Membrane renewal after dibucaine deciliation of Tetrahymena: Freeze-fracture technique,cilia, membrane structure

Axenic late log phase cultures of Tetrahymena pyriformis DN-B3 are deciliated by treatment with dibucaine. Deciliation occurs first at the anterior end of the cell and then progresses posteriorly. Concomitantly, all mature mucocysts are induced to discharge by the drug. The exact point of scission of each cilium is found to be a very localized region, between two specialized membrane arrays: the ciliary necklace and the ciliary patches, situated at the base of the cilium. Isolated cilia retain the patches, while the necklaces remain with the deciliated bodies. The cell membrane seals over the stubs. The new ciliary membrane then grows out above the necklace without the patches, which do not generally appear for several hours. Membrane renewal is therefore asynchronous, with bulk growth preceding the formation of specialized intramembrane particle arrays. During regrowth, the cilia also first return at the anterior end of the cell. This suggests that underlying gradients, perhaps related to Ca²⁺, are significant in the deciliation process.     

Mice lacking the mitochondrial exonuclease MGME1 develop inflammatory kidney disease with glomerular dysfunction

Mitochondrial DNA (mtDNA) maintenance disorders are caused by mutations in ubiquitously expressed nuclear genes and lead to syndromes with variable disease severity and tissue-specific phenotypes. Loss of function mutations in the gene encoding the mitochondrial genome and maintenance exonuclease 1 (MGME1) result in deletions and depletion of mtDNA leading to adult-onset multisystem mitochondrial disease in humans. To better understand the in vivo function of MGME1 and the associated disease pathophysiology, we characterized a Mgme1 mouse knockout model by extensive phenotyping of ageing knockout animals. We show that loss of MGME1 leads to de novo formation of linear deleted mtDNA fragments that are constantly made and degraded. These findings contradict previous proposal that MGME1 is essential for degradation of linear mtDNA fragments and instead support a model where MGME1 has a critical role in completion of mtDNA replication. We report that Mgme1 knockout mice develop a dramatic phenotype as they age and display progressive weight loss, cataract and retinopathy. Surprisingly, aged animals also develop kidney inflammation, glomerular changes and severe chronic progressive nephropathy, consistent with nephrotic syndrome. These findings link the faulty mtDNA synthesis to severe inflammatory disease and thus show that defective mtDNA replication can trigger an immune response that causes age-associated progressive pathology in the kidney.     

Genome-Wide Association Studies in an Isolated Founder Population from the Pacific Island of Kosrae

It has been argued that the limited genetic diversity and reduced allelic heterogeneity observed in isolated founder populations facilitates discovery of loci contributing to both Mendelian and complex disease. A strong founder effect, severe isolation, and substantial inbreeding have dramatically reduced genetic diversity in natives from the island of Kosrae, Federated States of Micronesia, who exhibit a high prevalence of obesity and other metabolic disorders. We hypothesized that genetic drift and possibly natural selection on Kosrae might have increased the frequency of previously rare genetic variants with relatively large effects, making these alleles readily detectable in genome-wide association analysis. However, mapping in large, inbred cohorts introduces analytic challenges, as extensive relatedness between subjects violates the assumptions of independence upon which traditional association test statistics are based. We performed genome-wide association analysis for 15 quantitative traits in 2,906 members of the Kosrae population, using novel approaches to manage the extreme relatedness in the sample. As positive controls, we observe association to known loci for plasma cholesterol, triglycerides, and C-reactive protein and to a compelling candidate loci for thyroid stimulating hormone and fasting plasma glucose. We show that our study is well powered to detect common alleles explaining ≥5% phenotypic variance. However, no such large effects were observed with genome-wide significance, arguing that even in such a severely inbred population, common alleles typically have modest effects. Finally, we show that a majority of common variants discovered in Caucasians have indistinguishable effect sizes on Kosrae, despite the major differences in population genetics and environment.     

