Reduced male fertility is common but highly variable in form and severity in a natural house mouse hybrid zone

Barriers to gene flow between naturally hybridizing taxa reveal the initial stages of speciation. Reduced hybrid fertility is a common feature of reproductive barriers separating recently diverged species. In house mice (Mus musculus), hybrid male sterility has been studied extensively using experimental crosses between subspecies. Here, we present the first detailed picture of hybrid male fertility in the European M. m. domesticus-M. m. musculus hybrid zone. Complete sterility appears rare or absent in natural hybrids but a large proportion of males (~30%) have sperm count or relative testis weight below the range in pure subspecies, and likely suffer reduced fertility. Comparison of a suite of traits related to fertility among subfertile males indicates reduced hybrid fertility in the contact zone is highly variable among individuals and ancestry groups in the type, number, and severity of spermatogenesis defects present. Taken together, these results suggest multiple underlying genetic incompatibilities are segregating in the hybrid zone, which likely contribute to reproductive isolation between subspecies.     

Calcium signaling during the plant-plant interaction of parasitic Cuscuta reflexa with its hosts

The plant parasite Cuscuta reflexa induces various responses in compatible and incompatible host plants. The visual reactions of both types of host plants including obvious morphological changes require the recognition of Cuscuta ssp. A consequently initiated signaling cascade is triggered which leads to a tolerance of the infection or, in the case of some incompatible host plants, to resistance. Calcium (Ca²⁺) release is the major second messenger during signal transduction. Therefore, we have studied Ca²⁺ spiking in tomato and tobacco during infection with C. reflexa. In our recently published study¹ Ca²⁺ signals were monitored as bioluminescence in aequorin-expressing tomato plants after the onset of C. reflexa infestation. Signals at the attachment sites were observed from 30 to 48 h after infection. In an assay with leaf disks of aequorin-expressing tomato which were treated with different C. reflexa plant extracts it turned out that the substance that induced Ca²⁺ release in the host plant was closely linked to the parasite''s haustoria.Key words: cuscut, odder, calcium signaling, plant parasitismThe genera Cuscuta, also known as dodder, includes 170 parasitic species with a worldwide distribution. Members of Cuscuta ssp. belong to the 1% of angiospermic plants that live as holoparasites and depend on nutrients, water and carbohydrates from other host plants.² Cuscuta spp. lack roots or leaves but possess specific penetrating organs, the so called haustoria, which are fully developed 5–6 days after the first contact, when an interaction between parasite and host is established.As for all dicotyledonous plants, the typical Cuscuta spp. life cycle starts with the germination of seeds. At the stage of a rootless seedling, Cuscuta ssp. has just a few days to find and successfully invade a host plant. Although Cuscuta ssp. seedlings appear to coil indiscriminately around any vertical elongated object, they seem to have an efficient “sense of smell” to recognize potential “victims” and are therefore able to infest host plants more rapidly and efficiently.³ As soon as a host is reachable, Cuscuta ssp. starts to wind around the host shoot and initiates the attachment process as well as the development of haustoria.^2,4 Already at this initial phase of infection (12–48 h post attachment) the host plant senses the parasite and initiates an onset of several signals which are only partially known. Amongst the several induced genes are for example those encoding AGPs (Arabinogalactan Proteins), proteins promoting the parasite''s adherence.⁵ Also proteins are produced which might be important for nutrient and water uptake⁶ or which modify the host cell wall.⁷In this addendum article, we focus on signals which occur in host plants within the early infection stage prior to a vascular bundle connection and refer to our article about Ca²⁺ signalling in C. reflexa infected tomato plants.