title: Molecular basis of the STIL coiled coil oligomerization explains its requirement for de-novo formation of centrosomes in mammalian cells.
abstract: The STIL protein is essential for centriole replication and for the non-templated, de novo centriole biogenesis that is required for mammalian embryogenesis. Here we performed quantitative biophysical and structural analysis of the central short coiled coil domain (CCD) of STIL that is critical for its function. Using biophysical, biochemical and cell biology approaches, we identified the specific residues in the CCD that mediate the oligomerization, centrosomal localization and protein interactions of STIL. We characterized the structural properties of the coiled coil peptide using circular dichroism spectroscopy and size exclusion chromatography. We identified two regions in this domain, containing eight hydrophobic residues, which mediate the coiled coil oligomerization. Mutations in these residues destabilized the coiled coil thermodynamically but in most cases did not affect its secondary structure. Reconstituting mouse embryonic fibroblasts lacking endogenous Stil, we show that STIL oligomerization mediated by these residues is not only important for the centrosomal functions of STIL during the canonical duplication process but also for de-novo formation of centrosomes.
authors.affiliation: Sheba Cancer Research Center and the Edmond and Lily Safra Children Hospital, Sheba Medical Center, Tel-Hashomer 52621, Israel.|Department of molecular genetics and biochemistry, Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel.
authors.forename: Ahuvit
authors.initials: A
authors.lastname: David
authors.affiliation: Institute of Chemistry, the Hebrew University of Jerusalem, Safra Campus, Givat Ram, Jerusalem 91904, Israel.
authors.forename: Hadar
authors.initials: H
authors.lastname: Amartely
authors.affiliation: Sheba Cancer Research Center and the Edmond and Lily Safra Children Hospital, Sheba Medical Center, Tel-Hashomer 52621, Israel.|Department of molecular genetics and biochemistry, Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel.
authors.forename: Noa
authors.initials: N
authors.lastname: Rabinowicz
authors.affiliation: Institute of Chemistry, the Hebrew University of Jerusalem, Safra Campus, Givat Ram, Jerusalem 91904, Israel.
authors.forename: Mai
authors.initials: M
authors.lastname: Shamir
authors.affiliation: Institute of Chemistry, the Hebrew University of Jerusalem, Safra Campus, Givat Ram, Jerusalem 91904, Israel.
authors.forename: Assaf
authors.initials: A
authors.lastname: Friedler
authors.affiliation: Sheba Cancer Research Center and the Edmond and Lily Safra Children Hospital, Sheba Medical Center, Tel-Hashomer 52621, Israel.|Department of molecular genetics and biochemistry, Faculty of Medicine, Tel Aviv University, Tel Aviv, Israel.
authors.forename: Shai
authors.initials: S
authors.lastname: Izraeli
mesh_terms: D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000737:chemistry* / Q000378:metabolism*; D055592:Biophysical Phenomena; D002478:Cells, Cultured; D018385:Centrosome / Q000378:metabolism*; D002850:Chromatography, Gel; D002942:Circular Dichroism; D006801:Humans; D051379:Mice; D011487:Protein Conformation; D025941:Protein Interaction Mapping; D055503:Protein Multimerization; D021381:Protein Transport; D011518:Proto-Oncogene Proteins / Q000737:chemistry* / Q000378:metabolism*; D000073942:T-Cell Acute Lymphocytic Leukemia Protein 1
chemical_list: D051792:Basic Helix-Loop-Helix Proteins; D011518:Proto-Oncogene Proteins; D000073942:T-Cell Acute Lymphocytic Leukemia Protein 1; C489366:Tal1 protein, mouse