Dispersal of first “workers” in social wasps: Causes and implications of an alternative reproductive strategy

Many “workers” in north temperate colonies of the eusocial paper wasp Polistes fuscatus disappear within a few days of eclosion. We provide evidence that these females are pursuing an alternative reproductive strategy, i.e., dispersing to overwinter and become nest foundresses the following spring, instead of helping to rear brood on their natal nests. A female is most likely to stay and help at the natal nest (i.e., least likely to disperse) when it is among the first workers to emerge and when it emerges on a nest with more pupae (even though worker-brood relatedness tends to be lower in such colonies). The latter cause may result from the fact that pupae-laden nests are especially likely to survive, and thus any direct or indirect reproductive payoffs for staying and working are less likely to be lost. Disappearing females are significantly smaller than predicted if dispersal tendency was independent of body size (emergence order-controlled), suggesting that the females likely to be most effective at challenging for reproductive rights within the natal colony (i.e., the largest females) are also most likely to stay. Thus, early dispersal is conditional on a female’s emergence order, the maturity of its natal nest, and its body size. Finally, we present evidence that foundresses may actively limit the sizes of first-emerging females, perhaps to decrease the probability that the latter can effectively challenge foundresses for reproductive rights. The degree to which foundresses limit the size of first-emerging females accords well with the predictions of the theory of staying incentives.

High relatedness and inbreeding at the origin of eusociality in gall-inducing thrips

Within the haplodiploid eusocial gall-inducing thrips, a species-level phylogeny combined with genetic data for five eusocial species enables an inference of levels of relatedness and inbreeding values for lineages at the origin of eusociality. Character optimization using data from five eusocial species indicates that the lineage or lineages where eusociality is inferred to have originated exhibit relatedness of 0.64–0.92, and FIS of 0.33–0.64. The high inbreeding coefficients found in these eusocial thrips have increased relatedness among and within both sexes and have reduced the haplodiploidy-induced relatedness asymmetries [Hamilton, W. D. (1964) J. Theor. Biol. 7, 1–52]. These results indicate that unusually high relatedness is associated with the origin of eusociality, and they suggest a role for inbreeding in the evolution of bisexual helping.

Manipulation of the phenolic chemistry of willows by gall-inducing sawflies

The ability to induce galls on plants has evolved independently in many insect orders, but the adaptive significance and evolutionary consequences of gall induction are still largely unknown. We studied these questions by analyzing the concentrations of various plant defense compounds in willow leaves and sawfly galls. We found that the galls are probably nutritionally beneficial for the sawfly larvae, because the concentrations of most defensive phenolics are substantially lower in gall interiors than in leaves. More importantly, changes in chemistry occur in a similar coordinated pattern in all studied willow species, which suggests that the insects control the phenolic biosynthesis in their hosts. The resulting convergence of the chemical properties of the galls both within and between host species indicates that the role of plant chemistry in the evolution of host shifts may be fundamentally less significant in gallers than in other phytophagous insects.

A Carboniferous insect gall: insight into early ecologic history of the Holometabola.

Although the prevalence or even occurrence of insect herbivory during the Late Carboniferous (Pennsylvanian) has been questioned, we present the earliest-known ecologic evidence showing that by Late Pennsylvanian times (302 million years ago) a larva of the Holometabola was galling the internal tissue of Psaronius tree-fern fronds. Several diagnostic cellular and histological features of these petiole galls have been preserved in exquisite detail, including an excavated axial lumen filled with fecal pellets and comminuted frass, plant-produced response tissue surrounding the lumen, and specificity by the larval herbivore for a particular host species and tissue type. Whereas most suggestions over-whelmingly support the evolution of such intimate and reciprocal plant-insect interactions 175 million years later, we provide documentation that before the demise of Pennsylvanian age coal-swamp forests, a highly stereotyped life cycle was already established between an insect that was consuming internal plant tissue and a vascular plant host responding to that herbivory. This and related discoveries of insect herbivore consumption of Psaronius tissues indicate that modern-style herbivores were established in Late Pennsylvanian coal-swamp forests.

How caterpillar-damaged plants protect themselves by attracting parasitic wasps.

