The Escherichia coli Outer Membrane ß-Barrel Assembly Machinery (BAM) Crosstalks with the Divisome.

The BAM is a macromolecular machine responsible for the folding and the insertion of integral proteins into the outer membrane of diderm Gram-negative bacteria. In Escherichia coli, it consists of a transmembrane ß-barrel subunit, BamA, and four outer membrane lipoproteins (BamB-E). Using BAM-specific antibodies, in E. coli cells, the complex is shown to localize in the lateral wall in foci. The machinery was shown to be enriched at midcell with specific cell cycle timing. The inhibition of septation by aztreonam did not alter the BAM midcell localization substantially. Furthermore, the absence of late cell division proteins at midcell did not impact BAM timing or localization. These results imply that the BAM enrichment at the site of constriction does not require an active cell division machinery. Expression of the Tre1 toxin, which impairs the FtsZ filamentation and therefore midcell localization, resulted in the complete loss of BAM midcell enrichment. A similar effect was observed for YidC, which is involved in the membrane insertion of cell division proteins in the inner membrane. The presence of the Z-ring is needed for preseptal peptidoglycan (PG) synthesis. As BAM was shown to be embedded in the PG layer, it is possible that BAM is inserted preferentially simultaneously with de novo PG synthesis to facilitate the insertion of OMPs in the newly synthesized outer membrane.

Molecular architecture and modifications of full-length myocilin.

Myocilin, a modular multidomain protein, is expressed broadly in the human body but is best known for its presence in the trabecular meshwork extracellular matrix, and myocilin misfolding is associated with glaucoma. Despite progress in comprehending the structure and misfolding of the myocilin olfactomedin domain, the structure and function of full-length myocilin, and contextual changes in glaucoma, remain unknown. Here we expressed and purified milligram-scale quantities of full-length myocilin from suspension mammalian cell culture (Expi293F), enabling molecular characterization in detail not previously accessible. We systematically characterized disulfide-dependent and -independent oligomerization as well as confirmed glycosylation and susceptibility to proteolysis. We identified oligomeric states with glycosylation sites that are inaccessible to enzymatic removal. Low-resolution single particle 2D class averaging from conventional transmission electron microscopy imaging confirms an extended arrangement of tetramers, truncated products consistent with dimers, and a higher-ordered state consistent with octamer. Taken together, our study reveals new myocilin misfolded states and layers of intrinsic heterogeneity, expands our knowledge of olfactomedin-family proteins and lays the foundation for a better molecular understanding of myocilin structure and its still enigmatic biological function.

Protein haploinsufficiency drivers identify MYBPC3 variants that cause hypertrophic cardiomyopathy.

Hypertrophic cardiomyopathy (HCM) is the most common inherited cardiac disease. Variants in MYBPC3, the gene encoding cardiac myosin-binding protein C (cMyBP-C), are the leading cause of HCM. However, the pathogenicity status of hundreds of MYBPC3 variants found in patients remains unknown, as a consequence of our incomplete understanding of the pathomechanisms triggered by HCM-causing variants. Here, we examined 44 nontruncating MYBPC3 variants that we classified as HCM-linked or nonpathogenic according to cosegregation and population genetics criteria. We found that around half of the HCM-linked variants showed alterations in RNA splicing or protein stability, both of which can lead to cMyBP-C haploinsufficiency. These protein haploinsufficiency drivers associated with HCM pathogenicity with 100% and 94% specificity, respectively. Furthermore, we uncovered that 11% of nontruncating MYBPC3 variants currently classified as of uncertain significance in ClinVar induced one of these molecular phenotypes. Our strategy, which can be applied to other conditions induced by protein loss of function, supports the idea that cMyBP-C haploinsufficiency is a fundamental pathomechanism in HCM.

Meiotic regulation of the Ndc80 complex composition and function.

This review describes the current models for how the subunit abundance of the Ndc80 complex, a key kinetochore component, is regulated in budding yeast and metazoan meiosis. The past decades of kinetochore research have established the Ndc80 complex to be a key microtubule interactor and a central hub for regulating chromosome segregation. Recent studies further demonstrate that Ndc80 is the limiting kinetochore subunit that dictates the timing of kinetochore activation in budding yeast meiosis. Here, we discuss the molecular circuits that regulate Ndc80 protein synthesis and degradation in budding yeast meiosis and compare the findings with those from metazoans. We envision the regulatory principles discovered in budding yeast to be conserved in metazoans, thereby providing guidance into future investigations on kinetochore regulation in human health and disease.

