Person: Bonni, Azad
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Bonni
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Azad
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Bonni, Azad
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Publication Promoter Decommissioning by the NuRD Chromatin Remodeling Complex Triggers Synaptic Connectivity in the Mammalian Brain(Elsevier BV, 2014) Yamada, Tomoko; Yang, Yue Bo; Hemberg, Martin; Yoshida, Toshimi; Cho, Ha Young; Murphy, J. Patrick; Fioravante, Diasynou; Regehr, Wade; Gygi, Steven; Georgopoulos, Katia; Bonni, AzadPrecise control of gene expression plays fundamental roles in brain development, but the roles of chromatin regulators in neuronal connectivity have remained poorly understood. We report that depletion of the NuRD complex by in vivo RNAi and conditional knockout of the core NuRD subunit Chd4 profoundly impairs the establishment of granule neuron parallel fiber/Purkinje cell synapses in the rodent cerebellar cortex in vivo. By interfacing genome-wide sequencing of transcripts and ChIP-seq analyses, we uncover a network of repressed genes and distinct histone modifications at target gene promoters that are developmentally regulated by the NuRD complex in the cerebellum in vivo. Finally, in a targeted in vivo RNAi screen of NuRD target genes, we identify a program of NuRD-repressed genes that operate as critical regulators of presynaptic differentiation in the cerebellar cortex. Our findings define NuRD-dependent promoter decommissioning as a developmentally regulated programming mechanism that drives synaptic connectivity in the mammalian brain.Publication FLEXIQinase, a mass spectrometry-based assay, to unveil multi-kinase mechanisms(2013) Singh, Sasha; Winter, Dominic; Bilimoria, Parizad; Bonni, Azad; Steen, Hanno; Steen, Judith A.We introduce a mass spectrometry-based method that provides residue-resolved quantitative information about protein phosphorylation. In this FLEXIQinase assay we combined our Full-Length Expressed Stable Isotope-labeled Protein for Quantification strategy (FLEXIQuant) with a traditional kinase assay to determine the mechanisms of multi-kinase substrate phosphorylation such as priming-dependent kinase activities. The assay monitors the decrease in signal intensity of the substrate peptides and the concomitant increase in the (n×80 Da)-shifted phosphorylated peptide. We analyzed the c-Jun N-terminal Kinase (JNK)-dependent glycogen synthase kinase 3β (GSK3β) activity on doublecortin (DCX) revealing mechanistic details about the role of phosphorylation cross-talk in GSK3β activity and permitting an advanced model for GSK3β-mediated signaling.Publication An OBSL1-Cul7\(^{\text{Fbxw8}}\) Ubiquitin Ligase Signaling Mechanism Regulates Golgi Morphology and Dendrite Patterning(Public Library of Science, 2011) Litterman, Nadia Kathryn; Ikeuchi, Yoshiho; Gallardo, Gilbert; O'Connell, Brenda C.; Sowa, Mathew E.; Gygi, Steven; Harper, Jeffrey; Bonni, AzadThe elaboration of dendrites in neurons requires secretory trafficking through the Golgi apparatus, but the mechanisms that govern Golgi function in neuronal morphogenesis in the brain have remained largely unexplored. Here, we report that the E3 ubiquitin ligase Cul7\(^{\text{Fbxw8}}\) localizes to the Golgi complex in mammalian brain neurons. Inhibition of Cul7\(^{\text{Fbxw8}}\) by independent approaches including Fbxw8 knockdown reveals that Cul7\(^{\text{Fbxw8}}\) is selectively required for the growth and elaboration of dendrites but not axons in primary neurons and in the developing rat cerebellum in vivo. Inhibition of Cul7\(^{\text{Fbxw8}}\) also dramatically impairs the morphology of the Golgi complex, leading to deficient secretory trafficking in neurons. Using an immunoprecipitation/mass spectrometry screening approach, we also uncover the cytoskeletal adaptor protein OBSL1 as a critical regulator of Cul7\(^{\text{Fbxw8}}\) in Golgi morphogenesis and dendrite elaboration. OBSL1 forms a physical complex with the scaffold protein Cul7 and thereby localizes Cul7 at the Golgi apparatus. Accordingly, OBSL1 is required for the morphogenesis of the Golgi apparatus and the elaboration of dendrites. Finally, we identify the Golgi protein Grasp65 as a novel and physiologically relevant substrate of Cul7\(^{\text{Fbxw8}}\) in the control of Golgi and dendrite morphogenesis in neurons. Collectively, these findings define a novel OBSL1-regulated Cul7\(^{\text{Fbxw8}}\) ubiquitin signaling mechanism that orchestrates the morphogenesis of the Golgi apparatus and patterning of dendrites, with fundamental implications for our understanding of brain development.