Parasitic and predatory arthropods often prevent plants from being severely damaged by killing herbivores as they feed on the plants. Recent studies show that a variety of plants, when injured by herbivores, emit chemical signals that guide natural enemies to the herbivores. It is unlikely that herbivore-damaged plants initiate the production of chemicals solely to attract parasitoids and predators. The signaling role probably evolved secondarily from plant responses that produce toxins and deterrents against herbivores and antibiotics against pathogens. To effectively function as signals for natural enemies, the emitted volatiles should be clearly distinguishable from background odors, specific for prey or host species that feed on the plant, and emitted at times when the natural enemies forage. Our studies on the phenomena of herbivore-induced emissions of volatiles in corn and cotton plants and studies conducted by others indicate that (i) the clarity of the volatile signals is high, as they are unique for herbivore damage, produced in relatively large amounts, and easily distinguishable from background odors; (ii) specificity is limited when different herbivores feed on the same plant species but high as far as odors emitted by different plant species and genotypes are concerned; (iii) the signals are timed so that they are mainly released during the daytime, when natural enemies tend to forage, and they wane slowly after herbivory stops.

Agrobacterium VirD2 protein interacts with plant host cyclophilins

Agrobacterium tumefaciens induces crown gall tumors on plants by transferring a nucleoprotein complex, the T-complex, from the bacterium to the plant cell. The T-complex consists of T-DNA, a single-stranded DNA segment of the tumor-inducing plasmid, VirD2, an endonuclease covalently bound to the 5′ end of the T-DNA, and perhaps VirE2, a single-stranded DNA binding protein. The yeast two-hybrid system was used to screen for proteins interacting with VirD2 and VirE2 to identify components in Arabidopsis thaliana that interact with the T-complex. Three VirD2- and two VirE2-interacting proteins were identified. Here we characterize the interactions of VirD2 with two isoforms of Arabidopsis cyclophilins identified by using this analysis. The VirD2 domain interacting with the cyclophilins is distinct from the endonuclease, omega, and the nuclear localization signal domains. The VirD2–cyclophilin interaction is disrupted in vitro by cyclosporin A, which also inhibits Agrobacterium-mediated transformation of Arabidopsis and tobacco. These data strongly suggest that host cyclophilins play a role in T-DNA transfer.

Tumor necrosis factor α plays a central role in immune-mediated clearance of adenoviral vectors

Adenovirus (Ad) gene transfer vectors are rapidly cleared from infected hepatocytes in mice. To determine which effector mechanisms are responsible for elimination of the Ad vectors, we infected mice that were genetically compromised in immune effector pathways [perforin, Fas, or tumor necrosis factor α (TNF-α)] with the Ad vector, Ad5-chloramphenicol acetyl transferase (CAT). Mice were sacrificed at 7–60 days postinfection, and the levels of CAT expression in the liver determined by a quantitative enzymatic assay. When the livers of infected mice were harvested 28 days postinfection, the levels of CAT expression revealed that the effectors most important for the elimination of the Ad vector were TNF-α > Fas > perforin. TNF-α did not have a curative effect on infected hepatocytes, as the administration of TNF-α to infected severe combined immunodeficient mice or to infected cultures in vitro had no specific effect on virus persistence. However, TNF-α-deficient mice demonstrated a striking reduction in the leukocytic infiltration early on in the infection, suggesting that TNF-α deficiency resulted in impaired recruitment of inflammatory cells to the site of inflammation. In addition, the TNF-deficient mice had a significantly reduced humoral immune response to virus infection. These results demonstrate a dominant role of TNF-α in elimination of Ad gene transfer vectors. This result is particularly important because viral proteins that disable TNF-α function have been removed from most Ad vectors, rendering them highly susceptible to TNF-α-mediated elimination.

Fluorescence-intensity distribution analysis and its application in biomolecular detection technology

A methodology, fluorescence-intensity distribution analysis, has been developed for confocal microscopy studies in which the fluorescence intensity of a sample with a heterogeneous brightness profile is monitored. An adjustable formula, modeling the spatial brightness distribution, and the technique of generating functions for calculation of theoretical photon count number distributions serve as the two cornerstones of the methodology. The method permits the simultaneous determination of concentrations and specific brightness values of a number of individual fluorescent species in solution. Accordingly, we present an extremely sensitive tool to monitor the interaction of fluorescently labeled molecules or other microparticles with their respective biological counterparts that should find a wide application in life sciences, medicine, and drug discovery. Its potential is demonstrated by studying the hybridization of 5′-(6-carboxytetramethylrhodamine)-labeled and nonlabeled complementary oligonucleotides and the subsequent cleavage of the DNA hybrids by restriction enzymes.