Arc self-association and formation of virus-like capsids are mediated by an N-terminal helical coil motif.

Activity-regulated cytoskeleton-associated protein (Arc) is a protein interaction hub with diverse roles in intracellular neuronal signaling, and important functions in neuronal synaptic plasticity, memory, and postnatal cortical development. Arc has homology to retroviral Gag protein and is capable of self-assembly into virus-like capsids implicated in the intercellular transfer of RNA. However, the molecular basis of Arc self-association and capsid formation is largely unknown. Here, we identified a 28-amino-acid stretch in the mammalian Arc N-terminal (NT) domain that is necessary and sufficient for self-association. Within this region, we identified a 7-residue oligomerization motif, critical for the formation of virus-like capsids. Purified wild-type Arc formed capsids as shown by transmission and cryo-electron microscopy, whereas mutant Arc with disruption of the oligomerization motif formed homogenous dimers. An atomic-resolution crystal structure of the oligomerization region peptide demonstrated an antiparallel coiled-coil interface, strongly supporting NT-NT domain interactions in Arc oligomerization. The NT coil-coil interaction was also validated in live neurons using fluorescence lifetime FRET imaging, and mutation of the oligomerization motif disrupted Arc-facilitated endocytosis. Furthermore, using single-molecule photobleaching, we show that Arc mRNA greatly enhances higher-order oligomerization in a manner dependent on the oligomerization motif. In conclusion, a helical coil in the Arc NT domain supports self-association above the dimer stage, mRNA-induced oligomerization, and formation of virus-like capsids. DATABASE: The coordinates and structure factors for crystallographic analysis of the oligomerization region were deposited at the Protein Data Bank with the entry code 6YTU.

Assembly properties of bacterial tubulin homolog FtsZ regulated by the positive regulator protein ZipA and ZapA from Pseudomonas aeruginosa.

Bacterial tubulin homolog FtsZ self-assembles into dynamic protofilaments, which forms the scaffold for the contractile ring (Z-ring) to achieve bacterial cell division. Here, we study the biochemical properties of FtsZ from Pseudomonas aeruginosa (PaFtsZ) and the effects of its two positive regulator proteins, ZipA and ZapA. Similar to Escherichia coli FtsZ, PaFtsZ had a strong GTPase activity, ~ 7.8 GTP min-1 FtsZ-1 at pH 7.5, and assembled into mainly short single filaments in vitro. However, PaFtsZ protofilaments were mixtures of straight and "intermediate-curved" (100-300 nm diameter) in pH 7.5 solution and formed some bundles in pH 6.5 solution. The effects of ZipA on PaFtsZ assembly varied with pH. In pH 6.5 buffer ZipA induced PaFtsZ to form large bundles. In pH 7.5 buffer PaFtsZ-ZipA protofilaments were not bundled, but ZipA enhanced PaFtsZ assembly and promoted more curved filaments. Comparable to ZapA from other bacterial species, ZapA from P. aeruginosa induced PaFtsZ protofilaments to associate into long straight loose bundles and/or sheets at both pH 6.5 and pH 7.5, which had little effect on the GTPase activity of PaFtsZ. These results provide us further information that ZipA functions as an enhancer of FtsZ curved filaments, while ZapA works as a stabilizer of FtsZ straight filaments.

Correlating STED and synchrotron XRF nano-imaging unveils cosegregation of metals and cytoskeleton proteins in dendrites.

Zinc and copper are involved in neuronal differentiation and synaptic plasticity but the molecular mechanisms behind these processes are still elusive due in part to the difficulty of imaging trace metals together with proteins at the synaptic level. We correlate stimulated-emission-depletion microscopy of proteins and synchrotron X-ray fluorescence imaging of trace metals, both performed with 40 nm spatial resolution, on primary rat hippocampal neurons. We reveal the co-localization at the nanoscale of zinc and tubulin in dendrites with a molecular ratio of about one zinc atom per tubulin-αß dimer. We observe the co-segregation of copper and F-actin within the nano-architecture of dendritic protrusions. In addition, zinc chelation causes a decrease in the expression of cytoskeleton proteins in dendrites and spines. Overall, these results indicate new functions for zinc and copper in the modulation of the cytoskeleton morphology in dendrites, a mechanism associated to neuronal plasticity and memory formation.

A bacterial cytolinker couples positioning of magnetic organelles to cell shape control.

