DNA methylation mutants in Physcomitrella patens elucidate individual roles of CG and non-CG methylation in genome regulation.

Cytosine (DNA) methylation in plants regulates the expression of genes and transposons. While methylation in plant genomes occurs at CG, CHG, and CHH sequence contexts, the comparative roles of the individual methylation contexts remain elusive. Here, we present Physcomitrella patens as the second plant system, besides Arabidopsis thaliana, with viable mutants with an essentially complete loss of methylation in the CG and non-CG contexts. In contrast to A. thaliana, P. patens has more robust CHH methylation, similar CG and CHG methylation levels, and minimal cross-talk between CG and non-CG methylation, making it possible to study context-specific effects independently. Our data found CHH methylation to act in redundancy with symmetric methylation in silencing transposons and to regulate the expression of CG/CHG-depleted transposons. Specific elimination of CG methylation did not dysregulate transposons or genes. In contrast, exclusive removal of non-CG methylation massively up-regulated transposons and genes. In addition, comparing two exclusively but equally CG- or CHG-methylated genomes, we show that CHG methylation acts as a greater transcriptional regulator than CG methylation. These results disentangle the transcriptional roles of CG and non-CG, as well as symmetric and asymmetric methylation in a plant genome, and point to the crucial role of non-CG methylation in genome regulation.

Diabetes mellitus is associated with increased elastin fiber loss in ligamentum flavum of patients with lumbar spinal canal stenosis: results of a pilot histological study.

PURPOSE: Lumbar spinal canal stenosis (LSCS) is associated with fibrosis, decreased elastin-to-collagen ratio, and hypertrophy of the ligamentum flavum (LF). Diabetes mellitus (DM) is known to cause metabolic disturbances within the extracellular matrix in multiple tissues. These alterations may play a major role in the severity of clinical symptoms of LSCS affecting diabetic patients. We aimed to examine the hypothesis that DM may contribute to the LF changes seen in patients with LSCS. METHODS: The study cohort included 29 patients: 23 with LSCS (10 with DM vs. 13 without DM) as well as six patients with lumbar disc herniation (LDH). Surgical LF specimens were retrieved for histological assessment. Morphologic quantification of confocal microscopy images using fast Fourier transform analysis allowed us to compare anisotropy and elastin fiber orientation between groups. RESULTS: There was a significant positive correlation between fasting plasma glucose values and degree of elastin degradation (r = 0.36, p = 0.043). The diabetic patients with LSCS showed a significantly greater loss of elastic fibers (2.3 ± 0.9 vs. 1.5 ± 0.55, p = 0.009), although fibrosis was shown to be similar (1.44 ± 0.7 vs. 1.43 ± 0.88, p = 0.98). There was no significant difference in the degree of calcification in the LSCS group between patients with and without diabetes (1.71 vs. 2.05%, p = 0.653). Fiber orientation was found to be less homogenous in the LSCS compared with the LDH group, although not significantly affected by DM. CONCLUSIONS: The present study points to a significant contribution of DM to the loss of elastin fibers that occurs in the LF of patients with LSCS.

Movement of protein and macromolecules between host plants and the parasitic weed Phelipanche aegyptiaca Pers.

Little is known about the translocation of proteins and other macromolecules from a host plant to the parasitic weed Phelipanche spp. Long-distance movement of proteins between host and parasite was explored using transgenic tomato plants expressing green fluorescent protein (GFP) in their companion cells. We further used fluorescent probes of differing molecular weights to trace vascular continuity between the host plant and the parasite. Accumulation of GFP was observed in the central vascular bundle of leaves and in the root phloem of transgenic tomato plants expressing GFP under the regulation of AtSUC2 promoter. When transgenic tomato plants expressing GFP were parasitized with P. aegyptiaca, extensive GFP was translocated from the host phloem to the parasite phloem and accumulated in both Phelipanche tubercles and shoots. No movement of GFP to the parasite was observed when tobacco plants expressing GFP targeted to the ER were parasitized with P. aegyptiaca. Experiments using fluorescent probes of differing molecular weights to trace vascular continuity between the host plant and the parasite demonstrated that Phelipanche absorbs dextrans up to 70 kDa in size from the host and that this movement can be bi-directional. In the present study, we prove for the first time delivery of proteins from host to the parasitic weed P. aegyptiaca via phloem connections, providing information for developing parasite resistance strategies.