scholarly journals ATP-dependent force generation and membrane scission by ESCRT-III and Vps4

2018 ◽  
Author(s):  
Johannes Schöneberg ◽  
Shannon Yan ◽  
Maurizio Righini ◽  
Mark Remec Pavlin ◽  
Il-Hyung Lee ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Atp Release ◽  
Force Production ◽  
Force Generation ◽  
Membrane Insertion ◽  
Giant Vesicles ◽  
Membrane Nanotubes ◽  
Membrane Scission ◽  
Correct Topology

AbstractThe ESCRTs catalyze reverse-topology scission from the inner face of membrane necks in HIV budding, multivesicular endosome biogenesis, cytokinesis, and other pathways. We encapsulated a minimal ESCRT module consisting of ESCRT-III subunits Snf7, Vps24, and Vps2, and the AAA+ ATPase Vps4 such that membrane nanotubes reflecting the correct topology of scission could be pulled from giant vesicles. Upon ATP release by photo-uncaging, this system was capable of generating forces within the nanotubes in a manner dependent upon Vps4 catalytic activity, Vps4 coupling to the ESCRT-III proteins, and membrane insertion by Snf7. At physiological concentrations, single scission events were observed that correlated with forces of ~6 pN, verifying predictions that ESCRTs are capable of exerting forces on membranes. Imaging of scission with subsecond resolution revealed Snf7 puncta at the sites of membrane cutting, directly verifying longstanding predictions for the ESCRT scission mechanism.One Sentence SummaryESCRT-III and Vps4 were reconstituted from within the interior of nanotubes pulled from giant vesicles, revealing that this machinery couples ATP-dependent force production for membrane scission.

scholarly journals ATP-dependent force generation and membrane scission by ESCRT-III and Vps4

Science ◽  
2018 ◽  
Vol 362 (6421) ◽  
pp. 1423-1428 ◽  
Author(s):  
Johannes Schöneberg ◽  
Mark Remec Pavlin ◽  
Shannon Yan ◽  
Maurizio Righini ◽  
Il-Hyung Lee ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Adenosine Triphosphatase ◽  
Atp Release ◽  
Force Generation ◽  
Giant Vesicles ◽  
Endosomal Sorting ◽  
Aaa Atpase ◽  
Membrane Nanotubes ◽  
Membrane Scission ◽  
Correct Topology

The endosomal sorting complexes required for transport (ESCRTs) catalyze reverse-topology scission from the inner face of membrane necks in HIV budding, multivesicular endosome biogenesis, cytokinesis, and other pathways. We encapsulated ESCRT-III subunits Snf7, Vps24, and Vps2 and the AAA+ ATPase (adenosine triphosphatase) Vps4 in giant vesicles from which membrane nanotubes reflecting the correct topology of scission could be pulled. Upon ATP release by photo-uncaging, this system generated forces within the nanotubes that led to membrane scission in a manner dependent upon Vps4 catalytic activity and Vps4 coupling to the ESCRT-III proteins. Imaging of scission revealed Snf7 and Vps4 puncta within nanotubes whose presence followed ATP release, correlated with force generation and nanotube constriction, and preceded scission. These observations directly verify long-standing predictions that ATP-hydrolyzing assemblies of ESCRT-III and Vps4 sever membranes.

scholarly journals Insertion of plastidic β-barrel proteins into the outer envelopes of plastids involves an intermembrane space intermediate formed with Toc75-V/OEP80

2021 ◽  
Author(s):  
Lucia E Gross ◽  
Anna Klinger ◽  
Nicole Spies ◽  
Theresa Ernst ◽  
Nadine Flinner ◽  
...  
Keyword(s):  
Membrane Proteins ◽  
Protein Import ◽  
Membrane Insertion ◽  
Intermembrane Space ◽  
Envelope Membrane ◽  
Outer Envelope ◽  
Pea Proteins ◽  
Outer Envelope Membrane ◽  
Correct Topology ◽  
Shed Light

Abstract The insertion of organellar membrane proteins with the correct topology requires the following: First, the proteins must contain topogenic signals for translocation across and insertion into the membrane. Second, proteinaceous complexes in the cytoplasm, membrane, and lumen of organelles are required to drive this process. Many complexes required for the intracellular distribution of membrane proteins have been described, but the signals and components required for the insertion of plastidic β-barrel-type proteins into the outer membrane are largely unknown. The discovery of common principles is difficult, as only a few plastidic β-barrel proteins exist. Here, we provide evidence that the plastidic outer envelope β-barrel proteins OEP21, OEP24, and OEP37 from pea (Pisum sativum) and Arabidopsis thaliana contain information defining the topology of the protein. The information required for translocation of pea proteins across the outer envelope membrane is present within the six N-terminal β-strands. This process requires the action of TOC (translocon of the outer chloroplast membrane). After translocation into the intermembrane space, β-barrel proteins interact with TOC75-V, as exemplified by OEP37 and P39, and are integrated into the membrane. The membrane insertion of plastidic β-barrel proteins is affected by mutation of the last β-strand, suggesting that this strand contributes to the insertion signal. These findings shed light on the elements and complexes involved in plastidic β-barrel protein import.

