Dicty News Electronic Edition Volume 24, number 14 June 3, 2005 Please submit abstracts of your papers as soon as they have been accepted for publication by sending them to dicty@northwestern.edu or by using the form at http://dictybase.org/db/cgi-bin/dictyBase/abstract_submit. Back issues of Dicty-News, the Dicty Reference database and other useful information is available at dictyBase - http://dictybase.org. ============= Abstracts ============= Transcriptional switch of the dia1 and impA promoter during the growth/differentiation transition Shigenori Hirose, Taira Mayanagi, Catherine Pears, Aiko Amagai, William F. Loomis, and Yasuo Maeda Eukaryotic Cell, in press When growth stops due to the depletion of nutrients, Dictyostelium cells rapidly turn off vegetative genes and start to express developmental genes. One of the early developmental genes, dia1, is adjacent to a vegetative gene, impA, on chromosome 4. An intergenic region of 654 bp separates the coding regions of these divergently transcribed genes. Constructs carrying the intergenic region express a reporter gene (GFP) that replaced impA in growing cells and a reporter gene that replaced dia1 (DsRed) during development. Deletion of a 112 bp region proximal to the transcriptional start site of impA resulted in complete lack of expression of both reporter genes during growth or development. At the other end of the intergenic region there are two copies of a motif that is also found in the carA regulatory region. Removing one copy of this repeat reduced impA expression 2-fold. Removing the second copy had no further consequences. Removing the central portion of the intergenic region resulted in high levels of expression of dia1 in growing cells indicating that this region contains a sequence involved in repression during the vegetative stage. Gel-shift experiments showed that a nuclear protein present in growing cells recognizes the sequence GAAGTTCTAATTGATTGAAG found in this region. This DNA binding activity is lost within the first 4 hours of development. Different nuclear proteins were found to recognize the repeated sequence proximal to dia1. One of these became prevalent after 4 hours of development. Together these regulatory components at least partially account for this aspect of the growth to differentiation transition. Submitted by: William F. Loomis [wloomis@ucsd.edu] ----------------------------------------------------------------------------- DNG1, a Dictyostelium homologue of tumor suppressor ING1 regulates differentiation of Dictyostelium cells. Taira Mayanagi, Aiko Amagai and Yasuo Maeda Department of Developmental Biology and Neurosciences, Graduate School of Life Sciences, Tohoku University, Aoba, Sendai 980-8578, Japan Cell Mol Life Sci., in press dng1 is a Dictyostelium homologue of the mammalian tumor suppressor ING gene. DNG1 protein localizes in the nucleus, and has a highly conserved PHD finger domain found in chromatin remodeling proteins. Both dng1 disruption and overexpression impaired cell proliferation. In dng1-null cells, the progression of differentiation was delayed in a cell-density dependent manner, and many tiny aggregates were formed. Exogenously applied cAMP pulses reversed the inhibitory effect caused by dng1 disruption on the aggregation during early development, but formation of tiny aggregates was not restored. dng1-overexpressing cells acquired the ability to chemotaxis to cAMP earlier and exhibited enhanced differentiation. These phenotypes were found to be coupled with altered expressions of early genes such as cAMP receptor 1 (car1) and contact site A (csA). Furthermore, disordered histone modifications were demonstrated in dng1-null cells. These results suggest a regulatory role of dng1 in the transition of cells from growth to differentiation. Submitted by: Taira Mayanagi [taira@nbiochem.med.osaka-u.ac.jp] ----------------------------------------------------------------------------- A collection of amino acid replacement matrices derived from clusters of orthologs Rolf Olsen and William F. Loomis Division of Biology. UCSD J. Molecular Evolution, in press Sequence divergence among orthologous proteins was characterized with 34 amino acid replacement matrices, sequence context analysis and a phylogenetic tree. The model was trained on very large datasets of aligned protein sequences drawn from 15 organisms including protists, plants, Dictyostelium, fungi and animals. Comparative tests with models currently used in phylogeny, i.e. with JTT+Gą F and WAG+Gą F, made on a test dataset of 380 multiple alignments containing protein sequences from all 5 of the major taxonomic groups mentioned, indicate our model should be preferred over the JTT+Gą F and WAG+Gą F models on datasets similar to the test dataset. The strong performance of our model of orthologous protein sequence divergence can be attributed to its ability to better approximate amino acid equilibrium frequencies to compositions found in alignment columns. Submitted by: William F. Loomis [wloomis@ucsd.edu] ----------------------------------------------------------------------------- Structural requirements of Dictyostelium differentiation-inducing factors for their stalk-cell-inducing activity in Dictyostelium cells and anti-proliferative activity in K562 human leukemic cells Naomi Gokan, Haruhisa Kikuchi, Koji Nakamura, Yoshiteru Oshima, Kohei Hosaka, and Yuzuru Kubohara* *Gunma University IMCR, Japan. Biochem. Pharmacol., in press The differentiation-inducing factor-1 (DIF-1) is a lipophilic signal molecule (chlorinated alkylphenone) that induces stalk cell differentiation in the cellular slime mould Dictyostelium discoideum. It has also been shown that DIF-1 and its derivative (DIF-3) suppress cell growth in mammalian tumor cells. In the present study, in order to assess the chemical structure-effect relationship of DIF derivatives and to develop useful agents for the study of both Dictyostelium development and cancer biology, we synthesized 28 analogues of DIF-1 and DIF-3 and investigated their stalk-cell-inducing activity in Dictyostelium HM44 cells (mutant strain) and anti-proliferative activity in human leukemia K562 cells. HM44 cells are defective in endogenous DIF-1 production and should be suitable for the assay for stalk-cell-inducing activity of DIF analogues. DIF-1 and some of its derivatives at nanomolar levels were good stalk-cell inducers in HM44 cells, whereas DIF-3 and some DIF-3 derivatives at micromolar levels were potent anti-proliferative agents in K562 cells. We also tried to search for antagonistic molecules against DIF-1 and DIF-3 but failed to find such molecules from the analogues used here. The present findings would give us hints for identifying the target molecule(s) of DIFs and also for developing novel anti-cancer drugs. Submitted by: Yuzuru Kubohara [kubohara@showa.gunma-u.ac.jp] ============================================================================== [End Dicty News, volume 24, number 14]