Finding the right mix: a framework for selecting seeding rates for cover crop mixtures.

Cover crop mixtures have the potential to provide more ecosystem services than cover crop monocultures. However, seeding rates that are typically recommended (i.e. seeding rate of monoculture divided by the number of species in the mixture) are non-optimized and often result in the competitive species dominating the mixture, and therefore limiting the amount of ecosystem services that are provided. We created an analytical framework for selecting seeding rates for cover crop mixtures that maximize multifunctionality while minimizing seed costs. The framework was developed using data from a field experiment, which included six response surface designs of two-species mixtures, as well as a factorial replacement design of three-species and four-species mixtures. We quantified intraspecific and interspecific competition among two grasses and two legume cover crop species with grass and legume representing two functional groups: pearl millet [Pennisetum glaucum (L.) R.Br.], sorghum sudangrass [Sorghum bicolor (L.) Moench × Sorghum sudanense (Piper) Stapf], sunn hemp (Crotalaria juncea L.), and cowpea [Vigna unguiculata (L.) Walp]. Yield-density models were fit to estimate intraspecific and interspecific competition coefficients for each species in biculture. The hierarchy from most to least competitive was sorghum sudangrass > sunn hemp > pearl millet > cowpea. Intraspecific competition of a less competitive species was the greatest when the biculture was composed of two species in the same functional group. Competition coefficients were used to build models that estimated the biomass of each cover crop species in three-species and four-species mixtures. The competition coefficients and models were validated with an additional nine site-years testing the same cover crop mixtures. The biomass of a species in a site-year was accurately predicted 69% of the time (low root mean square error, correlation > 0.5, not biased, r2 > 0.5). Applying the framework, we designed three-species and four-species mixtures by identifying relative seeding rates that produced high biomass with high species evenness (i.e. high multifunctionality) at low seed costs based on a Pareto front analysis of 10,418 mixtures. Accounting for competition when constructing cover crop mixtures can improve the ecosystem services provided, and such an advancement is likely to lead to greater farmer adoption.

To meet grand challenges, agricultural scientists must engage in the politics of constructive collective action.

Agriculture now faces grand challenges, with crucial implications for the global future. These include the need to increase production of nutrient-dense food, to improve agriculture's effects on soil, water, wildlife, and climate, and to enhance equity and justice in food and agricultural systems. We argue that certain politics of constructive collective action-and integral involvement of agricultural scientists in these politics-are essential for meeting grand challenges and other complex problems facing agriculture in the 21st century. To spur reflection and deliberation about the role of politics in the work of agricultural scientists, we outline these politics of constructive collective action. These serve to organize forceful responses to grand challenges through coordinated and cooperative action taken by multiple sectors of society. In essence, these politics entail (1) building bonds of affinity within a heterogenous network, (2) developing a shared roadmap for collective action, and (3) taking sustained action together. These emerging politics differ markedly from more commonly discussed forms of political activity by scientists, e.g., policy advisory, policy advocacy, and protest. We present key premises for our thesis, and then describe and discuss a politics of constructive collective action, the necessary roles of agricultural scientists, and an agenda for exploring and expanding their engagement in these politics.

Sex differences in verbal and performance IQ's of children undergoing open-heart surgery.

Boys with congenital heart defects had essentially normal Verbal and Performnance IQ's on preoperative and postoperative tests; but girls' Verbal IQ's were significantly lower than those of boys, and significantly lower than girls' own Performance IQ's. This sex difference among congenital heart cases reverses the usual finding that girls excel on Verbal tests.

Retroviral vector-mediated gene transfer into human hematopoietic progenitor cells.

The transfer of the human gene for hypoxanthine phosphoribosyltransferase (HPRT) into human bone marrow cells was accomplished by use of a retroviral vector. The cells were infected in vitro with a replication-incompetent murine retroviral vector that carried and expressed a mutant HPRT complementary DNA. The infected cells were superinfected with a helper virus and maintained in long-term culture. The production of progeny HPRT virus by the bone marrow cells was demonstrated with a colony formation assay on cultured HPRT-deficient, ouabain-resistant murine fibroblasts. Hematopoietic progenitor cells able to form colonies of granulocytes or macrophages (or both) in semisolid medium in the presence of colony stimulating factor were present in the nonadherent cell population. Colony forming units cloned in agar and subsequently cultured in liquid medium produced progeny HPRT virus, indicating infection of this class of hematopoietic progenitor cell.

