Plants contain highly divergent actin isovariants

Actin protein isovariants have been identified in animals with distinct cytoplasmic or muscle specific patterns of expression. Analysis of vascular plant actin gene sequences suggests that an even greater diversity should exist within the plant actin protein families, but previous studies on plant proteins have not demonstrated the presence of multiple actin isovariants. Antibodies recognizing a conserved amino-terminal plant actin peptide, a family of plant actin peptides from a variable region, and two monoclonal antibodies to conserved epitopes within animal actins were used to identify isovariants of soybean actin resolved by two-dimensional isoelectric focusing (IEF) sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Approximately six to eight actin isovariants with pI values ranging from 5.1 to 5.8 have been identified from soybean hypocotyls, stems, leaves, and roots with varying amounts of most isovariants present in all four organs. Acidic isovariants were present in much higher levels in leaves and stems. Antisera with lambda-class actin specificity detected a subset of three isovariants in all organs examined. One monoclonal and one antipeptide antisera are shown to react well with a wide variety of plant actin isovariants. Similar patterns of actin isovariants were detected in the distant angiosperms, Arabidopsis, petunia, and maize. It is likely that many of these diverse classes of isovariants have been preserved throughout vascular plant evolution and reflect the ancient diversity within plant actin gene families. The extreme difference among isovariants implies the presence of a complex actin-based cytoskeletal system in plants.     

Divergence and differential expression of soybean actin genes

DNA sequence analysis as well as genomic blotting experiments using cloned soybean actin DNA sequences as probes show that large sequence heterogeneity exists among members of the soybean actin multigene family. This heterogeneity suggested that the members of this family might be diverged in function and/or regulation. Five of the six soybean actin gene family members examined are shown to be significantly more diverged from one another than members of other known actin gene families. This high level of divergence was utilized in the preparation of actin gene-specific probes in the analysis of the complexity and expression of these members of the soybean actin gene family. Hybridization studies indicate that the six soybean actin genes fall into three classes with a pair of genes in each class. These six genes account for all but two actin gene fragments detected in the soybean genome. We have compared the relative steady state mRNA levels of these classes of soybean actin genes in three organs of soybean. We find that actin genes SAc6 and SAc7 are most highly expressed accounting for 80% of all actin mRNA with respect to the six soybean actin genes examined. Actin genes SAc3 and SAc1 are expressed at intermediate and low levels respectively; and SAc2 and SAc4 are expressed at barely detectable levels. Four of the six soybean actin genes appear to be expressed at the same level in root, shoot and hypocotyl. SAc3 and SAc7 genes appear to be more highly expressed in shoot and 2,4-dichlorophenoxyacetic acid-induced hypocotyl than in root and hypocotyl.(ABSTRACT TRUNCATED AT 250 WORDS)     

THE QUANTITATIVE GENETICS OF SEXUAL DIMORPHISM IN SILENE LATIFOLIA (CARYOPHYLLACEAE). II. RESPONSE TO SEX-SPECIFIC SELECTION

A well-established theoretical relationship exists between genetic correlations between the sexes and the dynamics of response to sex-specific selection. The present study investigates the response to sex-specific selection for two sexually dimorphic traits that have been documented to be genetically variable, calyx diameter and flower number, in Silene latifolia. Following the establishment of a base generation with a known genetic background, selection lines were established and two generations of sex-specific selection were imposed. Calyx diameter responded directly to sex-specific selection, and the positive genetic correlation between the sexes was reflected in correlated responses in the sex that was not the basis for selection within a particular line. Flower number showed a more erratic response to sex-specific selection in that selection in some lines was initially in the wrong direction, that is, selection for a decrease in flower number resulted in an increase. These erratic responses were attributable to genotype-environment interaction as reflected in significant heteroscedasticity in variance among families. Correlated responses to selection in the sex that was not the immediate basis for selection indicated the possible existence of a negative genetic correlation between the sexes for this trait. These results test for the first time the impact of genetic correlations between the sexes on the evolutionary dynamics of sexually dimorphic traits in a plant species.     

Structure, Expression and Phylogenetic Analysis of the Gene Encoding Actin I in Pneumocystis Carinii

Actin is a major component of the cytoskeleton and one of the most abundant proteins found in eukaryotic cells. Comparative sequence analysis shows that this essential gene has been highly conserved throughout eukaryotic evolution making it useful for phylogenetic analysis. Complete cDNA clones for the actin-encoding gene were isolated and characterized from Pneumocystis carinii purified from immunosuppressed rat lungs. The nucleotide sequence encodes a protein of 376 amino acids. The predicted actin protein of P. carinii shares a high degree of conservation to other known actins. Only one major actin gene was found in P. carinii. The P. carinii actin sequence was compared with 30 other actin sequences. Gene phylogenies constructed using both neighbor-joining and protein parsimony methods places the P. carinii actin sequence closest to the majority of the fungi. Since the phylogenetic relationship of P. carinii to fungi and protists has been questioned, these data on the actin gene phylogeny support the grouping of P. carinii with the fungi.     

