Coelomomyces opifexi (Pillai & Smith). Coelomomycetaceae: Blastocladiales II. Experiments in sporangial germination

Summary Laboratory experiments on sporangial germination and zoospore activity in Coelomomyces opifexi which utilises a suparlittoral environment are described. Sporangial germination depends upon (a) salinity of the medium used and (b) whether the sporangia were derived from living or deceased larvae. Sporangia from living larvae germinated almost instantaneously in distilled, tap, brackish pond and sea water with a salinity of 4.2 There was only partial germination at a salinity of 17, and none at all in 35 (full sea water). Sporangia from deceased larvae required a conditioning of 7 days or more under moisture at 23°C or 28°C before germination. Sporangia from living and moribund larvae became thick-walled and darker when exposed to a salinity of 8.5 or higher. These, likewise, required a conditioning period for germination. The biological and ecological significance of these observations are discussed.

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Zusammenfassung Die in vitro-Versuche an das Keimen und die Zootätigkeit von der supralitteral lebenden spezies Coelomomyces opifexi sind hier beschrieben worden. Das sporenbeheltische Keimen ist abhängig von (a) der Salzhaltigkeit der Umgebung und (b) ob die Sporenbehalter von lebenden oder toten Puppen erhalten sind. Die von lebenden Puppen herstammenden Sporenbehalter keimen sofort in distilliertem, Leitings,- Brack- und 4.2 tigem Salzwasser, nur zum Teil in 17 tigem Salzwasser und gar nicht in Meereswasser (35). Die Wänder der von toten Puppen herstammende Sporenbehalter waren dichter und schwarzer und brauchten mindestens 7 Tage zum keimen in einer feuchten Umgebung von 23° bis zu 28°C. Wenn der Salzgehalt stieg über 8.5, so worden die Wänder beider Arten dichter und schwarzer und brauchten ebenso eine bedingte Periode zum keimen. Die biologische und ekologische Bedeutung dieser Beobachtungen sind diskutiert worden.

     

Effects of Lead on Adenylate Cyclase Activity in Rat Cerebral Cortex

Lead decreased in a dose dependent manner the basal AC activity in membranes of rat cerebral cortex (IC₅₀ = 2.5 ± 0.1 M). In membranes preincubated under basal conditions, AC activity was stimulated by approximately two and fourfold by 10 M Gpp(NH)p or forskolin, respectively. Under basal conditions, lead (3 M) inhibited enzyme activity up to 50%, but was not able to inhibit the Gpp(NH)p- or the forskolin-stimulated AC activity. However, in membranes preincubated with Gpp(NH)p (10 M), lead (3 M) had no significant effect on enzyme activity, but it partly blocked the stimulation of AC activity elicited by forskolin (10 M). In membranes preincubated with 10 M lead, the addition of 10 M Gpp(NH)p or forskolin in the incubation medium did not stimulate AC activity. However, when added together in the incubation medium Gpp(NH)p + forskolin produced an increase in enzyme activity. In membranes preincubated with 10 M lead + 10 M Gpp(NH)p, Gpp(NH)p (10 M) or forskolin (10 M) added alone or in combination to the incubation medium did not stimulate AC activity. Moreover, under these latter conditions lead had no further effect on enzyme activity. These results indicate that lead may interact with G-proteins and with the catalytic subunit of cerebral cortical AC to produce inhibition of the enzyme activity.     

Inheritance of mitochondrial DNA in lentil (Lens culinaris Medik.)

Restriction fragment analysis was used to examine the inheritance of lentil mitochondrial DNA (mtDNA) in F₁ and F₅ progeny from intrasubspecific (Lens culinaris ssp. culinaris) crosses and in F₁ progeny from intersubspecific (Lens culinaris ssp. orientalis x L. culinaris ssp. culinaris) crosses. Southern blots of digested parental and progeny DNA were hybridized to heterologous maize mtDNA probes specific to coxI and atp6 genes. Two restriction fragment polymorphisms separated L.c. ssp. culinaris Laird and Eston from L.c. ssp. culinaris ILL5588, and one restriction fragment polymorphism distinguished L.c. ssp. culinaris Laird and Eston from L.c. ssp. orientalis LO4. Twelve of 13 f₁ progeny and all F₅ progeny from the intrasubspecific crosses, and all F₁ progeny from intersubspecific crosses had only maternal mtDNA restriction fragments. One f₁ plant from an Eston x ILL5588 cross inherited mtDNA fragments from both parents. Nuclear DNA inheritance was biparental in all F₁ progeny.NRCC No. 38451     