Magnetotactic bacteria maneuver within the geomagnetic field by means of intracellular magnetic organelles, magnetosomes, which are aligned into a chain and positioned at midcell by a dedicated magnetosome-specific cytoskeleton, the "magnetoskeleton." However, how magnetosome chain organization and resulting magnetotaxis is linked to cell shape has remained elusive. Here, we describe the cytoskeletal determinant CcfM (curvature-inducing coiled-coil filament interacting with the magnetoskeleton), which links the magnetoskeleton to cell morphology regulation in Magnetospirillum gryphiswaldense Membrane-anchored CcfM localizes in a filamentous pattern along regions of inner positive-cell curvature by its coiled-coil motifs, and independent of the magnetoskeleton. CcfM overexpression causes additional circumferential localization patterns, associated with a dramatic increase in cell curvature, and magnetosome chain mislocalization or complete chain disruption. In contrast, deletion of ccfM results in decreased cell curvature, impaired cell division, and predominant formation of shorter, doubled chains of magnetosomes. Pleiotropic effects of CcfM on magnetosome chain organization and cell morphology are supported by the finding that CcfM interacts with the magnetoskeleton-related MamY and the actin-like MamK via distinct motifs, and with the cell shape-related cytoskeleton via MreB. We further demonstrate that CcfM promotes motility and magnetic alignment in structured environments, and thus likely confers a selective advantage in natural habitats of magnetotactic bacteria, such as aquatic sediments. Overall, we unravel the function of a prokaryotic cytoskeletal constituent that is widespread in magnetic and nonmagnetic spirilla-shaped Alphaproteobacteria.

Microtubule Attachment and Centromeric Tension Shape the Protein Architecture of the Human Kinetochore.

The nanoscale protein architecture of the kinetochore plays an integral role in specifying the mechanisms underlying its functions in chromosome segregation. However, defining this architecture in human cells remains challenging because of the large size and compositional complexity of the kinetochore. Here, we use Förster resonance energy transfer to reveal the architecture of individual kinetochore-microtubule attachments in human cells. We find that the microtubule-binding domains of the Ndc80 complex cluster at the microtubule plus end. This clustering occurs only after microtubule attachment, and it increases proportionally with centromeric tension. Surprisingly, Ndc80 complex clustering is independent of the organization and number of its centromeric receptors. Moreover, this clustering is similar in yeast and human kinetochores despite significant differences in their centromeric organizations. These and other data suggest that the microtubule-binding interface of the human kinetochore behaves like a flexible "lawn" despite being nucleated by repeating biochemical subunits.

WDR90 is a centriolar microtubule wall protein important for centriole architecture integrity.

Centrioles are characterized by a nine-fold arrangement of microtubule triplets held together by an inner protein scaffold. These structurally robust organelles experience strenuous cellular processes such as cell division or ciliary beating while performing their function. However, the molecular mechanisms underlying the stability of microtubule triplets, as well as centriole architectural integrity remain poorly understood. Here, using ultrastructure expansion microscopy for nanoscale protein mapping, we reveal that POC16 and its human homolog WDR90 are components of the microtubule wall along the central core region of the centriole. We further found that WDR90 is an evolutionary microtubule associated protein. Finally, we demonstrate that WDR90 depletion impairs the localization of inner scaffold components, leading to centriole structural abnormalities in human cells. Altogether, this work highlights that WDR90 is an evolutionary conserved molecular player participating in centriole architecture integrity.

Cells are made up of compartments called organelles that perform specific roles. A cylindrical organelle called the centriole is important for a number of cellular processes, ranging from cell division to movement and signaling. Each centriole contains nine blades made up of protein filaments called microtubules, which link together to form a cylinder. This well-known structure can be found in a variety of different species. Yet, it is unclear how centrioles are able to maintain this stable architecture whilst carrying out their various different cell roles. In early 2020, a group of researchers discovered a scaffold protein at the center of centrioles that helps keep the microtubule blades stable. Further investigation suggested that another protein called WDR90 may also help centrioles sustain their cylindrical shape. However, the exact role of this protein was poorly understood. To determine the role of WDR90, Steib et al. ­ including many of the researchers involved in the 2020 study ­ used a method called Ultrastructure Expansion Microscopy to precisely locate the WDR90 protein in centrioles. This revealed that WDR90 is located on the microtubule wall of centrioles in green algae and human cells grown in the lab. Further experiments showed that the protein binds directly to microtubules and that removing WDR90 from human cells causes centrioles to lose their scaffold proteins and develop structural defects. This investigation provides fundamental insights into the structure and stability of centrioles. It shows that single proteins are key components in supporting the structural integrity of organelles and shaping their overall architecture. Furthermore, these findings demonstrate how ultrastructure expansion microscopy can be used to determine the role of individual proteins within a complex structure.