In vivo analysis of plant RNA structure: soybean 18S ribosomal and ribulose-1,5-bisphosphate carboxylase small subunit RNAs

A method to investigate the structure of RNA molecules within intact plant tissues has been developed. The RNA structures are analyzed using dimethyl sulfate (DMS), which modifies substituents of adenine and cytosine residues within single-stranded regions of RNA molecules. Reactive sites are identified by primer extension analysis. Using this procedure, an analysis of the secondary structure of the cytoplasmic 18S ribosomal RNA in soybean seedling leaves has been completed. DMS modification data are in good agreement with the phylogenetic structure predicted for soybean 18S rRNA. However, there are a few notable exceptions where residues thought to be involved in double-stranded regions in all 18S rRNAs are strongly modified in soybean leaf samples. These data taken together with the phylogenetic structure suggest that alternate structures may exist in vivo.The further applicability of this technique is demonstrated by comparing the modification pattern obtained in vivo to that obtained in vitro for a particular mRNA molecule encoding the small subunit of ribulose-1,5-bisphosphate carboxylase. The results obtained are compared to a predicted minimum energy secondary structure. The data indicate that the conformation of RNA molecules within the cell may not be reflected in a structural analysis of purified mRNA molecules.     

Single pulses of cytoplasmic calcium associated with phagocytosis of individual zymosan particles by macrophages

We have measured cytosolic free calcium levels ([Ca++]i) in individual macrophages during the phagocytosis of single zymosan particles. We report here that the contact of a macrophage with zymosan results in a rapid transient elevation of [Ca++]i. Each [Ca++]i pulse is symmetrical lasting for up to 30 seconds. In contrast, macrophage spreading is associated with repetitive [Ca++]i spiking occurring in salvos of up to four smaller spikes, each lasting for between 8 and 18 seconds. These qualitative and kinetic differences might suggest that the role of [Ca++]i in phagocytosis is distinct from its role in spreading.     

Ethanol tolerance and carbohydrate metabolism in lactobacilli

Summary This paper describes the ethanol tolerance and metabolism of 31 strains ofLactobacillus on glucose, xylose, lactose, cellobiose and starch. The purpose of this work was to determine the suitability of the 31 strains as potential host for the ethanol producing genes, pyruvate decarboxylase and aldehyde dehydrogenase, fromZymomonas mobilis. The 31 strains were screened for their ability to grow in 0 to 8% v/v ethanol on all five carbohydrates. Those strains that were able to grow to an OD of 1.0 in 8% ethanol were evaluated at ethanol concentrations up to 16%. v/v. The fermentative products from the five carbohydrates were analyzed to determine the ratios of lactic acid, ethanol, and acetic acid.Published as Paper No. 9786, Journal Series Nebraska Agricultural Experiment Station, Lincoln, NE 68583-0704.     

Does seawater acidification affect zooxanthellae density and health in the invasive upside‐down jellyfish,Cassiopea spp.?

Ocean acidification is the decline in seawater pH that results from the absorption of atmospheric carbon dioxide (CO₂). Decreased pH has negative effects on survivability, growth, and development in many marine calcifiers, potentially resulting in reduced coral species richness. This reduction in richness could open new niche space, allowing the spread of invasive species, such as the upside‐down jellyfish (Cassiopea spp.). Like corals, this jellyfish forms symbiotic relationships with zooxanthellae, photosynthetic dinoflagellates. This study focused on the effect of seawater acidification in Cassiopea spp. We monitored zooxanthellae density and two measures of health (bell diameter and volume) in individuals of Cassiopea sp. at three pH levels chosen to mimic different open‐ocean average conditions: 8.2, representing pre‐industrial revolution conditions; and 7.9 and 7.6, representing predicted declines in pH in the next century. Zooxanthellae density and health of the jellyfish were measured twice—prior to experimental manipulations and after four weeks of exposure to experimental pHs—in three consecutive trials. The effects of pH and Trial on proportional change in jellyfish attributes were analyzed using generalized linear mixed models. We found no significant effects of either factor. These results indicate that decreasing seawater pH has no apparent negative effect on zooxanthellae density or health in Cassiopea, which suggests that these jellyfish may be relatively insensitive to the impacts of ocean acidification, heightening its potential as an invasive species.