Intracellular sites of the synthesis of sweet potato mitochondrial F1ATPase subunits

Summary Five subunits (-, -, -, - and -subunits) of the six -and -subunits) in the F₁ portion (F₁ATPase) of sweet potato (Ipomoea batatas) mitochondrial adenosine triphosphatase were isolated by an electrophoretic method. The - and -subunits were not distinguishable immunologically but showed completely different tryptic peptide maps, indicating that they were different molecular species. In vitro protein synthesis with isolated sweet potato root mitochondria produced only the -subunit when analyzed with anti-sweet potato F₁ATPase antibody reacting with all the subunits except the -subunit. Sweet potato root poly(A)⁺RNA directed the synthesis of six polypeptides which were immunoprecipitated by the antibody: two of them immunologically related to the -subunit and the others to the - and -subunits. We conclude that the -subunit of the F₁ATPase is synthesized only in the mitochondria and the -, - and -subunits are in the cytoplasm.     

α-Thalassemia and the production of different α chain variants in heterozygotes

The production of five chain variants (Hb G-Georgia, Hb St. Luke's, Hb Lloyd, Hb Montgomery, and Hb G-Philadelphia) in heterozygotes was evaluated through hematological observations, hemoglobin quantification, and biosynthetic studies. All heterozygotes for Hb St. Luke's and Hb Lloyd and most heterozygotes with Hb G-Georgia and Hb Montgomery had normal hematology and average / values of about 1.1. They were assigned a normal genotype (^G/), although the proportions of Hb St. Luke's and Hb G-Georgia were low (10 to 13%) and those of Hb Lloyd and Hb Montgomery twice as high (20%). Data from short-term incubations confirmed this genotype for some of these heterozygotes. Isolated Hb St. Luke's and Hb G-Georgia gave low ^G/ values (0.2 and 0.3) indicating that these Hb variants were defective at the level of Hb assembly. Isolated Hb Montgomery and Hb G-Philadelphia, however, gave higher ^G/ values of 0.6 and 0.8, respectively. A second type of variability existed among Hb G-Georgia (20 vs. 13%), Hb Montgomery (28 vs. 20%), and Hb G-Philadelphia (47 vs. 34%) heterozygotes, in whom the levels of Hb G differed. The occurrence of higher levels of these three chain heterozygosities was associated with hematological or biosynthetic evidence of a mild or moderate chain deficiency due to an -thalassemia-2 heterozygosity (^G/⁰ or ^0G/) or a homozygosity (^0G/⁰), respectively.This study was supported in part by USPHS Research Grants HLB-05168 and HLB-15158.     

Kinetics of growth and lactic acid production in continuous and batch culture

Summary In a lactic acid fermentation by Streptococcus faecalis, the specific consumption rate of glucose (v) and the specific production rate of lactic acid () were represented by the following simple equations as functions of the specific growth rate (): 1/=(1/) + 1/ = (1/) + By use of data from a batch culture, these two equations were derived from enzyme kinetics of the product inhibition. These equations were successfully applied to the results of batch culture and chemostat culture. In addition, calculation of ATP yield by these equations agreed with the experimental results better than the conventional Leudeking-Piret type equation, which includes two terms associated with growth and not with growth. Correspondence to: H. Ohara     

Mapping the α-globin genes in HB J Mexico carriers

Summary the organization of the -globin genes was studied by restriction endonuclease mapping, in subjects carrying the variant Hb J Mexico. A subject homozygous for Hb J synthesized both Hb J (about 55%) and Hb A and had two loci per chromosome. His restriction site map was found to be identical to that obtained with a normal DNA, except for a mutant Bgl II site which was observed on the Hb J chromosome proximal to the 5-locus. We have also mapped the DNA of a compound heterozygote for Hb J and -thalassemia, who synthesizes 38% Hb J and we have found a single gene corresponding to a –^3.7 haplotype on one chromosome and two genes, respectively ^J and ^A, on the other.     

