VCP-dependent muscle degeneration is linked to defects in a dynamic tubular lysosomal network in vivo.

Lysosomes are classically viewed as vesicular structures to which cargos are delivered for degradation. Here, we identify a network of dynamic, tubular lysosomes that extends throughout Drosophila muscle, in vivo. Live imaging reveals that autophagosomes merge with tubular lysosomes and that lysosomal membranes undergo extension, retraction, fusion and fission. The dynamics and integrity of this tubular lysosomal network requires VCP, an AAA-ATPase that, when mutated, causes degenerative diseases of muscle, bone and neurons. We show that human VCP rescues the defects caused by loss of Drosophila VCP and overexpression of disease relevant VCP transgenes dismantles tubular lysosomes, linking tubular lysosome dysfunction to human VCP-related diseases. Finally, disruption of tubular lysosomes correlates with impaired autophagosome-lysosome fusion, increased cytoplasmic poly-ubiquitin aggregates, lipofuscin material, damaged mitochondria and impaired muscle function. We propose that VCP sustains sarcoplasmic proteostasis, in part, by controlling the integrity of a dynamic tubular lysosomal network

CK1 is required for a mitotic checkpoint that delays cytokinesis

Failure to accurately partition genetic material during cell division causes aneuploidy and drives tumorigenesis [1]. Cell-cycle checkpoints safeguard cells from such catastrophes by impeding cell-cycle progression when mistakes arise. FHA-RING E3 ligases, including human RNF8 [2] and CHFR [3] and fission yeast Dma1 [4], relay checkpoint signals by binding phosphorylated proteins via their FHA domains and promoting ubiquitination of downstream targets [5]. Upon mitotic checkpoint activation, S. pombe Dma1 concentrates at spindle pole bodies (SPBs) in an FHA-dependent manner and ubiquitinates Sid4, a scaffold of Polo kinase, to suspend cytokinesis [6]. However, the kinase or kinases that phosphoprime Sid4 for Dma1-mediated ubiquitination are unknown. Here, we report that the highly conserved protein kinase CK1 transmits the signal necessary to stall cytokinesis by phosphopriming Sid4 for Dma1-mediated ubiquitination. Like Dma1, CK1 accumulates at SPBs during a mitotic arrest and associates stably with SPB components, including Sid4. Our results establish CK1 as an integral component of a mitotic, ubiquitin-mediated checkpoint pathway. © 2013 Elsevier Ltd

Fission yeast Dma1 requires RING domain dimerization for its ubiquitin ligase activity and mitotic checkpoint function

In fission yeast (Schizosaccharomyces pombe), the E3 ubiquitin ligase Dma1 delays cytokinesis if chromosomes are not properly attached to the mitotic spindle.Dma1contains a C-terminal RING domain, and we have found that the Dma1 RING domain forms a stable homodimer. Although the RING domain is required for dimerization, residues in the C-terminal tail are also required to help form or stabilize the dimeric structure because mutation of specific residues in this region disrupts Dma1 dimerization. Further analyses showed that Dma1 dimerization is required for proper localization at spindle pole bodies and the cell division site, E3 ligase activity, and mitotic checkpoint function. Thus, Dma1 forms an obligate dimer via its RING domain, which is essential for efficient transfer of ubiquitin to its substrate(s). This study further supports the mechanistic paradigm that many RING E3 ligases function as RING dimers. © 2012 by The American Society for Biochemistry and Molecular Biology, Inc

Cdk1 promotes cytokinesis in fission yeast through activation of the septation initiation network

In Schizosaccharomyces pombe, late mitotic events are coordinated with cytokinesis by the septation initiation network (SIN), an essential spindle pole body (SPB)-associated kinase cascade, which controls the formation, maintenance, and constriction of the cytokinetic ring. It is not fully understood how SIN initiation is temporally regulated, but it depends on the activation of the GTPase Spg1, which is inhibited during interphase by the essential bipartite GTPase-activating protein Byr4-Cdc16. Cells are particularly sensitive to the modulation of Byr4, which undergoes cell cycle-dependent phosphorylation presumed to regulate its function. Polo-like kinase, which promotes SIN activation, is partially responsible for Byr4 phosphorylation. Here we show that Byr4 is also controlled by cyclin-dependent kinase (Cdk1)-mediated phosphorylation. A Cdk1 nonphosphorylatable Byr4 phosphomutant displays severe cell division defects, including the formation of elongated, multinucleate cells, failure to maintain the cytokinetic ring, and compromised SPB association of the SIN kinase Cdc7. Our analyses show that Cdk1-mediated phosphoregulation of Byr4 facilitates complete removal of Byr4 from metaphase SPBs in concert with Plo1, revealing an unexpected role for Cdk1 in promoting cytokinesis through activation of the SIN pathway. © 2014 Rachfall et al