Concerted biosynthesis of an insect elicitor of plant volatiles

A variety of agricultural plant species, including corn, respond to insect herbivore damage by releasing large quantities of volatile compounds and, as a result, become highly attractive to parasitic wasps that attack the herbivores. An elicitor of plant volatiles, N-(17-hydroxylinolenoyl)-l-glutamine, named volicitin and isolated from beet armyworm caterpillars, is a key component in plant recognition of damage from insect herbivory. Chemical analysis of the oral secretion from beet armyworms that have fed on 13C-labeled corn seedlings established that the fatty acid portion of volicitin is plant derived whereas the 17-hydroxylation reaction and the conjugation with glutamine are carried out by the caterpillar by using glutamine of insect origin. Ironically, these insect-catalyzed chemical modifications to linolenic acid are critical for the biological activity that triggers the release of plant volatiles, which in turn attract natural enemies of the caterpillar.

The Stem-Loop Binding Protein (SLBP1) Is Present in Coiled Bodies of the Xenopus Germinal Vesicle

The stem-loop binding protein (SLBP1) binds the 3′ stem-loop of histone pre-mRNA and is required for efficient processing of histone transcripts in the nucleus. We examined the localization of SLBP1 in the germinal vesicle of Xenopus laevis oocytes. In spread preparations of germinal vesicle contents, an anti-SLBP1 antibody stained coiled bodies and specific chromosomal loci, including terminal granules, axial granules, and some loops. After injection of myc-tagged SLBP1 transcripts into the oocyte cytoplasm, newly translated myc-SLBP1 protein was detectable in coiled bodies within 4 h and in terminal and axial granules by 8 h. To identify the region(s) of SLBP1 necessary for subnuclear localization, we subcloned various parts of the SLBP1 cDNA and injected transcripts of these into the cytoplasm of oocytes. We determined that 113 amino acids at the carboxy terminus of SLBP1 are sufficient for coiled body localization and that disruption of a previously defined RNA-binding domain did not alter this localization. Coiled bodies also contain the U7 small nuclear ribonucleoprotein particle (snRNP), which participates in cleavage of the 3′ end of histone pre-mRNA. The colocalization of SLBP1 and the U7 snRNP in the coiled body suggests coordinated control of their functions, perhaps through a larger histone-processing particle. Some coiled bodies are attached to the lampbrush chromosomes at the histone gene loci, consistent with the view that coiled bodies in the oocyte recruit histone-processing factors to the sites of histone pre-mRNA transcription. The non-histone chromosomal sites at which SLBP1 is found include the genes coding for 5 S rRNA, U1 snRNA, and U2 snRNA, suggesting a wider role for SLBP1 in the biosynthesis of small non-spliced RNAs.

Assembly of Lampbrush Chromosomes from Sperm Chromatin

We have examined the behavior of demembranated sperm heads when injected into the germinal vesicle (GV) of amphibian oocytes. Xenopus sperm heads injected into Xenopus GVs swelled immediately and within hours began to stain with an antibody against RNA polymerase II (Pol II). Over time each sperm head became a loose mass of chromosome-like threads, which by 24–48 h resolved into individually recognizable lampbrush chromosomes (LBCs). Although LBCs derived from sperm are unreplicated single chromatids, their morphology and immunofluorescent staining properties were strikingly similar to those of the endogenous lampbrush bivalents. They displayed typical transcriptionally active loops extending from an axis of condensed chromomeres, as well as locus-specific “landmarks.” Experiments with [3H]GTP and actinomycin D demonstrated that transcription was not necessary for the initial swelling of the sperm heads and acquisition of Pol II but was required for maintenance of the lampbrush loops. Splicing was not required at any stage during formation of sperm LBCs. When Xenopus sperm heads were injected into GVs of the newt Notophthalmus, the resulting sperm LBCs displayed very long loops with pronounced Pol II axes, like those of the endogenous newt LBCs; as expected, they stained with antibodies against newt-specific proteins. Other heterologous injections, including sperm heads of the frog Rana pipiens and the zebrafish Danio rerio in Xenopus GVs, confirm that LBCs can be derived from taxonomically distant organisms. The GV system should help identify both cis- and trans-acting factors needed to convert condensed chromatin into transcriptionally active LBCs. It may also be useful in producing cytologically analyzable chromosomes from organisms whose oocytes do not go through a typical lampbrush phase or cannot be manipulated by current techniques.