Neuromuscular drive and force production are not altered during bilateral contractions

1998 ◽  
Vol 84 (1) ◽  
pp. 200-206 ◽  
Author(s):  
J. M. Jakobi ◽  
E. Cafarelli
Keyword(s):  
Motor Unit ◽  
Force Production ◽  
Young Male ◽  
Neuromuscular Control ◽  
Force Generation ◽  
Isometric Contractions ◽  
Firing Rates ◽  
Significant Limitation ◽  
Motor Unit Firing ◽  
Male Subjects

Jakobi, J. M., and E. Cafarelli. Neuromuscular drive and force production are not altered during bilateral contractions. J. Appl. Physiol. 84(1): 200–206, 1998.—Several investigators have studied the deficit in maximal voluntary force that is said to occur when bilateral muscle groups contract simultaneously. A true bilateral deficit (BLD) would suggest a significant limitation of neuromuscular control; however, some of the data from studies in the literature are equivocal. Our purpose was to determine whether there is a BLD in the knee extensors of untrained young male subjects during isometric contractions and whether this deficit is associated with a decreased activation of the quadriceps, increased activation of the antagonist muscle, or an alteration in motor unit firing rates. Twenty subjects performed unilateral (UL) and bilateral (BL) isometric knee extensions at 25, 50, 75, and 100% maximal voluntary contraction. Total UL and BL force (Δ3%) and maximal rate of force generation (Δ2.5%) were not significantly different. Total UL and BL maximal vastus lateralis electromyographic activity (EMG; 2.7 ± 0.28 vs. 2.6 ± 0.24 mV) and coactivation (0.17 ± 0.02 vs. 0.20 ± 0.02 mV) were also not different. Similarly, the ratio of force to EMG during submaximal UL and BL contractions was not different. Analysis of force production by each leg in UL and BL conditions showed no differences in force, rate of force generation, EMG, motor unit firing rates, and coactivation. Finally, assessment of quadriceps activity with the twitch interpolation technique indicated no differences in the degree of voluntary muscle activation (UL: 93.6 ± 2.51 Hz, BL: 90.1 ± 2.43 Hz). These results provide no evidence of a significant limitation in neuromuscular control between BL and UL isometric contractions of the knee extensor muscles in young male subjects.

scholarly journals Neck linker docking is critical for Kinesin-1 force generation in cells but at a cost to motor speed and processivity

eLife ◽  
2019 ◽  
Vol 8 ◽  
Author(s):  
Breane G Budaitis ◽  
Shashank Jariwala ◽  
Dana N Reinemann ◽  
Kristin I Schimert ◽  
Guido Scarabelli ◽  
...  
Keyword(s):  
Optical Trap ◽  
Force Production ◽  
Force Generation ◽  
Motor Speed ◽  
Force Output ◽  
Mechanical Element ◽  
Membrane Bound ◽  
Dynamics Simulations ◽  
Run Lengths ◽  
In Cells

Kinesin force generation involves ATP-induced docking of the neck linker (NL) along the motor core. However, the roles of the proposed steps of NL docking, cover-neck bundle (CNB) and asparagine latch (N-latch) formation, during force generation are unclear. Furthermore, the necessity of NL docking for transport of membrane-bound cargo in cells has not been tested. We generated kinesin-1 motors impaired in CNB and/or N-latch formation based on molecular dynamics simulations. The mutant motors displayed reduced force output and inability to stall in optical trap assays but exhibited increased speeds, run lengths, and landing rates under unloaded conditions. NL docking thus enhances force production but at a cost to speed and processivity. In cells, teams of mutant motors were hindered in their ability to drive transport of Golgi elements (high-load cargo) but not peroxisomes (low-load cargo). These results demonstrate that the NL serves as a mechanical element for kinesin-1 transport under physiological conditions.

scholarly journals Membrane Nanotubes Increase the Robustness of Giant Vesicles

ACS Nano ◽  
2018 ◽  
Vol 12 (5) ◽  
pp. 4478-4485 ◽  
Author(s):  
Tripta Bhatia ◽  
Jaime Agudo-Canalejo ◽  
Rumiana Dimova ◽  
Reinhard Lipowsky
Keyword(s):  
Giant Vesicles ◽  
Membrane Nanotubes

scholarly journals Contributions of Stretch Activation to Length-dependent Contraction in Murine Myocardium