⁸ Besides phytohormones or other initial signalling molecules, such cellular calcium signals might be involved in controlling the expression of important genes for developmental or resistance related processes.In our approach, Cuscuta reflexa shoots of ∼25 cm length were wrapped around transgenic constitutively aeqourin-expressing tomato (Solanum lycopersicum) and tobacco (Nicotiana benthamiana) plants. With a highly sensitive ccd-camera we then monitored the two interacting organisms. The Ca²⁺-signals which are released by the host-plant could be detected as light-emission. The first cytosolic calcium signals were observed 24–48 h after the parasite attachment when the haustoria formation was already initiated. Light, indicating a cytosolic calcium influx was clearly visible directly where the parasite started to penetrate host tissue via its haustoria (Fig. 1) and often appeared several times within 1–6 h. As the light signals per recorded picture were collected for 10 min it is not clear if the duration of such cytosolic calcium influx comprises 10 ms or 10 min. An additional experiment in our study was the usage of a Cuscuta reflexa haustorium extract which was applied to aequorin expressing tomato leaf discs. Here it turned out that the Ca²⁺-ion influx happened steadily and slowly, because signals were only visible when summed up from 2 h recording. The finding that both boiled haustoria extract and control extract, made from Cuscuta reflexa shoots without haustoria, are inactive, suggests that a protein which is expressed during the infection process might be the direct or indirect trigger of such Ca²⁺-signals. These results overcome furthermore the theory that Calcium signals are induced by pressure, which might also be a step during Cuscuta ssp. infection.Open in a separate windowFigure 1Cuscuta reflexa infection induces calcium-signals in aequorin-expressing tomato. Left: Bright field; middle: light emission representing Ca²⁺-signals at the infection site ∼30 h post onset of the parasite; signals were monitored with a ccd camera. Right: overlay.The fact that calcium fluxes act as a second messenger in several stress responses such as cold shock, wind, touch, osmotic stress,⁹ phytohormone signalling pathways,¹⁰ plant—symbiotic interactions^10–12 or also plant pathogen interaction^13–15 complicates the interpretation of the signals that are induced by Cuscuta reflexa. One possibility could be that visible Ca²⁺-signals are part of a signalling pathway where also SA (salicylic acid) or/and JA (jasmonic acid) play an important role. Recently, Runyon et al.¹⁶ could show that tomato plants infected with Cuscuta pentagona respond with a strong induction of JA and SA 24–36 h post infections. This time frame correlates with our described calcium signals and it has been previously described that calcium fluxes might be a part of the JA- and SA-signalling cascade.The tomato—dodder interaction, however, represents an exception among dicotyledonous plants because tomato generates a hypersensitive response which is part of a successful resistance reaction.^7,16 In this particular case characteristic components of C. reflexa must be sensed by its host plant. These factors indicate “non-self” for the host plant, probably following a model comparable to the MAMP concept where characteristic molecular patterns of a pathogen are recognized in host plants via pattern recognition receptors and subsequently trigger defence responses.^17,18 But sensing and signalling in host plants takes place not only in the case of an “incompatible” interaction. The developmental phenomenons of a dodder—plant interaction in a “compatible” interaction are nearly a miracle. In this case, the parasite is completely tolerated and achieves the attachment and the penetration of the host plant. It interferes in developmental processes and manipulates its host to develop vascular tissues, to build up chimerical cell walls and interspecific symplastic cell connections.^16,17 Finally, it is connected to the host plant and starts to withdraw nutrients and carbohydrates^19,20 by mimicking endogenous sinks. Such a tolerated interaction reminds of an interaction of plants with bacterial or fungal symbionts, where also Ca²⁺-signals have been described and well characterized.^11,12 In the case of Cuscuta ssp.—host interaction a lot of further studies have to be done to discover all important steps of signalling cascades.     

A Defined and Xeno-Free Culture Method Enabling the Establishment of Clinical-Grade Human Embryonic,Induced Pluripotent and Adipose Stem Cells

Background

The growth of stem cells in in vitro conditions requires optimal balance between signals mediating cell survival, proliferation, and self-renewal. For clinical application of stem cells, the use of completely defined conditions and elimination of all animal-derived materials from the establishment, culture, and differentiation processes is desirable.

