Distinct translatome changes in specific neural populations precede electroencephalographic changes in prion-infected mice.

Selective vulnerability is an enigmatic feature of neurodegenerative diseases (NDs), whereby a widely expressed protein causes lesions in specific cell types and brain regions. Using the RiboTag method in mice, translational responses of five neural subtypes to acquired prion disease (PrD) were measured. Pre-onset and disease onset timepoints were chosen based on longitudinal electroencephalography (EEG) that revealed a gradual increase in theta power between 10- and 18-weeks after prion injection, resembling a clinical feature of human PrD. At disease onset, marked by significantly increased theta power and histopathological lesions, mice had pronounced translatome changes in all five cell types despite appearing normal. Remarkably, at a pre-onset stage, prior to EEG and neuropathological changes, we found that 1) translatomes of astrocytes indicated reduced synthesis of ribosomal and mitochondrial components, 2) glutamatergic neurons showed increased expression of cytoskeletal genes, and 3) GABAergic neurons revealed reduced expression of circadian rhythm genes. These data demonstrate that early translatome responses to neurodegeneration emerge prior to conventional markers of disease and are cell type-specific. Therapeutic strategies may need to target multiple pathways in specific populations of cells, early in disease.

Translation of the prion protein mRNA is robust in astrocytes but does not amplify during reactive astrocytosis in the mouse brain.

Prion diseases induce neurodegeneration in specific brain areas for undetermined reasons. A thorough understanding of the localization of the disease-causing molecule, the prion protein (PrP), could inform on this issue but previous studies have generated conflicting conclusions. One of the more intriguing disagreements is whether PrP is synthesized by astrocytes. We developed a knock-in reporter mouse line in which the coding sequence of the PrP expressing gene (Prnp), was replaced with that for green fluorescent protein (GFP). Native GFP fluorescence intensity varied between and within brain regions. GFP was present in astrocytes but did not increase during reactive gliosis induced by scrapie prion infection. Therefore, reactive gliosis associated with prion diseases does not cause an acceleration of local PrP production. In addition to aiding in Prnp gene activity studies, this reporter mouse line will likely prove useful for analysis of chimeric animals produced by stem cell and tissue transplantation experiments.

Tracing a key player in the regulation of plant architecture: the columnar growth habit of apple trees (Malus × domestica).

Plant architecture is regulated by a complex interplay of some key players (often transcription factors), phytohormones and other signaling molecules such as microRNAs. The columnar growth habit of apple trees is a unique form of plant architecture characterized by thick and upright stems showing a compaction of internodes and carrying short fruit spurs instead of lateral branches. The molecular basis for columnar growth is a single dominant allele of the gene Columnar, whose identity, function and gene product are unknown. As a result of marker analyses, this gene has recently been fine-mapped to chromosome 10 at 18.51-19.09 Mb [according to the annotation of the apple genome by Velasco (2010)], a region containing a cluster of quantitative trait loci associated with plant architecture, but no homologs to the well-known key regulators of plant architecture. Columnar apple trees have a higher auxin/cytokinin ratio and lower levels of gibberellins and abscisic acid than normal apple trees. Transcriptome analyses corroborate these results and additionally show differences in cell membrane and cell wall function. It can be expected that within the next year or two, an integration of these different research methodologies will reveal the identity of the Columnar gene. Besides enabling breeders to efficiently create new apple (and maybe related pear, peach, cherry, etc.) cultivars which combine desirable characteristics of commercial cultivars with the advantageous columnar growth habit using gene technology, this will also provide new insights into an elevated level of plant growth regulation.

Evaluation of the hormonal state of columnar apple trees (Malus x domestica) based on high throughput gene expression studies.

The columnar phenotype of apple trees (Malus x domestica) is characterized by a compact growth habit with fruit spurs instead of lateral branches. These properties provide significant economic advantages by enabling high density plantings. The columnar growth results from the presence of a dominant allele of the gene Columnar (Co) located on chromosome 10 which can appear in a heterozygous (Co/co) or homozygous (Co/Co) state. Although two deep sequencing approaches could shed some light on the transcriptome of columnar shoot apical meristems (SAMs), the molecular mechanisms of columnar growth are not yet elaborated. Since the influence of phytohormones is believed to have a pivotal role in the establishment of the phenotype, we performed RNA-Seq experiments to study genes associated with hormone homeostasis and clearly affected by the presence of Co. Our results provide a molecular explanation for earlier findings on the hormonal state of columnar apple trees. Additionally, they allow hypotheses on how the columnar phenotype might develop. Furthermore, we show a statistically approved enrichment of differentially regulated genes on chromosome 10 in the course of validating RNA-Seq results using additional gene expression studies.

The transcriptomes of columnar and standard type apple trees (Malus x domestica) - a comparative study.

Columnar apple trees (Malus x domestica) provide several economic advantages due to their specific growth habit. The columnar phenotype is the result of the dominant allele of the gene Co and is characterized by thick stems with short internodes and reduced lateral branching. Co is located on chromosome 10 and often appears in a heterozygous state (Co/co). The molecular explanation of columnar growth is not well established. Therefore, we studied the transcriptomes of columnar and standard type apple trees using 454 and Illumina next generation sequencing (NGS) technologies. We analyzed the transcriptomes of shoot apical meristems (SAMs) because we expect that these organs are involved in forming the columnar growth phenotype. The results of the comparative transcriptome analysis show significant differences in expression levels of hundreds of genes. Many of the differentially expressed genes are associated with membrane and cell wall growth or modification and can be brought in line with the columnar phenotype. Additionally, earlier findings on the hormonal state of shoots of columnar apples could be affirmed. Our study resulted in a large number of genes differentially expressed in columnar vs. standard type apple tree SAMs. Although we have not unraveled the nature of the Co gene, we could show that the modified expression of these genes, most likely due to the presence of Co, can determine the columnar phenotype. Furthermore, the usefulness of NGS for the analysis of the molecular basis of complex phenotypes is discussed.