Coilin Can Form a Complex with the U7 Small Nuclear Ribonucleoprotein

Coiled bodies (CBs) in the amphibian oocyte nucleus are spherical structures up to 10 μm or more in diameter, much larger than their somatic counterparts, which rarely exceed 1 μm. Oocyte CBs may have smaller granules attached to their surface or embedded within them, which are identical in structure and composition to the many hundreds of B-snurposomes found free in the nucleoplasm. The matrix of the CBs contains the diagnostic protein p80-coilin, which is colocalized with the U7 small nuclear ribonucleoprotein (snRNP), whereas the attached and embedded B-snurposomes contain splicing snRNPs. A few of the 50–100 CBs in the oocyte nucleus are attached to lampbrush chromosomes at the histone gene loci. By coimmunoprecipitation we show that coilin and the U7 snRNP can form a weak but specific complex in the nucleoplasm, which is dependent on the special U7 Sm-binding site. Under the same conditions coilin does not associate with the U1 and U2 snRNPs. Coilin is a nucleic acid-binding protein, as shown by its interaction with single-stranded DNA and with poly r(U) and poly r(G). We suggest that an important function of coilin is to form a transient complex with the U7 snRNP and accompany it to the CBs. In the case of CBs attached to chromosomes at the histone gene loci, the U7 snRNP is thus brought close to the actual site of histone pre-mRNA transcription.

Coilin Shuttles between the Nucleus and Cytoplasm In Xenopus Oocytes

Coiled bodies are discrete nuclear organelles often identified by the marker protein p80-coilin. Because coilin is not detected in the cytoplasm by immunofluorescence and Western blotting, it has been considered an exclusively nuclear protein. In the Xenopus germinal vesicle (GV), most coilin actually resides in the nucleoplasm, although it is highly concentrated in 50–100 coiled bodies. When affinity-purified anti-coilin antibodies were injected into the cytoplasm of oocytes, they could be detected in coiled bodies within 2–3 h. Coiled bodies were intensely labeled after 18 h, whereas other nuclear organelles remained negative. Because the nuclear envelope does not allow passive diffusion of immunoglobulins, this observation suggests that anti-coilin antibodies are imported into the nucleus as an antigen–antibody complex with coilin. Newly synthesized coilin is not required, because cycloheximide had no effect on nuclear import and subsequent targeting of the antibodies. Additional experiments with myc-tagged coilin and myc-tagged pyruvate kinase confirmed that coilin is a shuttling protein. The shuttling of Nopp140, NO38/B23, and nucleolin was easily demonstrated by the targeting of their respective antibodies to the nucleoli, whereas anti-SC35 did not enter the germinal vesicle. We suggest that coilin, perhaps in association with Nopp140, may function as part of a transport system between the cytoplasm and the coiled bodies.

Assembly of the Nuclear Transcription and Processing Machinery: Cajal Bodies (Coiled Bodies) and Transcriptosomes

We have examined the distribution of RNA transcription and processing factors in the amphibian oocyte nucleus or germinal vesicle. RNA polymerase I (pol I), pol II, and pol III occur in the Cajal bodies (coiled bodies) along with various components required for transcription and processing of the three classes of nuclear transcripts: mRNA, rRNA, and pol III transcripts. Among these components are transcription factor IIF (TFIIF), TFIIS, splicing factors, the U7 small nuclear ribonucleoprotein particle, the stem–loop binding protein, SR proteins, cleavage and polyadenylation factors, small nucleolar RNAs, nucleolar proteins that are probably involved in pre-rRNA processing, and TFIIIA. Earlier studies and data presented here show that several of these components are first targeted to Cajal bodies when injected into the oocyte and only subsequently appear in the chromosomes or nucleoli, where transcription itself occurs. We suggest that pol I, pol II, and pol III transcription and processing components are preassembled in Cajal bodies before transport to the chromosomes and nucleoli. Most components of the pol II transcription and processing pathway that occur in Cajal bodies are also found in the many hundreds of B-snurposomes in the germinal vesicle. Electron microscopic images show that B-snurposomes consist primarily, if not exclusively, of 20- to 30-nm particles, which closely resemble the interchromatin granules described from sections of somatic nuclei. We suggest the name pol II transcriptosome for these particles to emphasize their content of factors involved in synthesis and processing of mRNA transcripts. We present a model in which pol I, pol II, and pol III transcriptosomes are assembled in the Cajal bodies before export to the nucleolus (pol I), to the B-snurposomes and eventually to the chromosomes (pol II), and directly to the chromosomes (pol III). The key feature of this model is the preassembly of the transcription and processing machinery into unitary particles. An analogy can be made between ribosomes and transcriptosomes, ribosomes being unitary particles involved in translation and transcriptosomes being unitary particles for transcription and processing of RNA.