The epidemic of obesity and diabetes have increased interest in pathways that cause an increase in metabolic rates. The classical pathway of adaptive thermogenesis involves the function of mitochondrial Uncoupling Protein 1 (UCP1) in brown fat. This new work by the Spiegelman lab and published recently in Cell, shows that beige fat cells have another pathway used to generate heat and dissipate chemical energy. Their mitochondria utilize a futile cycle of creatine phosphorylation/dephosphorylation to elevate respiration and expend calories as heat. This new pathway may offer new insights into thermal regulation and offer new targets for human metabolic disorders.
NCOA4 is a selective cargo receptor for the autophagic turnover of ferritin, a process critical for regulation of intracellular iron bioavailability. However, how ferritinophagy flux is controlled and the roles of NCOA4 in iron-dependent processes are poorly understood. Through analysis of the NCOA4-FTH1 interaction, the Harper lab demonstrated that direct association via a key surface arginine in FTH1 and a C-terminal element in NCOA4 is required for delivery of ferritin to the lysosome via autophagosomes. Moreover, NCOA4 abundance is under dual control via autophagy and the ubiquitin proteasome system. Ubiquitin-dependent NCOA4 turnover is promoted by excess iron and involves an iron-dependent interaction between NCOA4 and the HERC2 ubiquitin ligase. In zebrafish and cultured cells, NCOA4 plays an essential role in erythroid differentiation. This work reveals the molecular nature of the NCOA4-ferritin complex and explains how intracellular iron levels modulate NCOA4-mediated ferritinophagy in cells and in an iron-dependent physiological setting.
Mancias, JD, Pontano Vaites L, Nissim S, Biancur DE, Kim AJ, Wang X, Liu Y, Goessling W, Kimmelman AC, Harper JW. (2015) Ferritinophagy via NCOA4 is required for erythroid development and is regulated by an iron dependent HERC2-mediated proteolysis. eLife
The AAA-ATPase VCP (also known as p97 or CDC48) uses ATP hydrolysis to ‘segregate’ ubiquitylated proteins from their binding partners, and has been implicated in numerous pathways ranging from ERAD to repair of damaged DNA. VCP acts through UBX-domain-containing adaptors that provide target specificity, but the targets and functions of UBXD proteins remain poorly understood. Through systematic proteomic analysis of UBXD proteins in human cells reported in Nature Cell Biology, the Harper lab revealed a network of over 195 interacting proteins, implicating VCP in diverse cellular pathways based on Gene Ontology analysis. Through detailed analysis of the interaction partners for the unstudied adaptor UBXN10, they uncover a role for this adaptor and VCP/p97 in ciliogenesis. UBXN10 associates with the intraflagellar transport B (IFT-B) complex, which regulates anterograde transport into cilia. Using TIRF microscopy in living cells, the demonstrate that UBXN10 is localized in cilia and traffics in cilia with the IFT-B complex. Moreover, deletion of UBXN10 using gene-editing renders cells unable to form cilia. Pharmacological inhibition of VCP destabilized they IFT-B complex and increased trafficking rates. Depletion of UBXN10 in zebrafish embryos causes defects in left–right asymmetry, which depends on functional cilia. This study provides a resource for exploring the landscape of UBXD proteins in biology and identifies an unexpected requirement for VCP–UBXN10 in ciliogenesis.
Raman, M, Sergeev, M, Garnaas M, Lydeard JR, Huttlin EL, Goessling W, Shah JV, and Harper JW. (2015) Systematic proteomics of the VCP–UBXD adaptor network identifies a role for UBXN10 in regulating ciliogenesis. Nature Cell Biology
Previous work has identified roles for the protein kinase PINK1 and the ubiquitin ligase PARKIN in the synthesis of ubiquitin chains on the surface of mitochondria in response to mitochondrial damage. But how these ubiquitin chains target damaged mitochondria to autophagosomes for “mitophagy” and the regulation of this process remained unclear. New work from the Harper Lab, published in Molecular Cell, defines a mechanism in which ubiquitin chains on mitochondria recruit 4 autophagy adaptors (OPTN, NDP52, p62, and TAX1BP1) through their ubiquitin binding domains to specific foci on damaged mitochondria. This recruitment leads to activation of the TBK1 kinase, which associates with OPTN, NDP52, and p62. In a concerted fashion, TBK1 phosphorylates OPTN, NDP52, and p62. Biochemical studies demonstrated that phosphorylation of OPTN by TBK1 further stimulates its intrinsic ubiquitin binding activity. Thus, this mechanism provides a self-reinforcing system that links these ubiquitin binding autophagy receptors with damaged mitochondria. Importantly, recent work has demonstrated that both TBK1 and OPTN are mutated in ALS. Thus, this work demonstrates that genes mutated in Parkinson’s Disease and ALS function in a common signal transduction pathway to help rid the cell of damaged mitochondria, and suggests that mechanisms for disposal of damaged mitochondria and possibly other toxic cellular components may be commonly targeted in such neurodegenerative diseases.
