The Impact of Chlorophyll-Retention Mutations,d1d2 and cyt-G1, during Embryogeny in Soybean

The ultrastructural, physiological, and molecular changes in developing and mature seeds were monitored in a control line (Glycine max [L.] Merr., cv Clark) that exhibited seed degreening and two mutant lines (d1d2 and cyt-G1) that retained chlorophyll upon seed maturation. Ultrastructural studies showed that the control line had no internal membranes, whereas stacked thylakoid membranes were detected in the green seed from the mutant lines. Pigment analyses indicated that total chlorophyll was lowest in the mature seeds of the control line. Mature d1d2 and cyt-G1 seed had elevated Chl a and Chl b levels, respectively. In both control and mutant lines, Lhcb1, Lhcb2, and RbcS mRNAs were abundant in embryos prior to cotyledon filling, declined after the onset of storage protein accumulation, and were barely detectable or undetectable in all later stages of seed development. Therefore, the chlorophyll-retention phenotype must be a result of the alteration of a process that occurs after translation of photosynthesis-related mRNAs to stabilize apoprotein and pigment levels. Furthermore, different elements controlling either the synthesis or turnover of Chl a and Chl b must be impaired in the d1d2 and cyt-G1 lines. No reproducible differences in total leaf, embryonic, and chloroplast protein profiles and plastid DNAs could be correlated with the mutations that induced chlorophyll retention.     

Characterization of a novel plant poly(A) polymerase

We have purified and characterized poly(A) polymerases (PAPs) from Pisum sativum, Brassica juncea, and Zea mays. Through chromatography on DEAE-Sepharose and heparin-Sepharose, these PAPs copurified as a single enzyme along with RNPs that could provide RNA substrates for the enzyme. More extensive purification by chromatography on MonoQ resulted in the resolution of the PAPs into as many as three fractions. One of these (PAP-I) contained a 43-kDa polypeptide immunologically related to the yeast PAP, and two others (PAP-II and PAP-III) contained RNAs that could serve as substrates for polyadenylation. These fractions by themselves possessed little PAP activity, but mixtures containing combinations of these displayed substantial activity. Similar PAP factors (PAP-I and PAP-III) were identified after fractionation of extracts prepared from Brassica juncea and Zea mays. The factors from one plant were completely interchangeable with those from different plants. We conclude that the poly(A) polymerases present in vegetative plant tissues consist of more than one component. In this respect, they are substantially different from other reported plant, mammalian, and yeast PAPs.     

Condition factor and hepatosomatic index as estimates of energy status in male three-spined stickleback

Seasonal variation in direct and indirect measures of energy status was examined using estimates of glycogen, lipid and protein levels in a single cohort of male three-spined sticklebacks from an annual population collected each month over one complete year. Condition factor, somatic condition factor and hepatosomatic index (HSI) were calculated as indirect indices of energy status and the accuracy of these indirect measures as predictors of energy status was investigated. Results indicate that both condition factors were significant predictors of energy reserves (lipid, protein, glycogen and total energy), but that the proportion of variance accounted for was small. Both condition factors perform better as predictors of energy content per unit body weight. The HSI was a significant, but a weak predictor of total glycogen levels over the whole year. On a seasonal basis the relationship between HSI and energy reserves was highly variable. These indices are therefore poor predictors of energy reserves in male three-spined sticklebacks.     

