Identification of new DNA targets of nuclear envelope proteins in C. elegans using the DamID technique

Resumen del póster presentado al 18th International C. elegans Meeting, celebrado en Los Angeles, California (US) del 22 al 26 de junio de 2011.The nuclear envelope (NE) has emerged as an important structure that serves numerous pivotal roles in the cell including compartmentalization, control of nuclear position and morphology, contribution to cell stability, chromatin organization and regulation of gene expression. The mechanism by which the NE controls gene expression is not well understood yet. Traditionally, the anchoring of chromatin to the nuclear periphery has been associated with silencing and heterochromatin formation. However, recent studies have shown that there is also actively transcribed chromatin at the NE, specially associated with the nuclear pore complexes. To unravel how the NE can regulate gene expression, we have developed tools to perform genome wide analysis using the DamID method. This method is based on the expression in vivo of chimeric proteins containing an adenine methyltransferase (Dam) from E. coli that methylates the DNA in the vicinity of native binding sites of a chromatin-interacting protein. Using the MosSCI technique, we have created Caenorhabditis elegans strains containing single copy insertions of Dam fused to NE and nuclear pore proteins such as emerin/EMR-1, lamin/LMN-1 and Nup98/NPP-10N. Employing a genetically amenable model system enables us to analyze nuclear architecture across several mutant backgrounds, and we are currently exploring methods to control expression of the Dam fusion proteins in a temporal manner and in specific tissues. We have confirmed the correct expression and localization of our fusion proteins and we are currently analyzing DNA binding sites using whole-genome tiling arrays and qPCR.Peer Reviewe