Nucleotide sequence of the rrnB 16S ribosomal RNA gene from Mycoplasma capricolum

Summary The nucleotide sequences of the rrnB 16S ribosomal RNA gene and its 5-and 3-flanking regions from Mycoplasma capricolum have been determined. The coding sequence is 1521 base pairs long, being 21 base pairs shorter than that of the Scherichia coli 16S rRNA gene. The 16S rRNA sequence of M. capricolum reveals 74% and 76% identity with that of E. coli and Anacystis nidulans, respectively. The secondary structure model constructed from the M. capricolum 16S rRNA.gene sequence resembles that proposed for E. coli 16S rRNA. A large stem structure can be constructed between the 5- and 3-flanking sequences of the 16S rRNA gene. The flanking regions are extremely rich in AT.     

The α2 cDNA sequence of human haptoglobin carries a bacterial promoter functionalin vivo

Various constructions of human haptoglobin (Hp) cDNA coding either for the complete ^2FS precursor protein or only for the subunit have been placed under the control of the P_R promoter in the bacterial expression vector pCQV2 (Queen, 1983). In addition to the expected 45,000 dalton polypeptide synthesized after induction of the P_R promoter, the complete ^2FS constructions constitutively express a smaller polypeptide of 30,000 dalton corresponding to a truncated Hp protein. Computer analysis of the HpcDNA revealed the presence of two strong potential bacterial promoters (²P_F and ²P_S) located in the duplicated ^2FS sequence. Both Hp promoter signals are followed by potential mRNA start sites and ribosome binding sites at a compatible distance from initiation codons. In addition, the Hp² cDNA sequence, when fused upstream to the cDNA coding for ¹-antitrypsin, constitutively promotesin vivo the efficient expression of an hybrid protein specifically recognized by antibodies raised against ¹-antitrypsin or haptoglobin.     

Acclimation of the Sea Cucumber Apostichopus japonicus to Decreased Salinity at the Blastula and Gastrula Stages: Its Effect on the Desalination Resistance of Larvae at Subsequent Stages of Development

Apostichopus (= Stichopus) japonicus blastulae and gastrulae were acclimated for 18 h to salinities of 32 (control), 24 and 22 (the lower limit of the range of tolerance), and 20 (below the range of tolerance). Acclimation to 20 resulted in the appearance of teratic larvae, most of which subsequently died. Acclimation to 24, 22, and 20 led to a shift in the range of tolerance of the larvae at further stages of development. With a decrease in salinity, acclimated larvae developed more successfully than unacclimated larvae. Acclimated larvae attained the pentactula stage and settled at a salinity range of 32–20; unacclimated larvae, at 32–22. At different stages of development, acclimated larvae survived greater decreases in salinity than unacclimated larvae. The acclimation effects could be traced up to metamorphosis and settling, i.e., two weeks after the end of the acclimation process.     

Genetic analysis of two wheat cultivars, ‘Sonalika’ and ‘WL 711’ for reaction to leaf rust (Puccinia recondita)

Summary Genetic analysis for leaf rust reaction of two widely adapted cultivars, Sonalika and WL 711, has been done using 21 near isogenic Lr lines and rust culture IL004 — avirulent on the two cultivars and all the Lr lines used. The segregation pattern in the F₂ generation indicated the presence of a recessive gene in Sonalika and of a dominant gene in WL 711. These genes in cultivars Sonalika and WL 711 have been identified as Lr 11 and Lr 13, respectively. Gene Lr 13 is no longer effective in WL 711 but it continues to give field resistance in the backgrounds of Chris, Prelude and Thatcher. There has been no significant change in the virulence spectrum of the leaf rust pathogen in India with the release of WL 711. High susceptibility of WL 711 seems to be due to the evolution of more aggressive forms of the pathogen to this cultivar. The gene Lr 11, which behaves as a recessive in Sonalika, was effective against leaf rust when this cultivar was released. The high susceptibility of Sonalika is probably due to an increase in the frequency of race 77 virulent on Lr 11. Lr 11 has shown a dominance reversal in the background of Sonalika. Present results suggest that interaction of resistance genes with the background genotype must be studied for their effective use in breeding programme.     

