Research Interests
The long-term goal of our research projects is to understand the molecular basis of gene silencing in eukaryotes. Gene activity is influenced by the proteins that package the DNA, by enzymes that modify these proteins or the DNA itself, and by RNA-mediated mechanisms (e.g., RNA interference). All these pathways play important roles in the control of gene expression during development (preventing cancer and other diseases) as well as in host defense responses against viruses and transposable elements. Thus, elucidation of these mechanisms will have an impact not only in basic biology but also in medicine and agriculture (http://www.pbs.org/wgbh/nova/sciencenow/3210/02.html). We are currently using the unicellular green alga Chlamydomonas reinhardtii and the higher plant Arabidopsis thaliana as model systems to identify and characterize molecular components of the gene silencing machineries. We are also interested in the applications of gene silencing for biotechnological and therapeutic purposes.
RNA Interference (RNAi)
Double-stranded RNA (dsRNA) is the trigger at the heart of RNAi and it can induce, in different organisms, a variety of outcomes such as the degradation of homologous RNAs, heterochromatin formation, DNA methylation, or even DNA elimination. The RNAi machinery has also been implicated in the processing and function of microRNAs, endogenous small RNAs that regulate gene expression by translational repression or mRNA cleavage. Despite rapid progress in understanding key steps of these pathways, many factors required for RNAi and related mechanisms have not been characterized as yet. We have isolated, by a variety of genetic screens, mutants in two classes of genes involved in RNAi. One group corresponds to factors that appear to be direct components of the RNAi machinery: MUT68p, a member of the DNA polymerase b-like nucleotidyltransferase superfamily; MUT70p, a putative RNA-binding protein; and MUT91p, a novel protein with a C2H2 zinc finger domain. Another group of genes, typified by MUT6 (encoding a DEAH-box RNA helicase), seems to regulate the pre-mRNA processing and, thus, the mature mRNA levels of RNAi components. We currently are characterizing the molecular role(s) of all these proteins as well as continuing with the isolation of RNAi-defective mutants.
Euchromatic Transcriptional Silencing
Post-translational histone modifications play an important role in determining chromatin states associated with gene expression or repression. These covalent modifications can be propagated mitotically, resulting in heritable epigenetic states that provide reversible cellular memory. However, relatively little is known about the role of post-translational histone modifications on the regulation of euchromatic genes in photosynthetic eukaryotes. Our lab is addressing the molecular mechanisms responsible for, and the role of, methylation and phosphorylation of core histones in the transcriptional silencing of euchromatin. We are also studying the function of these modifications in plant developmental programs and in the responses to abiotic stresses.
Representative Publications
Casas-Mollano J. A., Jeong B.-r., Xu J., Moriyama H. and Cerutti H. 2008. The MUT9p kinase phosphorylates histone H3 threonine 3 and is necessary for heritable epigenetic silencing in Chlamydomonas. Proc. Natl. Acad. Sci. USA 105: 6486-6491.
Casas-Mollano J. A., Rohr J., Kim E.-J., Balassa E., van Dijk K. and Cerutti H. 2008. Diversification of the core RNAi machinery in Chlamydomonas reinhardtii and the role of DCL1 in transposon silencing. Genetics 179: 69-81.
Merchant S.S., Prochnik S.E., Vallon O., Harris E.H., Karpowicz S.J. et al. 2007. The Chlamydomonas genome reveals the evolution of key animal and plant functions. Science 318: 245-250.
Casas-Mollano J. A., van Dijk K., Eisenhart J. and Cerutti H. 2007. SET3p monomethylates histone H3 on lysine 9 and is required for the silencing of tandemly repeated transgenes in Chlamydomonas. Nucleic Acids Research 35: 939-950.
Ibrahim F., Rohr J., Jeong W.-J., Hesson J. and Cerutti H. 2006. Untemplated oligoadenylation promotes degradation of RISC-cleaved transcripts. Science, 314: 1893.
Cerutti H. and Casas-Mollano A. 2006. On the Origin and Functions of RNA-Mediated Silencing: From Protists to Man. Curr Genet 50: 81-99.
van Dijk K.V., Marley K.E., Jeong B.-r., Xu J., Hesson J., Cerny R.L., Waterborg J.H. and Cerutti H. 2005. Monomethyl histone H3 lysine 4 as an epigenetic mark for silenced euchromatin in Chlamydomonas. Plant Cell, 17: 2439-2453.
Sarkar N., Lemaire S., Wu-Scharf D., Issakidis-Bourguet E., and Cerutti H. 2005. Functional Specialization of Chlamydomonas Cytosolic Thioredoxin h1 in the Response to Alkylation-Induced DNA Damage. Eukaryotic Cell 4: 262-273.
Rohr J., Sarkar N., Balenger S., Jeong B.-r. and Cerutti H. 2004. Tandem inverted repeat system for selection of effective transgenic RNAi strains in Chlamydomonas. Plant J. 40: 611-621.
van Dijk K.V. and Cerutti H. 2004. RNA-mediated silencing. In Encyclopedia of Plant and Crop Science; Robert M. Goodman, ed.; Marcel Dekker: New York; pp. 1242-1244.
Cerutti H. 2003. RNA interference: traveling in the cell and gaining functions? Trends Genet. 19, 39-46.
Zhang C., Wu-Scharf D., Jeong B.-r. and Cerutti H. 2002. A WD40-repeat containing protein, similar to a fungal co-repressor, is required for transcriptional gene silencing in Chlamydomonas. Plant J. 31: 25-36.
Jeong B.-r., Wu-Scharf D., Zhang C. and Cerutti H. 2002. Suppressors of transcriptional transgenic silencing in Chlamydomonas are sensitive to DNA-damaging agents and reactivate transposable elements. Proc. Natl. Acad. Sci. USA 99: 1076-1081.
Wu-Scharf D., Jeong B.-r., Zhang C. and Cerutti H. 2000. Transgene and transposon silencing in Chlamydomonas reinhardtii by a DEAH-box RNA helicase. Science 290: 1159-1162.
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