Quantification of the zygotic barrier between interbreeding taxa using gene flow data

Hybridization and introgression via interspecific gene flow are common processes in the plant kingdom. The effectiveness of these processes is governed by the strengths of multiple zygotic barriers. These barriers have often been quantified in artificial settings using laborious and time‐consuming hand‐pollination experiments, but their quantification is nonexistent in the landscape. In this study, we utilized gene flow data within a spatially explicit simulation to assess the strengths of zygotic barriers. Our model system consisted of Populus nigra and its hybrid, P. × canadensis, which interbreed under natural conditions. The study population was located in the floodplain of the Eder River in central Germany. Pollen‐mediated introgression rates from hybrid males into the seeds of individual female trees were used as the target pattern using an inverse modeling approach. Simulations that treated pollen from both taxa equally revealed a large discrepancy between the observed and modeled rates of introgression for both taxa. The discrepancy was reduced by introducing a zygotic barrier against the pollen from the hybrid males. The best model outcome values indicated comparably strong zygotic barriers acting against pollen‐mediated introgressive gene flow into the two parental taxa, P. nigra and P. × canadensis. The sensitivity of our model was tested by applying different dispersal functions. Four common probability density functions were used along with a pollen dispersal function that had previously been fitted to gene flow data from the same dataset. The best barrier value was almost independent of the dispersal functions used here. Moreover, it was within the range previously determined in hand‐pollination‐based investigations, validating our model. These data indicate that the inverse modeling approach is a powerful method for quantifying hidden processes, and we discuss its use as a valuable tool for generating new insights into plant mating systems that are relevant to evolutionary biology and risk analyses in conservation efforts.     

The bacterial paromomycin resistance gene, aphH, as a dominant selectable marker in Volvox carteri

The aminoglycoside antibiotic paromomycin that is highly toxic to the green alga Volvox carteri is efficiently inactivated by aminoglycoside 3′-phosphotransferase from Streptomyces rimosus. Therefore, we made constructs in which the bacterial aphH gene encoding this enzyme was combined with Volvox cis-regulatory elements in an attempt to develop a new dominant selectable marker – paromomycin resistance (Pm^R) – for use in Volvox nuclear transformation. The construct that provided the most efficient transformation was one in which aphH was placed between a chimeric promoter that was generated by fusing the Volvox hsp70 and rbcS3 promoters and the 3′ UTR of the Volvox rbcS3 gene. When this plasmid was used in combination with a high-impact biolistic device, the frequency of stable Pm^R transformants ranged about 15 per 10⁶ target cells. Due to rapid and sharp selection, Pm^R transformants were readily isolated after six days, which is half the time required for previously used markers. Co-transformation of an unselected marker ranged about 30%. The chimeric aphH gene was stably integrated into the Volvox genome, frequently as tandem multiple copies, and was expressed at a level that made selection of Pm^R transformants simple and unambiguous. This makes the engineered bacterial aphH gene an efficient dominant selection marker for the transformation and co-transformation of a broad range of V. carteri strains without the recurring need for using auxotrophic recipient strains.     

Effects of Alpine land‐use changes: Soil macrofauna community revisited

Although soil invertebrates play a decisive role in maintaining ecosystem functioning, little is known about their structural composition in Alpine soils and how their abundances are affected by the currently ongoing land‐use changes. In this study, we re‐assessed the soil macrofauna community structure of managed and abandoned Alpine pastureland, which has already been evaluated 14 years earlier. Our results confirm clear shifts in the community composition after abandonment, in that (1) Chilopoda and Diplopoda were recorded almost exclusively on the abandoned sites, (2) Coleoptera larvae and Diptera larvae were more abundant on the abandoned than on the managed sites, whereas (3) Lumbricidae dominated on the managed sites. By revisiting managed and abandoned sites, we infer community patterns caused by abandonment such as changes in the epigeic earthworm community structure, and we discuss seasonal and sampling effects. Our case study improves the still limited understanding of spatio‐temporal biodiversity patterns of Alpine soil communities.