Dissatisfaction encodes a tailless-like nuclear receptor expressed in a subset of CNS neurons controlling Drosophila sexual behavior.

The dissatisfaction (dsf) gene is necessary for appropriate sexual behavior and sex-specific neural development in both sexes. dsf males are bisexual and mate poorly, while mutant females resist male courtship and fail to lay eggs. Males and females have sex-specific neural abnormalities. We have cloned dsf and rescued both behavioral and neural phenotypes. dsf encodes a nuclear receptor closely related to the vertebrate Tailless proteins and is expressed in both sexes in an extremely limited set of neurons in regions of the brain potentially involved in sexual behavior. Expression of a female transformer cDNA under the control of a dsf enhancer in males leads to dsf-like bisexual behavior.

Activated T cells express a novel gene on chromosome 8 that is closely related to the murine ecotropic retroviral receptor.

A novel cDNA clone (20.5) which is differentially expressed between two closely related T-lymphoma cell clones was isolated by subtraction-enriched differential screening. SL12.4 cells, from which the cDNA was isolated, have characteristics of thymocytes at an intermediate stage in development. A sister cell clone derived from the same tumor, SL12.3, does not express this mRNA, has a distinct phenotype, and expresses fewer genes required for mature T-cell function. The cDNA sequence predicts a highly hydrophobic protein (approximately 49.5 kilodaltons) which contains seven putative membrane spanning domains. The gene was expressed on concanavalin A-activated T lymphocytes and was designated Tea (T-cell early activation gene). The Tea gene mapped to chromosome 8 and appeared to be conserved among mammalian and avian species. The Tea gene is distinct from, but bears extensive amino acid and DNA sequence similarity with, the murine ecotropic retroviral receptor which is encoded by the Rec-1 gene. Neither gene product displayed significant homology with other known transmembrane-spanning proteins. Thus, the Tea and Rec-1 genes establish a new family encoding multiple membrane-spanning proteins.

Stress differentially induces cationic amino acid transporter gene expression.

The amino acid l-arginine plays a central role in several adaptive metabolic pathways and we postulate that regulated L-arginine transport contributes to important physiological responses. The majority of L-arginine flux is mediated by transport system y+ that is encoded by at least three genes, Cat1, Cat2 and Cat3. Cat2 encodes two distinct protein isoforms (CAT2/CAT2a) that differ by 10-fold in their apparent substrate affinity. Cat2 transcription is controlled by four widely spaced promoters. The expression of CAT2/2a transcripts was tested in skeletal muscle and macrophages following specific stresses or activators. Unexpectedly, CAT2a transcripts accumulated in skeletal muscle in response to surgical trauma (hepatectomy and splenectomy) as well as food deprivation, although neither high affinity CAT2 nor CAT1 were detectably altered. Activated macrophages decreased CAT1 levels, but accumulated CAT2 and iNOS mRNA and protein with parallel kinetics suggesting that CAT2 mediated L-arginine transport might regulate the L-arginine:nitric oxide pathway. In macrophages, liver and skeletal muscle, the most distal CAT2 promoter was predominant. No change in promoter usage was apparent under any stress conditions tested nor was alternate splicing of the CAT2 transcript dictated by promoter usage. The differential regulation of the Cat genes indicates their encoded transporter proteins meet different requirements for cationic amino acids in the intact animal.

Female oxytocin gene-knockout mice, in a semi-natural environment, display exaggerated aggressive behavior.

