Rekonstruktion der Evolution von DNA-Bindespezifitäten von Transkriptionsfaktoren in rezenten Arten und ihren gemeinsamen Vorfahren
Genetik und Genomik der Pflanzen
Zusammenfassung der Projektergebnisse
Transcription factors (TFs) are the main regulators of gene transcription. To understand how they recognize their DNA targets, and how they gain or lose target sites during evolution, is fundamental for understanding how gene networks are controlled and diverge in function over evolutionary time. With this aim is important to develop tools to characterize the DNA binding specificity of any TF. In the past, we have stablished a methodology (SELEX-seq) to characterize relative DNA specificities of TFs. However, we realized that this method suffer of some bias as the DNA binding specificity calculated using different round of enrichment is not always reproducible. Therefore, in this project we stablished a new method where we do in parallel several round of enrichment using different DNA concentration for the initial pool of DNA oligos. In this way, we can precisely model the Kd of the TF to a particular DNA sequence observing the enrichment of that particular sequence depending of the TF/DNA ratio. We apply these methodology to characterize the AP1/FUL subfamily of TFs in Arabidopsis and other species. In particular, we could propose a model of how FUL TFs achieve different biological functions in different plant tissues. In this way, using our SELEX-seq approach, we identified that FUL TF change in DNA specificity depending of the presence of other TFs in their protein complex. This prompt us to ask the question: Can the different target genes of FUL TF complexes in different plant tissues be explained by a change in the DNA binding specificity of the FUL complex? By combining FUL ChIP-seq in flower meristem and pistils, and SELEX-seq experiments, we could show that tissue-specific FUL DNA binding observed by ChIP-seq could be explained by a different composition of FUL TF complex. To correctly predict the flower meristems binding sites, it is needed that FUL form complexes with SOC1 in this tissue, and to predict the DNA binding sites in pistils it is needed that FUL form complexes with SEP1, SEP3 and AG in this tissue. This was validated by isolating and characterizing the proteins associated with FUL in vivo (LC-MS/MS of FUL IPs in flower meristems and psitils). This challenges the traditional idea that the dynamics of chromatin is the main driver of target tissue-specificity of FUL TF complexes, and uncovers the importance of the dynamics of FUL TF complex composition in reshaping the regulatory function of FUL during flower development.
Projektbezogene Publikationen (Auswahl)
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“Combining ChIP-seq and SELEX-seq experiments: a tool to identify the target regions of particular transcription factor complexes”. Presentation at Workshop on Molecular Mechanisms controlling flower development (Cote d’Azur, FRANCE)
Muino J. M., Smaczniak C., Angenent G. C. & Kaufmann K.
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“Reconstructing the evolution of plant transcription factor DNA binding specificity across living species and their common ancestors” Poster at International Plant Systems Biology (Virtual)
Peilin, C., Smaczniak, C., , Kaufmann, K. & Muino J. M.