Catalytic components of proteasomes and the regulation of proteinase activity

The proteasome (multicatalytic proteinase complex) is a large multimeric complex which is found in the nucleus and cytoplasm of eukaryotic cells. It plays a major role in both ubiquitin-dependent and ubiquitin-independent nonlysosomal pathways of protein degradation. Proteasome subunits are encoded by members of the same gene family and can be divided into two groups based on their similarity to the and subunits of the simpler proteasome isolated fromThermoplasma acidophilum. Proteasomes have a cylindrical structure composed of four rings of seven subunits. The 26S form of the proteasome, which is responsible for ubiquitin-dependent proteolysis, contains additional regulatory complexes. Eukaryotic proteasomes have multiple catalytic activities which are catalysed at distinct sites. Since proteasomes are unrelated to other known proteases, there are no clues as to which are the catalytic components from sequence alignments. It has been assumed from studies with yeast mutants that -type subunits play a catalytic role. Using a radiolabelled peptidyl chloromethane inhibitor of rat liver proteasomes we have directly identified RC7 as a catalytic component. Interestingly, mutants in Prel, the yeast homologue of RC7, have already been reported to have defective chymotrypsin-like activity. These results taken together confirm a direct catalytic role for these -type subunits. Proteasome activities are sensitive to conformational changes and there are several ways in which proteasome function may be modulatedin vivo. Our recent studies have shown that in animal cells at least two proteasome subunits can undergo phosphorylation, the level of which is likely to be important for determining proteasome localization, activity or ability to form larger complexes. In addition, we have isolated two isoforms of the 26S proteinase.     

Interspecies recombination In nature: a meningococcus that has acquired a gonococcal PIB porin

A. vaginal isolate of Neisseria has been reported to resemble Neisseria meningitidis in biochemical characteristics but to react with serological reagents that are specific to the PI porin from Neisseria gonorrhoeae. We have confirmed that this isolate has the biochemical attributes of a meningococcus and have shown that it clusters among meningococcal Isolates on a dendrogram based on isoenzyme variation within housekeeping enzymes from populations of N. meningitidis and N. gonorrhoeae. Furthermore, the sequences of the fbp and adk genes were typical of those of N. meningitidis and were distinct from those of N. gonorrhoeae. However, the porB gene was very similar to the por genes of N. gonorrhoeae isolates that express the PIB class of outer-membrane porin (differing from one gonococcal por allele at only a single nucleotide site), and was clearly distinct from the porB genes of N. meningitidis. The isolate therefore appears to be a typical meningococcus, except that its porB gene has been replaced with the por gene from a gonococcus.     

AT-1.1: A thymus-specific alloantigen of chickens

A thymocyte-specific alloantigen, designated AT (avian thymus) –1.1, has been detected in Cornell C strain (CS) and Obese strain (OS) chickens, the latter being a strain derived from CS which develops a spontaneous form of autoimmune thyroiditis (SAT). Antisera specific for this antigen were developed first in a turkey immunized with thymocytes from an OS chicken and, later, in AT-1.1-negative CS chickens immunized with AT-1.1-positive thymocytes. AT-1.1 was detected in 50–70% of cells in a thymus cell suspension, but was not seen on peripheral blood lymphocytes, erythrocytes, or cells from bursa, spleen, kidney, liver, or brain. It was present on thymocytes of chickens at all ages tested, from 1 day to 6 months of age. AT-1.1 was not detected in six chicken lymphoid tumor cell lines tested, and birds expressing it were found to be negative for the presence of Marek's disease viral antigens. Pedigree studies on 287 (OS × CS)F₂ chickens demonstrated that AT-1.1 is expressed in a dominant or codominant manner, and the gene coding for this antigen was not linked to the B (major histocompatibility) complex. The genetics and tissue distributions of AT-1.1 indicate that it differs from thymus cell surface antigens, avian or mammalian, previously described.     

Application of the Indirect Immunoperoxidase Stain Technique to the Flagella of Azospirillum brasilense

An indirect immunoperoxidase stain was used to demonstrate by electron microscopy that an antigenic difference exists between the polar flagellum and the lateral flagella of Azospirillum brasilense ATCC 29145.     

The genotypic distribution among non-insulin-dependent diabetes mellitus (NIDDM) patients of a restriction fragment length polymorphism

The genotypic distribution among patients of a marker allele, or restriction fragment length polymorphism (RFLP), can be used to determine the mode of inheritance of a disease-predisposing gene, if the association of the marker with the disease is sufficiently high. In the case of noninsulin-dependent diabetes mellitus (NIDDM), the RFLP in the 5'-flanking region of the human insulin gene does not allow discrimination between dominant and recessive modes of inheritance, or between any intermediate model. Also, it is demonstrated, in general, that the observation of a higher odds ratio for individuals with two copies of a marker allele than for individuals with at least one copy does not in itself imply a gene-dosage model.