Compared to results from a generation of neuropharmacological work, the phenotype of mice lacking the oxytocin (OT) peptide gene was remarkably normal. An important component of the current experiments was to assay OT-knockout (OTKO) and wild-type (WT) littermate control mice living under controlled stressful conditions designed to mimic more closely the environment for which the mouse genome evolved. Furthermore, our experimental group was comprised of an all-female population, in contrast to previous studies which have focused on all-male populations. Our data indicated that aggressive behaviors initiated by OTKO during a food deprivation feeding challenge were considerably more intense and diverse than aggressive behaviors initiated by WT. From the measures of continuous social interaction in the intruder paradigm, it emerged that OTKO mice were more offensively aggressive (attacking rumps and tails) than WT. In a test of parental behaviors, OTKO mice were 100% infanticidal while WT were 16% infanticidal and 50% maternal. Finally, 'alpha females' (always OTKO) were identified in each experiment. They were the most aggressive, the first to feed and the most dominant at nesting behaviors. Semi-natural environments are excellent testing environments for elucidating behavioral differences between transgenic mice and their WT littermates which may not be ordinarily discernible. Future studies of mouse group behavior should include examining female groupings in addition to the more usual all-male groups.

Mammalian integral membrane receptors are homologous to facilitators and antiporters of yeast, fungi, and eubacteria.

We demonstrate that three integral membrane receptors of mammals--the ecotropic retroviral leukemia receptor (ERR), the human retroviral receptor (HRR), and the T-cell early activator (Tea)--are homologous to a family of transporters specific for amino acids, polyamines, and choline (APC), which catalyze solute uniport, solute:cation symport, or solute:solute antiport in yeast, fungi, and eubacteria. Interestingly, the ERR membrane protein was recently shown to function as a cation:amino acid cotransporter. A binary sequence similarity matrix and an evolutionary tree of the 14 members of this family, illustrating their sequence similarities and divergences, were constructed. Other proteins, including the developmentally controlled GerAII spore germination protein of Bacillus subtilis and the acetylcholine receptor of Drosophila melanogaster gave sequence comparison scores of a sufficiently large magnitude to suggest (but not to establish) a common evolutionary origin with members of the APC family. We report an extended and corrected Tea cDNA sequence and show that the mammalian Tea and ERR encoding genes are differentially expressed in tissues and cell lines. Furthermore, the two mammalian cDNA sequences hybridize with other vertebrate and yeast genomic DNAs under stringent conditions. These observations support the notion that cell surface receptor proteins in mammals are transport proteins that share a common origin with transport proteins of single-celled organisms. Thus, permeases of essential metabolites may function pathologically as viral receptors.

Three-color imaging using fluorescent proteins in living zebrafish embryos.

The zebrafish embryo is especially valuable for cell biological studies because of its optical clarity. In this system, use of an in vivo fluorescent reporter has been limited to green fluorescent protein (GFP). We have examined other fluorescent proteins alone or in conjunction with GFP to investigate their efficacy as markers for multi-labeling purposes in live zebrafish. By injecting plasmid DNA containing fluorescent protein expression cassettes, we generated single-, double-, or triple-labeled embryos using GFP, blue fluorescent protein (BFP, a color-shifted GFP), and red fluorescent protein (DsRed, a wild-type protein structurally related to GFP). Fluorescent imaging demonstrates that GFP and DsRed are highly stable proteins, exhibiting no detectable photoinstability, and a high signal-to-noise ratio. BFP demonstrated detectable photoinstability and a lower signal-to-noise ratio than either GFP or DsRed. Using appropriate filter sets, these fluorescent proteins can be independently detected even when simultaneously expressed in the same cells. Multiple labels in individual zebrafish cells open the door to a number of biological avenues of investigation, including multiple, independent tags of transgenic fish lines, lineage studies of wild-type proteins expressed using polycistronic messages, and the detection of protein-protein interactions at the subcellular level using fluorescent protein fusions.

In vivo measurement accuracy in vital and necrotic canals with the Endex apex locator.

Currently apex locators are being used to determine working length. This study was undertaken to see what is actually being measured and if the pulp status, i.e. vital or necrotic, makes a difference in the determination. In this in vivo study, 33 teeth, both vital and necrotic, were measured by the Endex apex locator and then radiographed. After the length determination, the file was cemented to place, the tooth extracted, and then shaved back until the file and the apex were exposed. The position of the file was measured in relation to the apical foramen. Results indicate that all measurements were within a narrow range (-0.86 mm to 0.50 mm). There was no statistical difference in measurements between vital and necrotic canals.

