The histone methyl transferase PR-Set7 mediates histone H4 Lys 20 methylation, a mark of constitutive and facultative heterochromatin. We isolated a null mutation in Drosophila PR-Set7 that suppresses position effect variegation, indicating that PR-Set7 indeed functions in silencing general gene expression. In PR-Set7 larval leg and eye discs, the number of cells is lower than normal, and the DNA content in these cells is significantly increased. These data show that PR-Set7-dependent methylation is essential for the process of mitosis. The methylation mark is highly stable and is maintained even in the absence of PR-Set7 protein.Supplemental material is available at http://www.genesdev.org.

Di-and trimethylation of histone H4 lysine20 (H4K20) are thought to play an important role in controlling gene expression in vertebrates and in Drosophila. By inducing a null mutation in Drosophila Suv4-20, we show that it encodes the histone H4 lysine20 di-and trimethyltransferase. In Suv4-20 mutants, the H4K20 di-and trimethyl marks are strongly reduced or absent, and the monomethyl mark is significantly increased. We find that even with this biochemical function, Suv4-20 is not required for survival and does not control position-effect variegation (PEV).

The epidermis serves as a protective barrier in animals. After epidermal injury, barrier repair requires activation of many wound response genes in epidermal cells surrounding wound sites. Two such genes in Drosophila encode the enzymes dopa decarboxylase (Ddc) and tyrosine hydroxylase (ple). In this paper we explore the involvement of the Toll/NF-κB pathway in the localized activation of wound repair genes around epidermal breaks. Robust activation of wound-induced transcription from ple and Ddc requires Toll pathway components ranging from the extracellular ligand Spätzle to the Dif transcription factor. Epistasis experiments indicate a requirement for Spätzle ligand downstream of hydrogen peroxide and protease function, both of which are known activators of wound-induced transcription. The localized activation of Toll a few cell diameters from wound edges is reminiscent of local activation of Toll in early embryonic ventral hypoderm, consistent with the hypothesis that the dorsal–ventral patterning function of Toll arose from the evolutionary cooption of a morphogen-responsive function in wound repair. Furthermore, the combinatorial activity of Toll and other signaling pathways in activating epidermal barrier repair genes can help explain why developmental activation of the Toll, ERK, or JNK pathways alone fail to activate wound repair loci.

Methylation of specific amino acids in histone tails is responsible for packaging DNA into condensed, repressed chromatin, and into open chromatin that is accessible to the transcription machinery. Monomethylation and trimethylation of histone H4-lysine 20 (H4-K20) control the formation of repressed chromatin. Using antibodies that specifically recognize the three methyl marks of histone H4-K20, we characterized their regulation during the cell cycle and throughout development. We find free mono- and trimethylated histone H4-K20 in unfertilized Drosophila eggs and in S2 tissue culture cells. Soluble mono-. di-, and trimethylated H4-K20 are also found in HeLa cells. These soluble modified histones may represent a pool of free histones that can rapidly be incorporated into chromatin. The three methyl marks are each regulated differentially during development and their detection on western blots does not overlap with their detection on chromosomes. Monomethylated H4-K20 is detected on condensed chromosomes throughout development, while di- and trimethylated H4-K20 are detected on metaphase chromosomes at specific stages. Our results suggest that the detection of methylated H4-K20 on chromosomes may reveal chromatin packaging rather than the distribution of the methyl marks.

No abstract

No abstract

No abstract

In previous work we demonstrated that various types of cultured cells with a limited life span could not reactivate DNA synthesis in the nuclei of mouse peritoneal macrophages in heterokaryons. We now investigate the role of telomerase in the process of the macrophage nucleus reactivation in heterokaryons with immortal telomerase-positive 3T3 Swiss mouse fibroblasts and human fibroblasts with introduced hTERT gene. We report that introduction of the hTERT gene into human diploid fibroblasts results in emergence of telomerase activity in these cells and the ability to induce the reactivation of DNA synthesis in the macrophage nuclei in heterokaryons. Inhibition of telomerase activity in heterokaryons by reverse transcriptase inhibitors (azidothymidine and guanosine polyphosphonate analogues) and by a 2'-O-methyl-RNA oligonucleotide anti-sense to the template region of telomerase RNA, block reactivation of DNA synthesis in macrophage nuclei without inhibiting DNA synthesis in the nuclei of fibroblasts. Our results suggest alterations (shortening or damage) in the macrophage telomere structure. As far as we know, heterokaryons with macrophages are the first cellular model for rapid investigation of the effects of telomerase inhibitors.