Crystal Structure of Keratin 1/10(C401A) 2B Heterodimer Demonstrates a Proclivity for the C-Terminus of Helix 2B to Form Higher Order Molecular Contacts.

We previously determined the crystal structure of the wild-type keratin 1/10 helix 2B heterodimer at 3.3 Å resolution. We proposed that the resolution of the diffraction data was limited due to the crystal packing effect from keratin 10 (K10) residue Cys401. Cys401K10 formed a disulfide-linkage with Cys401 from another K1/10 heterodimer, creating an "X-shaped" structure and a loose crystal packing arrangement. We hypothesized that mutation of Cys401K10 to alanine would eliminate the disulfide-linkage and improve crystal packing thereby increasing resolution of diffraction and enabling a more accurate side chain electron density map. Indeed, when a K10 Cys401Ala 2B mutant was paired with its native keratin 1 (K1) 2B heterodimer partner its x-ray crystal structure was determined at 2.07 Å resolution; the structure does not contain a disulfide linkage. Superposition of the K1/K10(Cys401Ala) 2B structure onto the wild-type K1/10 2B heterodimer structure had a root-mean-square-deviation of 1.88 Å; the variability in the atomic positions reflects the dynamic motion expected in this filamentous coiled-coil complex. The electrostatic, hydrophobic, and contour features of the molecular surface are similar to the lower resolution wild-type structure. We postulated that elimination of the disulfide linkage in the K1/K10(Cys401Ala) 2B structure could allow for the 2B heterodimers to bind/pack in the A22 tetramer configuration associated with mature keratin intermediate filament assembly. Analysis of the crystal packing revealed a half-staggered anti-parallel tetrameric complex of 2B heterodimers; however, their register is not consistent with models of the A22 mode of tetrameric alignment or prior biochemical cross-linking studies.

Identification of a Structurally Dynamic Domain for Oligomer Formation in Rootletin.

Rootletin is the main component of the ciliary rootlet and functions as a centriole linker connecting the two mother centrioles. Despite the functional importance of rootletin, the molecular architecture of the rootletin filament and its assembly mechanism are poorly understood. Here, we identify the coiled-coil domain 3 (CCD3) of rootletin as the key domain for its cellular function. The crystal structure of the CCD31108-1317 fragment containing 28 heptad repeats and 1 hendecad repeat reveals that it forms a parallel coiled-coil dimer spanning approximately 300 Šin length. Crosslinking experiments and biophysical analyses of the minimal functional region of CCD3 (CCD3-6) suggest that CCD3-6 is structurally dynamic and may be important for oligomer formation. We also show that oligomerization-defective CCD3 mutants fail in centrosomal localization and centriole linkage, suggesting that rootletin oligomerization may be important for its function.

A multipronged approach to understanding the form and function of hStaufen protein.

Staufen is a dsRNA-binding protein involved in many aspects of RNA regulation, such as mRNA transport, Staufen-mediated mRNA decay and the regulation of mRNA translation. It is a modular protein characterized by the presence of conserved consensus amino acid sequences that fold into double-stranded RNA binding domains (RBDs) as well as degenerated RBDs that are instead involved in protein-protein interactions. The variety of biological processes in which Staufen participates in the cell suggests that this protein associates with many diverse RNA targets, some of which have been identified experimentally. Staufen binding mediates the recruitment of effectors via protein-protein and protein-RNA interactions. The structural determinants of a number of these interactions, as well as the structure of full-length Staufen, remain unknown. Here, we present the first solution structure models for full-length hStaufen155, showing that its domains are arranged as beads-on-a-string connected by flexible linkers. In analogy with other nucleic acid-binding proteins, this could underpin Stau1 functional plasticity.

Structural Evidence for an Octameric Ring Arrangement of SARM1.