A mutation in separase causes genome instability and increased susceptibility to epithelial cancer.

Proper chromosome segregation is essential for maintenance of genomic integrity and instability resulting from failure of this process may contribute to cancer. Here, we demonstrate that a mutation in the mitotic regulator separase is responsible for the cell cycle defects seen in the zebrafish mutant, cease&desist (cds). Analysis of cds homozygous mutant embryos reveals high levels of polyploidy and aneuploidy, spindle defects, and a mitotic exit delay. Carcinogenesis studies demonstrated that cds heterozygous adults have a shift in tumor spectrum with an eightfold increase in the percentage of fish bearing epithelial tumors, indicating that separase is a tumor suppressor gene in vertebrates. These data strongly support a conserved cross-species role for mitotic checkpoint genes in genetic stability and epithelial carcinogenesis.

Assembly of GMPCPP-bound tubulin into helical ribbons and tubes and effect of colchicine.

Microtubule assembly and disassembly is a complex structural process that does not proceed by simple addition and subtraction of individual subunits to and from a helical polymer, as would be the case for actin and other helical assemblies. The dynamic process of microtubule growth and shrinking involves short-lasting polymer forms that differ substantially from the microtubule itself and constitute crucial assembly and disassembly intermediates. Structural characterization thus depends on the stabilization of these brief intermediates and their preservation as polymeric assemblies. This paper gives experimental details on the polymerization of GMPCPP-tubulin into low-temperature, stable polymers that we propose to correspond to the early stages in microtubule assembly, and includes new data on the effect of colchicine on GMPCPP-tubulin polymerization. Finally, we add our thoughts on the possible biological meaning of tubulin polymerization versatility.

A zebrafish bmyb mutation causes genome instability and increased cancer susceptibility.

A major goal of cancer research has been to identify genes that contribute to cancer formation. The similar pathology between zebrafish and human tumors, as well as the past success of large-scale genetic screens in uncovering human disease genes, makes zebrafish an ideal system in which to find such new genes. Here, we show that a zebrafish forward genetic screen uncovered multiple cell proliferation mutants including one mutant, crash&burn (crb), that represents a loss-of-function mutation in bmyb, a transcriptional regulator and member of a putative proto-oncogene family. crb mutant embryos have defects in mitotic progression and spindle formation, and exhibit genome instability. Regulation of cyclin B levels by bmyb appears to be the mechanism of mitotic accumulation in crb. Carcinogenesis studies reveal increased cancer susceptibility in adult crb heterozygotes. Gene-expression signatures associated with loss of bmyb in zebrafish are also correlated with conserved signatures in human tumor samples, and down-regulation of the B-myb signature genes is associated with retention of p53 function. Our findings show that zebrafish screens can uncover cancer pathways, and demonstrate that loss of function of bmyb is associated with cancer.

y(+)-type cationic amino acid transport: expression and regulation of the mCAT genes.

The transport of cationic amino acids across animal cell membranes is largely mediated by a small group of well-described transport system (y+, bo,+, Bo,+). Only recently have genes encoding transport proteins in some of these systems been isolated. Two genes, mCAT-1 and mCAT-2, encode related multiple membrane-spanning proteins that share substantial amino acid sequence identity and virtually superimposable hydrophilicity profiles. mCAT-1 and mCAT-2 proteins expressed in Xenopus oocytes are functionally indistinguishable and similar to transport system y+, but have distinct tissue distribution patterns. mCAT-1 expression is nearly ubiquitous and produces a single protein, while mCAT-2 is highly tissue-specific, has two distinct protein isoforms encoded by a single gene and is expressed in different tissues using at least two widely separated promoters. All three proteins facilitate the ion-independent transport of arginine, lysine and ornithine. Both mCAT-1 and mCAT-2 proteins have low amino acid sequence similarity but strikingly similar hydrophilicity profiles with amino acid antiporters, uniporters and symporters of yeast, fungi and eubacteria. Current work will elucidate whether any of the mCAT proteins interact with members of a newly identified family of single membrane-spanning proteins, such as rBAT, 4F2 and NAA-Tr, which are thought to modulate or activate y+L and/or bo,+ transport systems.