. It was found on one hand that in vitro stimulation of antigen-presenting cell (APC)-depleted normal responding population with TNP-self prepared from cells of normal or tumor-bearing mice produced comparable anti-TNP CTL responses. On the other hand, APC-depleted responding cells from tumor-bearing mice were unable to induce anti-TNP CTL responses under conditions in which APC-depleted normal responding cells induced an effective TNP-CTL response. These results indicate that selective impairment of L3T4+ T cell-mediated immunity is induced in the tumor-bearing state and that such an impairment is ascribed to the dysfunction of L3T4+ T cells themselves, but not of APC required for the activation of L3T4+ T-cell subset.

Renal interaction of atrial natriuretic peptide with angiotensin II: glomerular and tubular effects.

1. The possible interactions between the renal effects of atrial natriuretic peptide (ANP) and angiotensin II (AII) were studied in normal sodium-replete human subjects. Recent investigations have suggested that ANP inhibits the pressor and volume-retaining effects of activation of the renin-angiotensin system. Thus, ANP may attenuate the effects of AII on renal haemodynamics or tubular transport. 2. ANP (0.1 micrograms/kg per min, 60 min) was intravenously infused into eight normal human subjects with and without pretreatment with enalapril (20 mg, per oral), an inhibitor of the converting enzyme, and during infusion of AII (10 mg/kg per min). 3. ANP infusion alone caused increases in the urine volume (from 96 +/- 23 to 229 +/- 44 mL/h, P less than 0.05) and urinary sodium excretion (from 11.5 +/- 1.6 to 20.9 +/- 4.2 mEq/h, P less than 0.05). These changes were accompanied by an increase in the glomerular filtration rate (from 127 +/- 9 to 158 +/- 9 mL/min, P less than 0.05). ANP infusion after enalapril administration lowered the mean blood pressure (from 76 +/- 2 to 71 +/- 3 mmHg, P less than 0.05) to a level similar to that observed during ANP infusion alone (from 84 +/- 2 to 74 +/- 2 mmHg, P less than 0.01), but did not result in a significant diuresis (from 139 +/- 23 to 174 +/- 51 mL/h) or natriuresis (from 19.7 +/- 2.5 to 14.3 +/- 3.4 mEq/h, P less than 0.05). This combined treatment with a converting enzyme inhibitor and ANP reduced both the glomerular filtration rate (160 +/- 9 to 141 +/- 10 mL/min) and the renal plasma flow (from 775 +/- 49 to 570 +/- 45 mL/min, P less than 0.01). 4. The antinatriuretic effects of exogenous AII were reversed by superimposed ANP infusion (urinary sodium excretion: from 4.8 +/- 1.0 to 24.3 +/- 5.2 mEq/h, P less than 0.01). Under these conditions, the glomerular filtration rate increased (from 114 +/- 6 to 156 +/- 7 mL/min, P less than 0.05) to levels similar to those observed with ANP infusion alone. In addition the increased tubular sodium reabsorption induced by AII was inhibited by concomitant ANP infusion (fractional proximal tubular sodium reabsorption: from 90.7 +/- 3.5 to 80.3 +/- 16.6%, P less than 0.05, fractional post-proximal tubular sodium reabsorption: from 91.5 +/- 9.8 to 87.6 +/- 8.8%, P less than 0.05).(ABSTRACT TRUNCATED AT 400 WORDS)

Role of angiotensin II in the renal response to atrial natriuretic peptide in normal subjects.

Atrial natriuretic peptide (ANP) has been shown to inhibit angiotensin II (Ang II)-induced steroidogenesis and vasoconstriction. To investigate the role of Ang II in the renal response to ANP, a synthetic ANP (0.1 micron/kg/min, 60 min) was infused for 1 h in eight subjects with or without pretreatment with an inhibitor of the converting enzyme, enalapril (20 mg, p.o.), or Ang II (10 ng/kg/min). ANP infusion alone caused increases in urinary volume, urinary sodium excretion, and glomerular filtration rate (GFR). However, enalapril treatment abolished these diuretic and natriuretic effects of ANP. In this group, GFR was decreased and no tubular effects, which was estimated by urinary excretion of sodium and phosphate, were observed. The anti-natriuretic effects of exogenous Ang II were reversed by concomitant ANP infusion, which inhibited both proximal and postproximal sodium reabsorption induced by Ang II without changing the GFR. These results indicate that endogenous Ang II plays an obligatory role in the natriuretic response to ANP and also suggested that ANP inhibits Ang II-stimulated tubular reabsorption of sodium.

Property of class I H-2 alloantigen-reactive Lyt-2+ helper T cell subset. Abrogation of its proliferative and IL-2-producing capacities by intravenous injection of class I H-2-disparate allogeneic cells.

The present study investigates the distinctiveness of Class I H-2 alloantigen-reactive Lyt-2+ helper/proliferative T cell subset in the aspect of tolerance induction. Primary mixed lymphocyte reactions (MLR) revealed that Lyt-2+ and L3T4+ T cell subsets from C57BL/6 (B6) mice were exclusively capable of responding to class I H-2 [B6-C-H-2bm1 (bm1)]- and class II H-2 [B6-C-H-2bm12 (bm12)]-alloantigens, respectively. Anti-bm12 MLR was not affected by i.v. injection of bm12 spleen cells into recipient B6 mice. In contrast, a single i.v. administration of bm1 spleen cells into B6 mice resulted in the abrogation of the capacity of recipient B6 spleen and lymph node cells to give anti-bm1 MLR. This suppression was bm1 alloantigen-specific, since lymphoid cells from B6 mice i.v. presensitized with bm1 cells exhibited comparable anti-bm12 primary MLR to that obtained by normal B6 lymphoid cells. Such tolerance was rapidly (24 h after the i.v. injection of bm1 cells) inducible and lasting for at shortest 3 wk. Addition of lymphoid cells from anti-bm1-tolerant B6 mice to cultures of normal B6 lymphoid cells did not suppress the proliferative responses of the latter cells, indicating that the tolerance is not due to the induction of suppressor cells but attributed to the elimination or functional impairment of anti-bm1 proliferative clones. The tolerance was also demonstrated by the failure of tolerant lymphoid cells to produce IL-2. It was, however, found that anti-bm1 CTL responses were generated by tolerant lymphoid cells which were unable to induce the anti-bm1 MLR nor to produce detectable level of IL-2. These results demonstrate that class I H-2 alloantigen-reactive Lyt-2+ Th cell subset exhibits a distinct property which is expressed by neither Lyt-2+ CTL directed to class I H-2 nor L3T4+ Th cells to class II H-2 alloantigens.