SARM1 induces axonal degeneration in response to various insults and is therefore considered an attractive drug target for the treatment of neuro-degenerative diseases as well as for brain and spinal cord injuries. SARM1 activity depends on the integrity of the protein's SAM domains, as well as on the enzymatic conversion of NAD+ to ADPR (ADP Ribose) products by the SARM1's TIR domain. Therefore, inhibition of either SAM or TIR functions may constitute an effective therapeutic strategy. However, there is currently no SARM1-directed therapeutic approach available because of an insufficient structural and mechanistic understanding of this protein. In this study, we found that SARM1 assembles into an octameric ring. This arrangement was not described before in other SAM proteins, but is reminiscent of the apoptosome and inflammasome-well-known apoptotic ring-like oligomers. We show that both SARM1 and the isolated tandem SAM1-2 domains form octamers in solution, and electron microscopy analysis reveals an octameric ring of SARM1. We determined the crystal structure of SAM1-2 and found that it also forms a closed octameric ring in the crystal lattice. The SAM1-2 ring interactions are mediated by complementing "lock and key" hydrophobic grooves and inserts and electrostatic charges between the neighboring protomers. We have mutated several interacting SAM1-2 interfaces and measured how these mutations affect SARM1 apoptotic activity in cultured cells, and in this way identified critical oligomerization sites that facilitate cell death. These results highlight the importance of oligomerization for SARM1 function and reveal critical epitopes for future targeted drug development.

Solving the structure of Lgl2, a difficult blind test of unsupervised structure determination.

In the companion paper by Ufimtsev and Levitt [Ufimtsev IS, Levitt M (2019) Proc Natl Acad Sci USA, 10.1073/pnas.1821512116], we presented a method for unsupervised solution of protein crystal structures and demonstrated its utility by solving several test cases of known structure in the 2.9- to 3.45-Å resolution range. Here we apply this method to solve the crystal structure of a 966-amino acid construct of human lethal giant larvae protein (Lgl2) that resisted years of structure determination efforts, at 3.2-Å resolution. The structure was determined starting with a molecular replacement (MR) model identified by unsupervised refinement of a pool of 50 candidate MR models. This initial model had 2.8-Å RMSD from the solution. The solved structure was validated by comparison with a model subsequently derived from an alternative crystal form diffracting to higher resolution. This model could phase an anomalous difference Fourier map from an Hg derivative, and a single-wavelength anomalous dispersion phased density map made from these sites aligned with the refined structure.

Regulation of Streptococcus pneumoniae FtsZ assembly by divalent cations: paradoxical effects of Ca2+ on the nucleation and bundling of FtsZ polymers.

The assembly and disassembly of the FtsZ ring drives the division of bacteria cells, including Streptococcus pneumoniae, which causes pneumonia and meningitis. In contrast to FtsZ from other bacterial species, Streptococcus pneumoniae (Spn) FtsZ contains two tryptophan residues. Here, we demonstrate that the assembly and disassembly of Streptococcus pneumoniae FtsZ (SpnFtsZ) monomers can be monitored by the intrinsic tryptophan fluorescence of FtsZ. We found that the assembly of SpnFtsZ is closely associated with its GTPase activity. Guanosine 5'-[ß,γ-imido]triphosphate, a nonhydrolyzable analog of GTP, stabilized the FtsZ filaments without inducing their bundling. Using intrinsic tryptophan fluorescence, light scattering, and electron microscopy, we could differentiate the effects of divalent calcium and magnesium on the assembly of FtsZ. Though Mg2+ increased the stability of the FtsZ filaments, it could not prevent the disassembly of the filaments under conditions where GTP was limiting. Thus, our results indicate that Mg2+ primarily enhances the longitudinal assembly of FtsZ. Low concentrations of Ca2+ strongly promoted the bundling of FtsZ filaments and inhibited the disassembly of the filaments, suggesting that low concentrations of Ca2+ enhance the lateral interactions between the FtsZ filaments. Interestingly, Ca2+ delayed the nucleation process of FtsZ assembly, indicating that Ca2+ exerts paradoxical effects on the assembly of FtsZ. However, higher concentrations of Ca2+ did not enhance the bundling of FtsZ filaments. In addition, Ca2+ altered the secondary structure of FtsZ and increased the fluorescence of the FtsZ-1-anilinonaphthalene-8-sulfonic acid complex, indicating that Ca2+ induces conformational changes in FtsZ. The study provides an interesting insight into the assembly of SpnFtsZ and its regulation by divalent cations.

Movement dynamics of divisome proteins and PBP2x:FtsW in cells of Streptococcus pneumoniae.

