The histone H3K4 demethylase JARID1A directly interacts with haematopoietic transcription factor GATA1 in erythroid cells through its second PHD domain

Chromatin remodelling and transcription factors play important roles in lineage commitment and development through control of gene expression. Activation of selected lineage-specific genes and repression of alternative lineage-affiliated genes results in tightly regulated cell differentiation transcriptional programmes. However, the complex functional and physical interplay between transcription factors and chromatin modifying enzymes remains elusive. Recent evidence has implicated histone demethylases in normal haematopoietic differentiation as well as in malignant haematopoiesis. Here we report an interaction between H3K4 demethylase JARID1A and the haematopoietic-specific master transcription proteins SCL and GATA1 in red blood cells. Specifically, we observe a direct physical contact between GATA1 and the second PHD domain of JARID1A. This interaction has potential implications for normal and malignant haematopoiesis

Structure of the DNA-bound T-box domain of human TBX1, a transcription factor associated with the DiGeorge syndrome

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Structure and mechanism of bactericidal mammalian perforin-2, an ancient agent of innate immunity

Perforin-2 (MPEG1) is thought to enable the killing of invading microbes engulfed by macrophages and other phagocytes, forming pores in their membranes. Loss of perforin-2 renders individual phagocytes and whole organisms significantly more susceptible to bacterial pathogens. Here, we reveal the mechanism of perforin-2 activation and activity using atomic structures of pre-pore and pore assemblies, high-speed atomic force microscopy, and functional assays. Perforin-2 forms a pre-pore assembly in which its pore-forming domain points in the opposite direction to its membrane-targeting domain. Acidification then triggers pore formation, via a 180° conformational change. This novel and unexpected mechanism prevents premature bactericidal attack and may have played a key role in the evolution of all perforin family proteins

Characterization of the interactions between the H3K4 demethylase JARID1A and the SCL-nucleated oncogenic DNA binding complex

Transcriptional regulation is of key importance to cellular processes such as development and differentiation, and it is dependent on the accessibility of DNA in the chromatin. Methylation and demethylation of histones alter properties of nucleosomes, thereby changing the accessibility of DNA. For many years, histone tail methylation was considered to be irreversible but in the last decade two families of histone demethylases were discovered, lysine specific demethylase 1 (LSD1) and Jumonji C (JMJC) demethylases. The work presented in this thesis focuses on the chromatin remodeling protein JARID1A, belonging to the JARID1 family of JMJC demethylases that demethylates lysine 4 (K4) on the tail of histone H3. Double and triple methylation of K4 on histone H3 are marks traditionally linked to transcriptionally active regions of DNA. Recently, a translocation fusing the third PHD domain (PHD3) of JARID1A to NUP98 (a common leukaemia fusion partner with transactivation activity) was identified in acute myeloid leukaemia patients (AML). Also, PHD3 was previously shown to mediate interactions between JARID1A and the second LIM domain of LMO2. LMO2 operates within the multiprotein DNA binding complex named "SCL complex" (including proteins SCL/TAL1, LDB1, E47, GATA1) that regulates gene expression at different stages of erythropoiesis. This thesis presents findings of an investigation into relations between JARID1A and the oncogenic multiprotein complex SCL-E47-LMO2-LDB1-GATA1. The study of the endogenous proteins from erythroid cells using techniques such as Co-immunoprecipitation, GST pull-down and Size-exclusion chromatography showed that JARID1A interacts with SCL and GATA1. These interactions were further characterized biochemically using analytical ultracentrifugation which revealed that the second PHD domain of JARID1A is involved in direct interaction with GATA1. Hence, JARID1A could play a role in regulation of erythropoiesis by modulating activities of transcription factors SCL and GATA1.</p

Flavivirus maturation leads to the formation of an occupied lipid pocket in the surface glycoproteins

Here, the authors provide cryo-EM structures of mature and immature Spondweni virus, defining the furin recognition site at high resolution, and identifying a lipid that binds E upon capsid maturation and is also present in Zika and Dengue virions

Structure of the leukemia oncogene LMO2: implications for the assembly of a hematopoietic transcription factor complex

The LIM only protein 2 (LMO2) is a key regulator of hematopoietic stem cell development whose ectopic expression in T cells leads to the onset of acute lymphoblastic leukemia. Through its LIM domains, LMO2 is thought to function as the scaffold for a DNA-binding transcription regulator complex, including the basic helix-loop-helix proteins SCL/TAL1 and E47, the zinc finger protein GATA-1, and LIM-domain interacting protein LDB1. To understand the role of LMO2 in the formation of this complex and ultimately to dissect its function in normal and aberrant hematopoiesis, we solved the crystal structure of LMO2 in complex with the LID domain of LDB1 at 2.4 Å resolution. We observe a largely unstructured LMO2 kept in register by the LID binding both LIM domains. Comparison of independently determined crystal structures of LMO2 reveals large movements around a conserved hinge between the LIM domains. We demonstrate that such conformational flexibility is necessary for binding of LMO2 to its partner protein SCL/TAL1 in vitro and for the function of this complex in vivo. These results, together with molecular docking and analysis of evolutionarily conserved residues, yield the first structural model of the DNA-binding complex containing LMO2, LDB1, SCL/TAL1, and GATA-1