Relief of the Dma1-mediated checkpoint requires Dma1 autoubiquitination and dynamic localization

© 2018 Jones, Chen, et al. Chromosome segregation and cell division are coupled to prevent aneuploidy and cell death. In the fission yeast Schizosaccharomyces pombe, the septation initiation network (SIN) promotes cytokinesis, but upon mitotic checkpoint activation, the SIN is actively inhibited to prevent cytokinesis from occurring before chromosomes have safely segregated. SIN inhibition during the mitotic checkpoint is mediated by the E3 ubiquitin ligase Dma1. Dma1 binds to the CK1-phosphorylated SIN scaffold protein Sid4 at the spindle pole body (SPB), and ubiquitinates it. Sid4 ubiquitination antagonizes the SPB localization of the Polo-like kinase Plo1, the major SIN activator, so that SIN signaling is delayed. How this checkpoint is silenced once spindle defects are resolved has not been clear. Here we establish that Dma1 transiently leaves SPBs during anaphase B due to extensive autoubiquitination. The SIN is required for Dma1 to return to SPBs later in anaphase. Blocking Dma1 removal from SPBs by permanently tethering it to Sid4 prevents SIN activation and cytokinesis. Therefore, controlling Dma1’s SPB dynamics in anaphase is an essential step in S. pombe cell division and the silencing of the Dma1-dependent mitotic checkpoint

A density functional theory analysis of the adsorption and surface chemistry of inorganic iodine species on graphitea

In the event of a nuclear accident, fission products may be released into the environment. The release of 131I is of particular concern to human health. Iodine can be captured using a number of materials and frequently, this is accomplished with activated carbon impregnated with organic bases. Previous studies have used DFT and the graphite (0001) surface as a surrogate for adsorption, those studies focus on the species I•, I2, and CH3I. In this work we perform an ab initio study of the adsorption onto the surface of a graphite sheet of I2, CH3I, and inorganic acidic iodine species (HI, HOI, HIO2, and HIO3), which were selected to examine the possible effect of oxidation state on adsorption. The PBE exchange-correlation functional with D3 dispersion was employed. It was found that for molecular iodine, the iodine atoms tended to either situate above the center of a hexagonal site on the graphite or directly atop a carbon atom with the lighter components resting closer to the graphite. For each species the relative binding energies spanned the range of 21–33 kJ mol-1 and graphite-iodine distance was in the range of 3.52–3.93 Å. In all cases we found no significant charge transfer between the iodine species and the graphite, thus we conclude that all the iodine species studied undergo strong physisorption to the graphite

Physiology and evolution of nitrate acquisition in Prochlorococcus

Prochlorococcus is the numerically dominant phototroph in the oligotrophic subtropical ocean and carries out a significant fraction of marine primary productivity. Although field studies have provided evidence for nitrate uptake by Prochlorococcus, little is known about this trait because axenic cultures capable of growth on nitrate have not been available. Additionally, all previously sequenced genomes lacked the genes necessary for nitrate assimilation. Here we introduce three Prochlorococcus strains capable of growth on nitrate and analyze their physiology and genome architecture. We show that the growth of high-light (HL) adapted strains on nitrate is ~17% slower than their growth on ammonium. By analyzing 41 Prochlorococcus genomes, we find that genes for nitrate assimilation have been gained multiple times during the evolution of this group, and can be found in at least three lineages. In low-light adapted strains, nitrate assimilation genes are located in the same genomic context as in marine Synechococcus. These genes are located elsewhere in HL adapted strains and may often exist as a stable genetic acquisition as suggested by the striking degree of similarity in the order, phylogeny and location of these genes in one HL adapted strain and a consensus assembly of environmental Prochlorococcus metagenome sequences. In another HL adapted strain, nitrate utilization genes may have been independently acquired as indicated by adjacent phage mobility elements; these genes are also duplicated with each copy detected in separate genomic islands. These results provide direct evidence for nitrate utilization by Prochlorococcus and illuminate the complex evolutionary history of this trait.Gordon and Betty Moore Foundation (Grant GBMF495)National Science Foundation (U.S.) (Grant OCE-1153588)National Science Foundation (U.S.) (Grant DBI-0424599