High-level Expression of Maize <Emphasis Type="Italic">γ</Emphasis>-zein Protein in Transgenic Soybean (<Emphasis Type="Italic">Glycine max</Emphasis>)

Primers were developed for 118 microsatellites isolated from grape (Vitis vinifera) genomic libraries enriched for (AC)n repeats. Only one microsatellite sequence matched other grape SSR-sequences in the GeneBank database. Genotyping was carried out in the parental lines and four offspring of two pseudo-test-cross populations, Cabernet Sauvignon x Seyval and Chardonnay x Bianca, and a further six other grape genotypes (V. vinifera Sultanina, Merlot, Syrah, Müller-Thurgau, Vitis Regent and V. riparia Gloire de Montpellier). A total of 108 microsatellites showed easily scorable alleles and 100 of them segregated according to a configuration suitable for mapping in either cross. A further 8 SSRs, although unsuitable for mapping in those crosses, showed polymorphism in the other genotypes tested. This set of markers was used, along with 75 microsatellites of other repeat-types, to fingerprint 46 offspring of the cross Chardonnay x Bianca. For each full-sib, individual heterozygosity and distance in repeat units between pairs of alleles at each locus (mean d²) were calculated as a tool for predicting highly outbred recombinant individuals. Six microsatellites with segregation ratios significantly distorted towards the lack of homozygous sibs were identified and mapped to linkage groups LG 3 and LG 5. Estimation of heterozygosity at genome-wide level and genotyping at loci for which homozygous sibs are discriminated against are discussed for marker-assisted background selection in outcrossing grapevines.     

C(alpha) chemical shift tensors in helical peptides by dipolar-modulated chemical shift recoupling NMR

The C chemical shift tensors of proteins contain information on the backbone conformation. We have determined the magnitude and orientation of the C chemical shift tensors of two peptides with -helical torsion angles: the Ala residue in G*AL (=–65.7°, =–40°), and the Val residue in GG*V (=–81.5°, =–50.7°). The magnitude of the tensors was determined from quasi-static powder patterns recoupled under magic-angle spinning, while the orientation of the tensors was extracted from C–H and C–N dipolar modulated powder patterns. The helical Ala C chemical shift tensor has a span of 36 ppm and an asymmetry parameter of 0.89. Its ₁₁ axis is 116° ± 5° from the C–H bond while the ₂₂ axis is 40° ± 5° from the C–N bond. The Val tensor has an anisotropic span of 25 ppm and an asymmetry parameter of 0.33, both much smaller than the values for -sheet Val found recently (Yao and Hong, 2002). The Val ₃₃ axis is tilted by 115° ± 5° from the C–H bond and 98° ± 5° from the C–N bond. These represent the first completely experimentally determined C chemical shift tensors of helical peptides. Using an icosahedral representation, we compared the experimental chemical shift tensors with quantum chemical calculations and found overall good agreement. These solid-state chemical shift tensors confirm the observation from cross-correlated relaxation experiments that the projection of the C chemical shift tensor onto the C–H bond is much smaller in -helices than in -sheets.     

Location of β-amylase sequences in wheat and its relatives

Summary A -amylase cDNA clone isolated from barley has been used to locate -amylase encoding sequences on wheat, rye, and Aegilops umbellulata chromosomes by hybridisation to restriction endonuclease digested DNA obtained from wheat aneuploid and wheat-alien addition lines. Structural genes were identified on homoeologous group 4 and 5 chromosomes, confirming the results of isozyme studies. In addition, a further set of structural genes was found on homoeologous group 2 chromosomes. It is proposed that there are two homoeoallelic series, -Amy-1 on group 4 or 5 chromosomes, and -Amy-2 on group 2 chromosomes. Evidence is presented that each locus contains one or two -amylase structural genes, and it is suggested that the large number of isozymes seen upon IEF are due to post-translational modifications.     