Methodology/Principal Findings

Here, we report the development of a fully defined xeno-free medium (RegES), capable of supporting the expansion of human embryonic stem cells (hESC), induced pluripotent stem cells (iPSC) and adipose stem cells (ASC). We describe the use of the xeno-free medium in the derivation and long-term (>80 passages) culture of three pluripotent karyotypically normal hESC lines: Regea 06/015, Regea 07/046, and Regea 08/013. Cardiomyocytes and neural cells differentiated from these cells exhibit features characteristic to these cell types. The same formulation of the xeno-free medium is capable of supporting the undifferentiated growth of iPSCs on human feeder cells. The characteristics of the pluripotent hESC and iPSC lines are comparable to lines derived and cultured in standard undefined culture conditions. In the culture of ASCs, the xeno-free medium provided significantly higher proliferation rates than ASCs cultured in medium containing allogeneic human serum (HS), while maintaining the differentiation potential and characteristic surface marker expression profile of ASCs, although significant differences in the surface marker expression of ASCs cultured in HS and RegES media were revealed.

Conclusion/Significance

Our results demonstrate that human ESCs, iPSCs and ASCs can be maintained in the same defined xeno-free medium formulation for a prolonged period of time while maintaining their characteristics, demonstrating the applicability of the simplified xeno-free medium formulation for the production of clinical-grade stem cells. The basic xeno-free formulation described herein has the potential to be further optimized for specific applications relating to establishment, expansion and differentiation of various stem cell types.     

Population parameters for resistance to Fusarium graminearum and Fusarium verticillioides ear rot among large sets of early, mid-late and late maturing European maize (Zea mays L.) inbred lines

Infection of maize ears with Fusarium graminearum (FG) and Fusarium verticillioides (FV) reduces yield and quality by mycotoxin contamination. Breeding and growing varieties resistant to both Fusarium spp. is the best alternative to minimize problems. The objectives of our study were to draw conclusions on breeding for ear rot resistance by estimating variance components, heritabilities and correlations between resistances to FV and FG severity and to investigate different inoculation methods. In 2007 and 2008, three maturity groups (early, mid-late, late) each comprising about 150 inbred lines were tested in Germany, France, Italy, and Hungary according to their maturity group. They were silk channel inoculated by FG (early) and FV (all groups). In the late maturity group, additionally kernel inoculation was applied in a separate trial. The percentage of mycelium coverage on the ear was rated at harvest (0–100%). Significant (P < 0.01) genotypic variances of ear rot severity were found in all groups. Inoculation was superior to natural infection because of higher disease severities and heritabilities. In early maturing flints and dents, FG caused significantly (P < 0.01) higher ear rot severity than FV (61.7 and 55.1% FG vs. 18.2 and 11.1% FV ear rot severity, respectively). FV inoculation in Southern Europe (mid-late, late) resulted in similar means between 10.3 and 14.0%. Selection is complicated by significant (P < 0.01) genotype × environment interactions. Correlation between FG and FV severity was moderate in flints and dents (r = 0.59 and 0.49, respectively) but lines resistant to both fungi exist. We conclude that chances for selecting improved European elite maize material within the existing germplasms is promising by multi-environmental inoculation trials.     

What stabilizes the 314‐helix in β3‐peptides? A conformational analysis using molecular simulation

β‐Peptides are analogs of natural α‐peptides and form a variety of remarkably stable structures. Having an additional carbon atom in the backbone of each residue, their folded conformation is not only influenced by the side‐chain sequence but also and foremost by their substitution pattern. The precise mechanism by which the side chains interact with the backbone is, however, hitherto not completely known. To unravel the various effects by which the side chains influence the backbone conformation, we quantify to which extent the dihedral angles of a β³‐substited peptide with an additional methyl group on the central C_α‐atom can be regarded as independent degrees of freedom and analyze the distributions of these dihedral angles. We also selectively capture the steric effect of substituents on the C_α‐ and C_β‐atoms of the central residue by alchemically changing them into dummy atoms, which have no nonbonded interactions. We find that the folded state of the β³‐peptide is primarily stabilized by a steric exclusion of large parts of the unfolded state (entropic effect) and only subsequently by mutual dependence of the ψ‐dihedral angles (enthalpic effect). The folded state of β‐peptides is stabilized by a different mechanism than that of α‐peptides. Proteins 2010. © 2010 Wiley‐Liss, Inc.