The stoichiometry of the outer kinetochore is modulated by microtubule-proximal regulatory factors
We report that Fin1 localizes to the microtubule-proximal edge of the kinetochore cluster during anaphase based on single-particle averaging of super-resolution images. Fin1 is required for the assembly of normal levels of Dam1 and Ndc80 submodules. Levels of Ndc80 further depend on the Dam1 microtubule binding complex. Our results suggest the stoichiometry of outer kinetochore submodules is strongly influenced by factors at the kinetochore–microtubule interface such as Fin1 and Dam1, and phosphorylation by cyclin-dependent kinase. Outer kinetochore stoichiometry is remarkably plastic and responsive to microtubule-pr...
Source: Journal of Cell Biology - June 30, 2019 Category: Cytology Authors: Dhatchinamoorthy, K., Unruh, J. R., Lange, J. J., Levy, M., Slaughter, B. D., Gerton, J. L. Tags: Cell Cycle and Division, Genetics Reports Source Type: research
XLF and H2AX function in series to promote replication fork stability
XRCC4-like factor (XLF) is a non-homologous end joining (NHEJ) DNA double strand break repair protein. However, XLF deficiency leads to phenotypes in mice and humans that are not necessarily consistent with an isolated defect in NHEJ. Here we show that XLF functions during DNA replication. XLF undergoes cell division cycle 7–dependent phosphorylation; associates with the replication factor C complex, a critical component of the replisome; and is found at replication forks. XLF deficiency leads to defects in replication fork progression and an increase in fork reversal. The additional loss of H2AX, which protects DNA ...
Source: Journal of Cell Biology - June 30, 2019 Category: Cytology Authors: Chen, B.-R., Quinet, A., Byrum, A. K., Jackson, J., Berti, M., Thangavel, S., Bredemeyer, A. L., Hindi, I., Mosammaparast, N., Tyler, J. K., Vindigni, A., Sleckman, B. P. Tags: Cell Cycle and Division, DNA Biology, Cancer, Genetics Reports Source Type: research
Beyond the Cell: The cell biology of the hepatocyte: A membrane trafficking machine
The liver performs numerous vital functions, including the detoxification of blood before access to the brain while simultaneously secreting and internalizing scores of proteins and lipids to maintain appropriate blood chemistry. Furthermore, the liver also synthesizes and secretes bile to enable the digestion of food. These diverse attributes are all performed by hepatocytes, the parenchymal cells of the liver. As predicted, these cells possess a remarkably well-developed and complex membrane trafficking machinery that is dedicated to moving specific cargos to their correct cellular locations. Importantly, while most epit...
Source: Journal of Cell Biology - June 30, 2019 Category: Cytology Authors: Schulze, R. J., Schott, M. B., Casey, C. A., Tuma, P. L., McNiven, M. A. Tags: Review Source Type: research
Expecto Patronin for slow and persistent minus end microtubule growth in dendrites
Microtubule plus ends are highly dynamic in neurons, while minus ends are often capped and stable. In this issue, Feng et al. (2019. J. Cell Biol. https://doi.org/10.1083/jcb.201810155) demonstrate that in dendrites, free minus ends undergo slow and processive growth mediated by the minus end–binding protein Patronin. (Source: Journal of Cell Biology)
Source: Journal of Cell Biology - June 30, 2019 Category: Cytology Authors: Broihier, H. T. Tags: Spotlight Source Type: research
The enemy of my enemy: PTEN and PLCXD collude to fight endosomal PtdIns(4,5)P2
Loss of the phosphoinositide 5-phosphatase OCRL causes accumulation of PtdIns(4,5)P2 on membranes and, ultimately, Lowe syndrome. In this issue, Mondin et al. (2019. J. Cell Biol. https://doi.org/10.1083/jcb.201805155) discover that a surprising partnership between PTEN and the phospholipase PLCXD can compensate for OCRL to suppress endosomal PtdIns(4,5)P2 accumulation. (Source: Journal of Cell Biology)
Source: Journal of Cell Biology - June 30, 2019 Category: Cytology Authors: Del Signore, S. J., Rodal, A. A. Tags: Spotlight Source Type: research
Defects in the origin licensing checkpoint stresses cells exiting G0
The full licensing of replication origins in late G1 is normally enforced by the licensing checkpoint. In this issue, Matson et al. (2019. J. Cell Biol. https://doi.org/10.1083/jcb.201902143) show that this checkpoint is inoperative in cells exiting from G0, resulting in incomplete origin licensing and consequent replicative stress. (Source: Journal of Cell Biology)
Source: Journal of Cell Biology - June 30, 2019 Category: Cytology Authors: Blow, J. J. Tags: Cell Cycle and Division, Biochemistry Spotlight Source Type: research
Loss of nuclear envelope integrity? No probLEM--BAF has it covered
Ruptures of the nuclear envelope can drive a catastrophic loss of nucleocytoplasmic compartmentalization. In this issue, Halfmann et al. (2019. J. Cell Biol. https://doi.org/10.1083/jcb.201901116) describe a mechanism for surveilling the integrity of the nuclear barrier and coupling the sensing of nuclear ruptures to the recruitment of the nuclear envelope repair machinery. (Source: Journal of Cell Biology)
Source: Journal of Cell Biology - June 30, 2019 Category: Cytology Authors: King, M. C., Lusk, C. P. Tags: Spotlight Source Type: research
XLF extends its range from DNA repair to replication
The close interplay between DNA replication and repair is underscored by a report from Chen et al. (2019. J. Cell Biol. https://doi.org/10.1083/jcb.201808134) in this issue. The authors demonstrate that the non-homologous end-joining factor XLF promotes the stability of replication forks. (Source: Journal of Cell Biology)