2006 ◽  
Vol 128 (4) ◽  
pp. 461-471 ◽  
Author(s):  
Julian E. Stelzer ◽  
Richard L. Moss
Keyword(s):  
Force Production ◽  
Muscle Length ◽  
Intrinsic Property ◽  
Work Capacity ◽  
Force Generation ◽  
Stretch Activation ◽  
Length Dependence ◽  
Systolic Ejection ◽  
End Diastolic Volume

The steep relationship between systolic force production and end diastolic volume (Frank-Starling relationship) in myocardium is a potentially important mechanism by which the work capacity of the heart varies on a beat-to-beat basis, but the molecular basis for the effects of myocardial fiber length on cardiac work are still not well understood. Recent studies have suggested that an intrinsic property of myocardium, stretch activation, contributes to force generation during systolic ejection in myocardium. To examine the role of stretch activation in length dependence of activation we recorded the force responses of murine skinned myocardium to sudden stretches of 1% of muscle length at both short (1.90 μm) and long (2.25 μm) sarcomere lengths (SL). Maximal Ca2+-activated force and Ca2+ sensitivity of force were greater at longer SL, such that more force was produced at a given Ca2+ concentration. Sudden stretch of myocardium during an otherwise isometric contraction resulted in a concomitant increase in force that quickly decayed to a minimum and was followed by a delayed development of force, i.e., stretch activation, to levels greater than prestretch force. At both maximal and submaximal activations, increased SL significantly reduced the initial rate of force decay following stretch; at submaximal activations (but not at maximal) the rate of delayed force development was accelerated. This combination of mechanical effects of increased SL would be expected to increase force generation during systolic ejection in vivo and prolong the period of ejection. These results suggest that sarcomere length dependence of stretch activation contributes to the steepness of the Frank-Starling relationship in living myocardium.

scholarly journals Formation of helical membrane tubes around microtubules by single-headed kinesin KIF1A

2015 ◽  
Vol 6 (1) ◽  
Author(s):  
David Oriola ◽  
Sophie Roth ◽  
Marileen Dogterom ◽  
Jaume Casademunt
Keyword(s):  
Motor Performance ◽  
Vesicular Transport ◽  
Force Generation ◽  
Theoretical Studies ◽  
Giant Vesicles ◽  
Traffic Conditions ◽  
Self Organized ◽  
Mechanochemical Cycle ◽  
Axonal Trafficking

Abstract The kinesin-3 motor KIF1A is in charge of vesicular transport in neuronal axons. Its single-headed form is known to be very inefficient due to the presence of a diffusive state in the mechanochemical cycle. However, recent theoretical studies have suggested that these motors could largely enhance force generation by working in teams. Here we test this prediction by challenging single-headed KIF1A to extract membrane tubes from giant vesicles along microtubule filaments in a minimal in vitro system. Remarkably, not only KIF1A motors are able to extract tubes but they feature a novel phenomenon: tubes are wound around microtubules forming tubular helices. This finding reveals an unforeseen combination of cooperative force generation and self-organized manoeuvreing capability, suggesting that the diffusive state may be a key ingredient for collective motor performance under demanding traffic conditions. Hence, we conclude that KIF1A is a genuinely cooperative motor, possibly explaining its specificity to axonal trafficking.

Mutability of Bifunctional Thigh Muscle Activity in Pedaling due to Contralateral Leg Force Generation

2002 ◽  
Vol 88 (3) ◽  
pp. 1308-1317 ◽  
Author(s):  
S. A. Kautz ◽  
D. A. Brown ◽  
H.F.M. Van der Loos ◽  
F. E. Zajac
Keyword(s):  
Muscle Activity ◽  
Isometric Force ◽  
Force Production ◽  
Rectus Femoris ◽  
Force Generation ◽  
Thigh Muscle ◽  
Work Output ◽  
Thigh Muscles ◽  
Sensorimotor Activity ◽  
Bifunctional Muscle