Mutagenic effectiveness and efficiency of EMS,DES and gamma-rays in rice

Summary Data on chlorophyll mutation frequency after treatment with EMS, DES and gamma-rays and sequential administration of gamma-rays and the two alkylating agents in three varieties of rice have been used to work out quantitatively the effectiveness and efficiency of each mutagen and combination treatment. For effectiveness, the order is EMS > DES and for efficiency it is EMS > DES > gamma-rays. In some sequential treatments (Gamma-rays + DES in IR8 and Basmati; DES + gamma-rays in IR8 and Jhona; Gamma-rays + EMS in IR8 and Basmati; and EMS + gamma-rays in IR8, Jhona and Basmati) mutation frequency is more than additive (synergistic) but these treatments are decisively less efficient because of their relatively high injurious effects in the M₁. generation. EMS induces more albinas than gamma-rays do. The mutational spectrum patterns induced by gamma-rays and DES are alike. In general, combination treatments tend to increase the frequency of albinas over other types of chlorophyll mutants.     

Hexahydroindanone derivatives of steroids formed by Rhodococcus equi

Summary A mutant strain of Rhodococcus equi accumulates three metabolites from the androst-4-ene-3,17-dione or from its degradation intermediate, 3a-H-4(3'-propionic acid)-7a-methylhexahydro-1,5-indanedione (MEPHIP). These three metabolites are: 3a-H-4a(3'-propionic acid)-5-hydroxy-7a-methylhexahydro-1-indanone--lactone (HIL); 3a-H-4(3'-trans acrylic acid)-5-hydroxy-7a-methylhexahydro-1-indanone (^2'-5-hydroxy-MEPHIP); and 3a-H-4(3'-hydroxy-3'-propionic acid)-5-hydroxy-7a-methylhexahydro-1-indanone (3'-hydroxy-HIL). The behaviour of this mutant allows us to propose a pathway for degradation of the intermediates, methylperhydroindanone propionates. However, during this degradation, the side-chain propionate was eliminated by a-oxidation mechanism. Offprint requests to: A. Miclo     

Analysis of primary food sources and trophic relationships of aquatic animals in a mangrove-fringed estuary,Khung Krabaen Bay (Thailand) using dual stable isotope techniques

Stable carbon (¹³C) and nitrogen (¹⁵N) isotopes were used to elucidate primary food sources and trophic relationships of organisms in Khung Krabaen Bay and adjacent offshore waters. The three separate sampling sites were mangroves, inner bay and offshore. The ¹³C values of mangrove leaves were –28.2 to –29.4, seagrass –10.5, macroalgae –14.9 to –18.2, plankton –20.0 to –21.8, benthic detritus –15.1 to –26.3, invertebrates –16.5 to –26.0, and fishes –13.4 to –26.3. The ¹⁵N values of mangrove leaves were 4.3 to 5.7, seagrass 4.3, macroalgae 2.2 to 4.4, plankton 5.7 to 6.4 , benthic detritus 5.1 to 5.3, invertebrates 7.2 to 12.2 , and fishes 6.3 to 15.9. The primary producers had distinct ¹³C values. The ¹³C values of animals collected from mangroves were more negative than those of animals collected far from shore. The primary carbon sources that support food webs clearly depended on location. The contribution of mangroves to food webs was confined only to mangroves, but a mixture of macroalgae and plankton was a major carbon source for organisms in the inner bay area. Offshore organisms clearly derived their carbon through the planktonic food web. The ¹⁵N values of consumers were enriched by 3–4 relative to their diets. The ¹⁵N data suggests that some of aquatic animals had capacity to change their feeding habits according to places and availability of foods and as a result, individuals of the same species could be assigned to different trophic levels at different places.     

