<<The>> elongator complex in plant: a study of its molecular networks

Abstract

The Elongator complex is a histone acetyltransferase complex associated with RNAPII to facilitate transcript elongation. It’s composed of six proteins (ELP1-6). ELP1-3 form the Elongator core subcomplex, while ELP4-6 form the accessory subcomplex. Elongator complex, firstly identified in yeast, was later isolated from animals and plants and all its six subunits are evolutionarily conserved. The Elongator activity is conferred by ELP3 that targets specifically histons H3 (lysine-14) and H4 (lysine-12) by acetylating histone in order to facilitate the RNAPII progresses through the nucleosome. In yeast, mutations in Elongator subunits induce delay in growth due to a slowly adaptation to changing environmental conditions. In human, mutations in Elongator components affect neuronal development and this leads to neuronal disease. Whereas, in plant Elongator stimulates plant growth acting a positive regulator of cell proliferation. At the phenotypic level, Elongator mutants, called elongata, are known for narrow leaves and short root. In the present work, by using the model plant Arabidopsis thaliana, we investigated some aspects of molecular networks underlying Elongator activity and its interaction with environmental factors, mainly focusing on light conditions. Based on previous unpublished data obtained through TAP analysis, in the first period of PhD project we focused the attention on the functional study of Sec31 gene encoding a protein involved in cell secretory pathway, identified as a putative direct interactor of Elongator complex. To add information on this interaction we analyzed phenotypic and developmental characteristics of sec31 mutants to compare with elo3-6 mutant. The histological expression pattern of Sec31 and ELO3 transcripts in wild type seedlings was also investigated, through multiprobe in situ hybridization, to compare organ/tissue specific expression domains. The obtained results showed that expression pattern of the two genes is quite similar while sec31 mutants do not resemble elo3-6 phenotypes. Moreover further TAP experiments and in silico analysis of protein/protein interaction did not confirm previous data, thus excluding a direct interaction between ELO3 and Sec31. However, expression analysis in sec31 mutants of some Elongator-related genes, performed by qRT-PCR, showed that Sec31 and ELO3 share common downstream target genes and both seem play a role in auxin pathway. Future trascriptomic analyses on auxin mutants on one side, and the identification of possible interactors/players of both genes on the other side, could be useful to deepen if the molecular circuits, by which Elongator complex and the secretory machinery act on auxin pathway, show some cross-talk or they work in an independent manner. A further aspect of Elongantor molecular network that we investigated deals with role of Elongator in the skoto/photomorphogenesis pathways. In particular we investigated the elo3-6 mutant in darkness and under light condition (red, far-red and blue light) through microarray and RNA-seq approaches. Gene ontology categories over representative in elo3-6 seedlins, identified by BINGO analysis, allowed us to discover the putative targets of Elongator both in darkness and in light, and to understand the position of whole Elongator complex along either pathways. The results suggested that Elongator complex takes part in the skotomorphogenesis and photomorphogenesis and is dependent on photoreceptors PHYA and PHYB. Microarray, RNA-seq, qRT-PCR and ChIP- qPCR analyses displayed that Elongator regulates transcription of some genes both in light and in darkness. In the specific, results displayed that Elongator complex participates in the skoto/photomorphogenic pathways by binding target genes such as HYH and LHY in light and darkness condition, respectively. Whereas it can regulate the activity of other putative targets such as Pifs gene (PIF4) in darkness and HY5 under light condition.