The Pex1 and Pex6 ATPases participate in peroxisome biogenesis. Mutations affecting these proteins are the most common cause of various generally fatal peroxisomal diseases. In a collaboration among the laboratories of Rapoport, Walz, and DiMaio/Baker, structures of the Pex1/Pex6 complex were determined in the presence of different nucleotides by cryo-electron microscopy (Blok et al., 2015). The structures support the hypothesis that the Pex1/Pex6 complex has a role in peroxisomal protein import analogous to that of another ATPase, p97, in ER-associated protein degradation.
Allied-Bristol Life Sciences, a joint venture between the university commercialization specialist Allied Minds and the US pharmaceutical company Bristol-Myers Squibb, is licensing research carried out by Malcolm Whitman on halofuginone, an active ingredient in blue evergreen hydrangea root. The Whitman Lab had showed several years ago that halofuginone could block the development of TH17-driven autoimmunity in a mouse model of multiple sclerosis by activating the amino acid response (AAR) pathway. They subsequently showed that halofuginone binds glutamyl-prolyl-tRNA synthetase (EPRS), which inhibits prolyl-tRNA synthetase activity; this inhibition of EPRSunderlies the broad bioactivities of this family of natural product derivatives. Now, Allied-Bristol Life Sciences is hoping to use this research to develop drugs to treat multipes types of autoimmune diseases.
Current and recent members of the Van Vactor and Perrimon laboratories at HMS have created a transgenic resource to allow conditional analysis of in vivo functions for over 140 microRNA genes in Drosophila with spatial and temporal precision. This resource, just published in Nature Communications, represents a highly collaborative project that will open the door to a broad variety of novel functional screens. Using this toolkit to explore the landscape of regulatory function in muscle tissue, they discovered a dozen microRNAs required for the maintenance of flight muscle form and function, suggesting that post-transcriptional mechanisms may be vital for protecting muscle from degeneration.
Using a combination of live cell imaging and single cell genome sequencing (Look-Seq), the Pellman laboratory has defined a mechanism for a new mutational process in cancer and congenital disease called chromothripsis (Zhang et al., Nature, 2015). In chromothripsis there is massive rearrangement of typically one of a cell’s chromosomes, leaving the rest of the genome unaltered. By recreating chromothripsis in the laboratory, the group shows that it can originate from abnormal nuclear structures, common in cancer cells, called micronuclei. The findings illustrate the importance of nuclear architecture and integrity for the maintenance of genome stability. The work was done in collaboration with Matthew Meyerson’s laboratory (Dana-Farber Cancer Institute & Department of Pathology at Harvard Medical School).
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Alternative splicing (AS) contributes to the proteomic diversity. In the brain, it emerges as a pervasive mechanism that plays a crucial role in the regulation of neuron maturation and activity. Chromatin modifying enzymes, that impact chromatin structure and globally control specific gene expression programs, are also subject to AS. However, at molecular level, it is poorly understood how AS can affect enzyme substrate specificity. In a recent publication in Molecular Cell (Laurent B. et al., PMID 25684206), Shi lab shed light on how AS can switch the enzymatic activity of the histone demethylase LSD1. In neurons, AS generates LSD1+8a, a LSD1 isoform containing an additional exon of 4 amino-acids (E8a). LSD1 has been reported to repress gene expression by demethylating histone H3K4. In their study, the authors show that the LSD1+8a isoform does not have the intrinsic capability to demethylate H3K4. Instead, LSD1+8a mediates H3K9 demethylation, in collaboration with the SVIL protein, and activate gene expression at its target promoters. Moreover, LSD1+8a and SVIL knockdowns increase H3K9 methylation levels at their target genes and compromise neuronal differentiation. These findings highlight AS as a means by which LSD1 acquires selective substrate specificities (H3K9 vs H3K4) to differentially control specific gene expression programs in neurons.
Note: This Shi lab publication was commented in its related Molecular Cell issue (Shin J. et al., PMID 25794611).
Breathing is essential for survival, and under precise neural control. The vagus nerve is a major connection between lung and brain required for normal respiration. In a recent publication in Cell, the Liberles Lab used molecular and genetic approaches to deconstruct the sensory vagus nerve, identifying two small populations of sensory neurons that exert powerful and opposing effects on breathing. Genetically guided anatomical mapping using Cre/LoxP technology revealed that these neurons densely innervate the lung and send long-range projections to different stereotyped brainstem targets. Optogenetic stimulation of one neuron type (P2ry1) acutely silences respiration, trapping animals in a state of exhalation, while activating another (Npy2r) causes rapid and shallow breathing. Activating P2ry1 neurons had no effect on heart rate and gastric pressure, other essential vagus nerve functions. Thus, the vagus nerve contains genetically definable labeled lines with different anatomical connections and autonomic roles. Specific manipulation of breathing-control neurons electrically or pharmacologically may impact airway diseases like asthma or apnea.