Microtubule Image Processing in CryoSPARC Pipeline - wieczoreklab/MiCSPARC

Sarkar et al. show that the CLASP protein Orbit stabilizes non-centrosomal microtubule plus ends in vivo, suppressing catastrophe and enabling persistent polymerization-based pushing forces. This stab...

The cellular distribution of mitochondria in response to stress and local energy needs is governed by the relative activities of the microtubule-based molecular motors kinesin and dynein. The mechanis...

The cellular distribution of mitochondria in response to stress and local energy needs is governed by the relative activities of the microtubule-based molecular motors kinesin and dynein. The mechanis...

The manchette is a transient microtubule (MT)-based structure that is vital for the correct shaping of sperm during spermiogenesis. Throughout spermiogenesis, the manchette retains structural integrity for several days, raising the question of how its MTs are regulated. Here, using cryo-electron tomography of manchettes isolated from rat testes, we find that manchette MT ends are structurally diverse. We show that the MT-binding protein CLASP2 is present throughout the manchette and likely regulates both MT ends. Using cryo-electron microscopy single particle analysis and super-resolution microscopy, we reveal that SPACA9 and MNMIP1 (SH3D21) bind to the seam of manchette MTs from the luminal side. SPACA9 binds to both α- and β-tubulin of protofilament 1 but does not interact directly with protofilament 13, while MNMIP1 binds directly to protofilament 13. MNMIP1 further extends and threads through the MT lattice at the seam. Our study reveals a novel seam MT inner protein complex with a unique binding mode, providing a plausible explanation for MT regulation that maintains manchette integrity over an extended period.

Primary cilia are microtubule-based organelles that mediate cellular responses to environmental cues. Although cilia disassemble and reassemble upon cell cycle entry and exit, respectively, in dividin...

Tau aggregation, a hallmark of Alzheimer’s disease, disrupts neuron structure. Aging weakens chaperone defenses like Hsp90. This study designs β-Hsp90, a small peptide mimicking Hsp90, to prevent Tau aggregation, offering promise for new amyloid disease drugs.

We developed an experimental platform in which CFE continuously synthesizes MT-associated proteins directly within a dense MT network, enabling the active-matter constituents themselves, including motors, cross-linkers and regulatory proteins, to be produced during the experiment rather than prescribed at assembly (Fig. 1a). Guanosine-5′-(α,β-methyleno)triphosphate (GMPCPP)-stabilized MTs were combined with a CFE transcription-translation system and sedimented into a mechanically quiescent planar sheet within a 3.3 mm × 3.3 mm well (1.2 mm liquid column), open to air for oxygen availability11,23,27 (Supplementary Fig. 1). Monomeric kinesin motors were continuously expressed from a 3 nM T7-driven plasmid encoding a codon-optimized kinesin gene (Supplementary Table 1). To sustain long-term coupling between gene expression and active matter, the CFE reaction was optimized to promote depletion-mediated MT bundling and minimize competition between protein synthesis and motor activity over shared energetic resources (Methods). At early times, t < 2 h, when kinesin concentrations remained low, the MT sheet behaved as a passive, load-bearing material with negligible internal motion (Fig. 1b and Supplementary Video 1). At longer times, t > 2 h, kinesin accumulated and promoted the formation of multimeric clusters. The emergence of tetrameric clusters, the minimal motor assemblies capable of generating flow, enabled crosslinking and extensile sliding...

Cryo-EM reveals the structural basis of kinesin-1 autoinhibition and its activation by MAP7D3.

The manchette is a transient microtubule (MT)-based structure that is vital for the correct shaping of sperm during spermiogenesis. Throughout spermiogenesis, the manchette retains structural integrit...

Kinesin-1 is a dynamic heterotetrameric assembly of two heavy and two light chains (KHC and KLC) that mediates microtubule-based intracellular transport of many different cargoes. The complex adopts a...