0389003 - ÚMG 2013 RIV GB eng C - Conference Paper (international conference)
Yildirim, Sukriye - Castano, Enrique - Philimonenko, Vlada - Dzijak, Rastislav - Sobol, Margaryta - Venit, Tomáš - Hozák, Pavel
Nuclear PIP2 and myosin I: new important players in DNA transcription.
emc2012 manchester, european microscopy congress. Manchester: RMS, IFSM, 2012, s. 77-77. ISBN 978-0-9502463-7-6.
[15th European Microscopy Congress. Manchester (GB), 16.09.2012-21.09.2012]
R&D Projects: GA ČR GAP305/11/2232; GA MŠMT LC545; GA MŠMT LC06063; GA MPO FR-TI3/588; GA ČR GD204/09/H084
Institutional research plan: CEZ:AV0Z50520514
Institutional support: RVO:68378050
Keywords : Myosin * PIP2 * transcription
Subject RIV: EB - Genetics ; Molecular Biology

Nuclear myosin I (NM1) is a 120 kDa molecular motor described in the cell nucleus. NM1 was shown to be involved in chromatin remodeling, repositioning of transcriptionally activated regions and also in transcription with RNA pol I. Myosin 1C binds to negatively charged phospholipids, specifically to phosphatidylinositol-(4,5)bisphosphate (PIP2) and this binding tethers myosin I to plasma membrane. Based on this we asked if NM1 also has binding properties to PIP2. We made single-point mutations in the pleckstrin homology (PH) domain of NM1 and applied fluorescence recovery after photobleaching (FRAP) and fluorescence correlation spectroscopy (FCS). Mutant NM1 became faster in mobility compared to its wild type NM1. We then depleted  PIP2 in the nucleus by co-transfecting the cells with inositol 5-phosphatase. After PIP2 depletion in nuclei, NM1 became faster showing once more that PIP2 binding reduces NM1 mobility. Phospholipase C delta (PLCδ) is an enzyme which binds to PIP2 via its PH domain and cleaves PIP2 into inositol (1,4,5) triphosphate and DAG. We mutated the PIP2 binding domain of PLCδPH and co-transfected cells with NM1and wild type or mutant PLCδPH. FRAP showed that NM1 mobility increased when PIP2 was occupied by wild type PLC compared to mutant one. All these data indicated that NM1 binds to PIP2 in the cell nucleus, and this was further confirmed by electron microscopy. We then focused on the function of PIP2 in ribosomal gene transcription and showed that PIP2 binds to the transcription machinery. Removal of PIP2 from in vitro transcription assays caused the inhibiton of transcription for ribosomal genes and it was possible to recover the transcription efficiency by adding back PIP2 to the transcription reaction. The data suggest that nucleolar PIP2 might serve as a transcription factor for ribosomal genes, and together with nucleolar myosin I it might form the structural core of nucleoli. Involvement of other structural proteins will be discussed.
Permanent Link: http://hdl.handle.net/11104/0220057