Source: Journal of Cell Biology - June 30, 2019 Category: Cytology Authors: Gonzalez-Rodriguez, Y., Bunting, S. F. Tags: Chromatin or Epigenetics, DNA Biology, Biochemistry, Genetics Spotlight Source Type: research
Greg Alushin: The shape of things to come
Alushin investigates the structural biology of biomechanical processes in the cytoskeleton. (Source: Journal of Cell Biology)
Source: Journal of Cell Biology - June 30, 2019 Category: Cytology Authors: ODonnell, M. A. Tags: People & amp;amp; Ideas Source Type: research
Correction: GGCX and VKORC1 inhibit osteocalcin endocrine functions
Vol. 208, No. 6, March 16, 2015. 10.1083/jcb.201409111. The authors noticed a mistake in the labeling of the genotyping primers in Fig. S1 A and Table S1. In Fig. S1 A, the primers P1 and P2 are... (Source: Journal of Cell Biology)
Source: Journal of Cell Biology - June 2, 2019 Category: Cytology Authors: Ferron, M., Lacombe, J., Germain, A., Oury, F., Karsenty, G. Tags: Corrections Source Type: research
Proteome of the central apparatus of a ciliary axoneme
Nearly all motile cilia have a "9+2" axoneme containing a central apparatus (CA), consisting of two central microtubules with projections, that is essential for motility. To date, only 22 proteins are known to be CA components. To identify new candidate CA proteins, we used mass spectrometry to compare axonemes of wild-type Chlamydomonas and a CA-less mutant. We identified 44 novel candidate CA proteins, of which 13 are conserved in humans. Five of the latter were studied more closely, and all five localized to the CA; therefore, most of the other candidates are likely to also be CA components. Our results reveal that the ...
Source: Journal of Cell Biology - June 2, 2019 Category: Cytology Authors: Zhao, L., Hou, Y., Picariello, T., Craige, B., Witman, G. B. Tags: Disease, Cilia Tools Source Type: research
Comprehensive knockout analysis of the Rab family GTPases in epithelial cells
The Rab family of small GTPases comprises the largest number of proteins (~60 in mammals) among the regulators of intracellular membrane trafficking, but the precise function of many Rabs and the functional redundancy and diversity of Rabs remain largely unknown. Here, we generated a comprehensive collection of knockout (KO) MDCK cells for the entire Rab family. We knocked out closely related paralogs simultaneously (Rab subfamily knockout) to circumvent functional compensation and found that Rab1A/B and Rab5A/B/C are critical for cell survival and/or growth. In addition, we demonstrated that Rab6-KO cells lack the basemen...
Source: Journal of Cell Biology - June 2, 2019 Category: Cytology Authors: Homma, Y., Kinoshita, R., Kuchitsu, Y., Wawro, P. S., Marubashi, S., Oguchi, M. E., Ishida, M., Fujita, N., Fukuda, M. Tags: Trafficking Tools Source Type: research
Peroxisome protein import recapitulated in Xenopus egg extracts
Peroxisomes import their luminal proteins from the cytosol. Most substrates contain a C-terminal Ser-Lys-Leu (SKL) sequence that is recognized by the receptor Pex5. Pex5 binds to peroxisomes via a docking complex containing Pex14, and recycles back into the cytosol following its mono-ubiquitination at a conserved Cys residue. The mechanism of peroxisome protein import remains incompletely understood. Here, we developed an in vitro import system based on Xenopus egg extracts. Import is dependent on the SKL motif in the substrate and on the presence of Pex5 and Pex14, and is sustained by ATP hydrolysis. A protein lacking an ...
Source: Journal of Cell Biology - June 2, 2019 Category: Cytology Authors: Romano, F. B., Blok, N. B., Rapoport, T. A. Tags: Organelles, Trafficking, Biochemistry, Biophysics Tools Source Type: research
Engineered SUMO/protease system identifies Pdr6 as a bidirectional nuclear transport receptor
Cleavage of affinity tags by specific proteases can be exploited for highly selective affinity chromatography. The SUMO/SENP1 system is the most efficient for such application but fails in eukaryotic expression because it cross-reacts with endogenous proteases. Using a novel selection system, we have evolved the SUMOEu/SENP1Eu pair to orthogonality with the yeast and animal enzymes. SUMOEu fusions therefore remain stable in eukaryotic cells. Likewise, overexpressing a SENP1Eu protease is nontoxic in yeast. We have used the SUMOEu system in an affinity-capture-proteolytic-release approach to identify interactors of the yeas...
Source: Journal of Cell Biology - June 2, 2019 Category: Cytology Authors: Vera Rodriguez, A., Frey, S., Görlich, D. Tags: Organelles, Trafficking, Biochemistry, Technology Tools Source Type: research
Necroptosis mediators RIPK3 and MLKL suppress intracellular Listeria replication independently of host cell killing
RIPK3, a key mediator of necroptosis, has been implicated in the host defense against viral infection primary in immune cells. However, gene expression analysis revealed that RIPK3 is abundantly expressed not only in immune organs but also in the gastrointestinal tract, particularly in the small intestine. We found that orally inoculated Listeria monocytogenes, a bacterial foodborne pathogen, efficiently spread and caused systemic infection in Ripk3-deficient mice while almost no dissemination was observed in wild-type mice. Listeria infection activated the RIPK3-MLKL pathway in cultured cells, which resulted in suppressio...
Source: Journal of Cell Biology - June 2, 2019 Category: Cytology Authors: Sai, K., Parsons, C., House, J. S., Kathariou, S., Ninomiya-Tsuji, J. Tags: Cell Death and Autophagy, Microbiology Articles Source Type: research