Bacterial cell division and peptidoglycan (PG) synthesis are orchestrated by the coordinated dynamic movement of essential protein complexes. Recent studies show that bidirectional treadmilling of FtsZ filaments/bundles is tightly coupled to and limiting for both septal PG synthesis and septum closure in some bacteria, but not in others. Here we report the dynamics of FtsZ movement leading to septal and equatorial ring formation in the ovoid-shaped pathogen, Streptococcus pneumoniae Conventional and single-molecule total internal reflection fluorescence microscopy (TIRFm) showed that nascent rings of FtsZ and its anchoring and stabilizing proteins FtsA and EzrA move out from mature septal rings coincident with MapZ rings early in cell division. This mode of continuous nascent ring movement contrasts with a failsafe streaming mechanism of FtsZ/FtsA/EzrA observed in a ΔmapZ mutant and another Streptococcus species. This analysis also provides several parameters of FtsZ treadmilling in nascent and mature rings, including treadmilling velocity in wild-type cells and ftsZ(GTPase) mutants, lifetimes of FtsZ subunits in filaments and of entire FtsZ filaments/bundles, and the processivity length of treadmilling of FtsZ filament/bundles. In addition, we delineated the motion of the septal PBP2x transpeptidase and its FtsW glycosyl transferase-binding partner relative to FtsZ treadmilling in S. pneumoniae cells. Five lines of evidence support the conclusion that movement of the bPBP2x:FtsW complex in septa depends on PG synthesis and not on FtsZ treadmilling. Together, these results support a model in which FtsZ dynamics and associations organize and distribute septal PG synthesis, but do not control its rate in S. pneumoniae.

Escherichia coli ZipA Organizes FtsZ Polymers into Dynamic Ring-Like Protofilament Structures.

ZipA is an essential cell division protein in Escherichia coli Together with FtsA, ZipA tethers dynamic polymers of FtsZ to the cytoplasmic membrane, and these polymers are required to guide synthesis of the cell division septum. This dynamic behavior of FtsZ has been reconstituted on planar lipid surfaces in vitro, visible as GTP-dependent chiral vortices several hundred nanometers in diameter, when anchored by FtsA or when fused to an artificial membrane binding domain. However, these dynamics largely vanish when ZipA is used to tether FtsZ polymers to lipids at high surface densities. This, along with some in vitro studies in solution, has led to the prevailing notion that ZipA reduces FtsZ dynamics by enhancing bundling of FtsZ filaments. Here, we show that this is not the case. When lower, more physiological levels of the soluble, cytoplasmic domain of ZipA (sZipA) were attached to lipids, FtsZ assembled into highly dynamic vortices similar to those assembled with FtsA or other membrane anchors. Notably, at either high or low surface densities, ZipA did not stimulate lateral interactions between FtsZ protofilaments. We also used E. coli mutants that are either deficient or proficient in FtsZ bundling to provide evidence that ZipA does not directly promote bundling of FtsZ filaments in vivo Together, our results suggest that ZipA does not dampen FtsZ dynamics as previously thought, and instead may act as a passive membrane attachment for FtsZ filaments as they treadmill.IMPORTANCE Bacterial cells use a membrane-attached ring of proteins to mark and guide formation of a division septum at midcell that forms a wall separating the two daughter cells and allows cells to divide. The key protein in this ring is FtsZ, a homolog of tubulin that forms dynamic polymers. Here, we use electron microscopy and confocal fluorescence imaging to show that one of the proteins required to attach FtsZ polymers to the membrane during E. coli cell division, ZipA, can promote dynamic swirls of FtsZ on a lipid surface in vitro Importantly, these swirls are observed only when ZipA is present at low, physiologically relevant surface densities. Although ZipA has been thought to enhance bundling of FtsZ polymers, we find little evidence for bundling in vitro In addition, we present several lines of in vivo evidence indicating that ZipA does not act to directly bundle FtsZ polymers.

Probing the mechanistic principles of bacterial cell division with super-resolution microscopy.

Bacterial cell division takes place almost entirely below the diffraction limit of light microscopy, making super-resolution microscopy ideally suited to interrogating this process. I review how super-resolution microscopy has advanced our understanding of bacterial cell division. I discuss the mechanistic implications of these findings, propose physical models for cell division compatible with recent data, and discuss key outstanding questions and future research directions.

Mitochondrial anchors: Positioning mitochondria and more.

The shape and position of mitochondria are intimately connected to both mitochondrial and cellular function. Mitochondrial anchors play a central role in mitochondrial positioning by exerting spatial, temporal, and contextual control over the cellular position of the organelle. Investigations into the molecular mechanisms of mitochondrial anchoring are still in the early stages, and we are beginning to appreciate the number and variety of anchors that exist. From the insight gained thus far, it is clear that mitochondrial anchoring has functional and physiological consequences that extend beyond mitochondrial positioning to other critical cellular processes.