Chromatin and siRNA pathways cooperate to maintain DNA methylation of small transposable elements in Arabidopsis

BACKGROUND: DNA methylation occurs at preferred sites in eukaryotes. In Arabidopsis, DNA cytosine methylation is maintained by three subfamilies of methyltransferases with distinct substrate specificities and different modes of action. Targeting of cytosine methylation at selected loci has been found to sometimes involve histone H3 methylation and small interfering (si)RNAs. However, the relationship between different cytosine methylation pathways and their preferred targets is not known. RESULTS: We used a microarray-based profiling method to explore the involvement of Arabidopsis CMT3 and DRM DNA methyltransferases, a histone H3 lysine-9 methyltransferase (KYP) and an Argonaute-related siRNA silencing component (AGO4) in methylating target loci. We found that KYP targets are also CMT3 targets, suggesting that histone methylation maintains CNG methylation genome-wide. CMT3 and KYP targets show similar proximal distributions that correspond to the overall distribution of transposable elements of all types, whereas DRM targets are distributed more distally along the chromosome. We find an inverse relationship between element size and loss of methylation in ago4 and drm mutants. CONCLUSION: We conclude that the targets of both DNA methylation and histone H3K9 methylation pathways are transposable elements genome-wide, irrespective of element type and position. Our findings also suggest that RNA-directed DNA methylation is required to silence isolated elements that may be too small to be maintained in a silent state by a chromatin-based mechanism alone. Thus, parallel pathways would be needed to maintain silencing of transposable elements

Polyploid genome of Camelina sativa revealed by isolation of fatty acid synthesis genes

<p>Abstract</p> <p>Background</p> <p><it>Camelina sativa</it>, an oilseed crop in the Brassicaceae family, has inspired renewed interest due to its potential for biofuels applications. Little is understood of the nature of the <it>C. sativa </it>genome, however. A study was undertaken to characterize two genes in the fatty acid biosynthesis pathway, <it>fatty acid desaturase (FAD) 2 </it>and <it>fatty acid elongase (FAE) 1</it>, which revealed unexpected complexity in the <it>C. sativa </it>genome.</p> <p>Results</p> <p>In <it>C. sativa</it>, Southern analysis indicates the presence of three copies of both <it>FAD2 </it>and <it>FAE1 </it>as well as <it>LFY</it>, a known single copy gene in other species. All three copies of both <it>CsFAD2 </it>and <it>CsFAE1 </it>are expressed in developing seeds, and sequence alignments show that previously described conserved sites are present, suggesting that all three copies of both genes could be functional. The regions downstream of <it>CsFAD2 </it>and upstream of <it>CsFAE1 </it>demonstrate co-linearity with the Arabidopsis genome. In addition, three expressed haplotypes were observed for six predicted single-copy genes in 454 sequencing analysis and results from flow cytometry indicate that the DNA content of <it>C. sativa </it>is approximately three-fold that of diploid <it>Camelina </it>relatives. Phylogenetic analyses further support a history of duplication and indicate that <it>C. sativa </it>and <it>C. microcarpa </it>might share a parental genome.</p> <p>Conclusions</p> <p>There is compelling evidence for triplication of the <it>C. sativa </it>genome, including a larger chromosome number and three-fold larger measured genome size than other <it>Camelina </it>relatives, three isolated copies of <it>FAD2</it>, <it>FAE1</it>, and the <it>KCS17-FAE1 </it>intergenic region, and three expressed haplotypes observed for six predicted single-copy genes. Based on these results, we propose that <it>C. sativa </it>be considered an allohexaploid. The characterization of fatty acid synthesis pathway genes will allow for the future manipulation of oil composition of this emerging biofuel crop; however, targeted manipulations of oil composition and general development of <it>C. sativa </it>should consider and, when possible take advantage of, the implications of polyploidy.</p

Prospective Analyses of Circulating B Cell Subsets in ABO-Compatible and ABO-Incompatible Kidney Transplant Recipients

Immunosuppressive strategies applied in renal transplantation traditionally focus on T cell inhibition. B cells were mainly examined in the context of antibody-mediated rejection, whereas the impact of antibody-independent B cell functions has only recently entered the field of transplantation. Similar to T cells, distinct B cell subsets can enhance or inhibit immune responses. In this study, we prospectively analyzed the evolution of B cell subsets in the peripheral blood of ABO-compatible (n = 27) and ABO-incompatible (n = 10) renal transplant recipients. Activated B cells were transiently decreased and plasmablasts were permanently decreased in patients without signs of rejection throughout the first year. In patients with histologically confirmed renal allograft rejection, activated B cells and plasmablasts were significantly elevated on day 365. Rituximab treatment in ABO-incompatible patients resulted in long-lasting B cell depletion and in a naive phenotype of repopulating B cells 1 year following transplantation. Acute allograft rejection was correlated with an increase of activated B cells and plasmablasts and with a significant reduction of regulatory B cell subsets. Our study demonstrates the remarkable effects of standard immunosuppression on circulating B cell subsets. Furthermore, the B cell compartment was significantly altered in rejecting patients. A specific targeting of deleterious B cell subsets could be of clinical benefit in renal transplantation