The effect of monoterpenes on astaxanthin production by a mutant ofPhaffia rhodozyma

Summary The total pigment and astaxanthin content ofPhaffia rhodozyma increased with increasing concentrations -pinene up to 500 l -pinene/l. Above this concentration the total pigment and astaxanthin content as well as the biomass production decreased. The addition of 500 l -pinene/l increased the total pigment content from 1652 g/g to 2201 g/g and the astaxanthin content from 1554 g/g to 1883 g/g. A sharp decrease in maximum specific growth rate occurred above 150 l -pinene/l.     

Synthetic substrate analogues for UDP-GlcNAc: Manα1-6R β(1-2)-N-acetylglucosaminyltransferase II. Substrate specificity and inhibitors for the enzyme

UDP-GlcNAc: Man1-6R (1-2)-N-acetylglucosaminyltransferase II (GlcNAc-T II; EC 2.4.1.143) is a key enzyme in the synthesis of complexN-glycans. We have tested a series of synthetic analogues of the substrate Man1-6(GlcNAc1-2Man1-3)Man-O-octyl as substrates and inhibitors for rat liver GlcNAc-T II. The enzyme attachesN-acetylglucosamine in 1-2 linkage to the 2-OH of the Man1-6 residue. The 2-deoxy analogue is a competitive inhibitor (K _i=0.13mm). The 2-O-methyl compound does not bind to the enzyme presumably due to steric hindrance. The 3-, 4- and 6-OH groups are not essential for binding or catalysis since the 3-, 4- and 6-deoxy and -O-methyl derivatives are all good substrates. Increasing the size of the substituent at the 3-position to pentyl and substituted pentyl groups causes competitive inhibition (K _i=1.0–2.5mm). We have taken advantage of this effect to synthesize two potentially irreversible GlcNAc-T II inhibitors containing a photolabile 3-O-(4,4-azo)pentyl group and a 3-O-(5-iodoacetamido)pentyl group respectively. The data indicate that none of the hydroxyls of the Man1-6 residue are essential for binding although the 2- and 3-OH face the catalytic site of the enzyme. The 4-OH group of the Man-O-octyl residue is not essential for binding or catalysis since the 4-deoxy derivative is a good substrate; the 4-O-methyl derivative does not bind. This contrasts with GlcNAc-T I which cannot bind to the 4-deoxy-Man- substrate analogue. The data are compatible with our previous observations that a bisectingN-acetylglucosamine at the 4-OH position prevents both GlcNAc-T I and GlcNAc-T II catalysis. However, in the case of GlcNAc-T II, the bisectingN-acetylglucosamine prevents binding due to steric hindrance rather than to removal of an essential OH group. The 3-OH of the Man1-3 is an essential group for GlcNAc-T II since the 3-deoxy derivative does not bind to the enzyme. The trisaccharide GlcNAc1-2Man1-3Man-O-octyl is a good inhibitor (K _i=0.9mm). The above data together with previous studies indicate that binding of the GlcNAc1-2Man1-3Man- arm of the branched substrate to the enzyme is essential for catalysis. Abbreviations: GlcNAc-T I, UDP-GlcNAc:Man1-3R (1-2)-N-acetylglucosaminyltransferase I (EC 2.4.1.101); GlcNAc-T II, UDP-GlcNAc:Man1-6R (1-2)-N-acetylglucosaminyltransferase II (EC 2.4.1.143); MES, 2-(N-morpholino)ethane sulfonic acid monohydrate.     

The association of the human ε-globin gene with the nuclear matrix: a reconsideration

The association of the human -globin gene with the nuclear matrix was studied in erythroid and non-erythroid cell lines. Using a high salt method to prepare histone depleted nuclei we studied the association of variety of fragments covering a 7.8 kb region which contains the human -globin gene. We furthermore studied the association of a set of DNA fragments covering the 13 kb human G/A-globin gene domain, the 16 kb /-globin gene domain and the 10 kb -globin gene domain with the nuclear matrix of K562 and Raji cells. The results show that all fragments studied are easily released from the nuclear matrix, indicating no specific association.Summarizing our results we could say that a region starting 5.7 kb 5 to the human -globin gene and ending 4 kb 3 to the human -globin gene seems to contain no attachment sites with the nuclear matrix of both erythroid and non-erythroid cells.