Locomotion requires uninterrupted transitions between limb extension and flexion. The role of contralateral sensorimotor signals in executing smooth transitions is little understood even though their participation is crucial to bipedal walking. However, elucidating neural interlimb coordinating mechanisms in human walking is difficult because changes to contralateral sensorimotor activity also affect the ipsilateral mechanics. Pedaling, conversely, is ideal for studying bilateral coordination because ipsilateral mechanics can be independently controlled. In pedaling, the anterior and posterior bifunctional thigh muscles develop needed anterior and posterior crank forces, respectively, to dominate the flexion-to-extension and extension-to-flexion transitions. We hypothesized that contralateral sensorimotor activity substantially contributes to the appropriate activation of these bifunctional muscles during the limb transitions. Bilateral pedal forces and surface electromyograms (EMGs) from four thigh muscles were collected from 15 subjects who pedaled with their right leg against a right-crank servomotor, which emulated the mechanical load experienced in conventional two-legged coupled-crank pedaling. In one pedaling session, the contralateral (left) leg pseudo-pedaled (i.e., EMG activity and pedal forces were pedaling-like, but pedal force was not allowed to affect crank rotation). In other sessions, the mechanically decoupled contralateral leg was first relaxed and then produced rhythmic isometric force trajectories during either leg flexion or one of the two limb transitions of the pedaling leg. With contralateral force production in the extension-to-flexion transition (predominantly by the hamstrings), rectus femoris activity and work output increased in the pedaling leg during its flexion-to-extension transition, which occurs simultaneously with contralateral extension-to-flexion in conventional pedaling. Similarly, with contralateral force production in the other transition (i.e., flexion-to-extension; predominantly by rectus femoris), hamstrings activity and work output increased in the pedaling leg during its extension-to-flexion transition. Therefore rhythmic isometric force generation in the contralateral leg supported the ongoing bifunctional muscle activity and resulting work output in the pedaling leg. The results suggest that neural interlimb coordinating mechanisms fine-tune bifunctional muscle activity in rhythmic lower-limb tasks to ensure limb flexion/extension transitions are executed successfully.

Dissecting the role of the myofilament in diaphragm dysfunction during the development of heart failure in mice

2016 ◽  
Vol 310 (5) ◽  
pp. H572-H586 ◽  
Author(s):  
Todd E. Gillis ◽  
Jordan M. Klaiman ◽  
Andrew Foster ◽  
Mathew J. Platt ◽  
Jason S. Huber ◽  
...  
Keyword(s):  
Heart Failure ◽  
Troponin T ◽  
Isometric Force ◽  
Force Production ◽  
Force Generation ◽  
Myosin Isoforms ◽  
Diaphragmatic Dysfunction ◽  
Diaphragmatic Function ◽  
Function Models

Dyspnea and reduced exercise capacity, caused, in part, by respiratory muscle dysfunction, are common symptoms in patients with heart failure (HF). However, the etiology of diaphragmatic dysfunction has not been identified. To investigate the effects of HF on diaphragmatic function, models of HF were surgically induced in CD-1 mice by transverse aortic constriction (TAC) and acute myocardial infarction (AMI), respectively. Assessment of myocardial function, isolated diaphragmatic strip function, myofilament force-pCa relationship, and phosphorylation status of myofilament proteins was performed at either 2 or 18 wk postsurgery. Echocardiography and invasive hemodynamics revealed development of HF by 18 wk postsurgery in both models. In vitro diaphragmatic force production was preserved in all groups while morphometric analysis revealed diaphragmatic atrophy and fibrosis in 18 wk TAC and AMI groups. Isometric force-pCa measurements of myofilament preparations revealed reduced Ca2+ sensitivity of force generation and force generation at half-maximum and maximum Ca2+ activation in 18 wk TAC. The rate of force redevelopment ( ktr) was reduced in all HF groups at high levels of Ca2+ activation. Finally, there were significant changes in the myofilament phosphorylation status of the 18 wk TAC group. This includes a decrease in the phosphorylation of troponin T, desmin, myosin light chain (MLC) 1, and MLC 2 as well as a shift in myosin isoforms. These results indicate that there are multiple changes in diaphragmatic myofilament function, which are specific to the type and stage of HF and occur before overt impairment of in vitro force production.

scholarly journals PFN2 and NAA80 cooperate to efficiently acetylate the N-terminus of actin

2020 ◽  
Author(s):  
Rasmus Ree ◽  
Laura Kind ◽  
Anna Kaziales ◽  
Sylvia Varland ◽  
Minglu Dai ◽  
...  
Keyword(s):  
Catalytic Activity ◽  
Actin Cytoskeleton ◽  
Actin Filament ◽  
Cell Shape ◽  
Specific Role ◽  
Force Generation ◽  
Filament Formation ◽  
Modus Operandi ◽  
N Terminus

AbstractThe actin cytoskeleton is of profound importance to cell shape, division, and intracellular force generation. Profilins bind to globular (G-)actin and regulate actin filament formation. Although profilins are well-established actin regulators, the distinct roles of the dominant profilin, profilin 1 (PFN1), versus the less abundant profilin 2 (PFN2) remain enigmatic. Here, we define a specific role for PFN2 as a stable interactor and regulator of the actin N-terminal acetyltransferase NAA80. PFN2 binding increases the intrinsic catalytic activity of NAA80. Furthermore, binding of PFN2 to NAA80 via its proline-rich loop promotes binding between the globular domains of actin and NAA80, and thus acetylation of actin. The majority of NAA80 is stably bound to PFN2, and we propose that this complex acetylates G-actin before it is incorporated into filaments. In conclusion, we reveal a functionally specific role of PFN2, and establish the modus operandi for NAA80-mediated actin N-terminal acetylation. Data are available via ProteomeXchange with identifier PXD020188.

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