Biology subjects

Mohd Rafiq, N.

Publications and source records attributed to Mohd Rafiq, N..

6 recordsLinked to original sources

bioRxiv2026

Curved microtubule regions mark sites of lattice compaction in cells and neurons

Angstrom-scale changes in microtubule (MT) lattice spacing regulate the selective recruitment of MT-associated proteins, yet how these structural states operate in cells remains poorly understood. Here, we show that MT lattice expansion, induced by protein-based expanders or microtubule-stabilizing agents such as Taxol and epothilone D, drives the relocalization of compact lattice-binding proteins, including tau, doublecortin (DCX), and the C1 domain-containing signaling protein GEF-H1, into highly curved MT-associated domains, whereas the compaction-inducing agent laulimalide suppresses this response. In contrast, the tumor suppressor RASSF1A preferentially associates with expanded lattice states, revealing differential lattice sensitivity among closely related C1 domain-containing proteins. These short, curved assemblies are enriched at MT intersections and discrete MT segments, revealing spatially heterogeneous lattice states within individual microtubules. At substoichiometric levels, compact lattice-binding proteins behave as both MT compactors and curvature sensors. Changes in osmotic pressure selectively promote dissociation of compact lattice-binding proteins, whereas expanded lattice-binding proteins remain largely unaffected. Using curved filament formation as an in-cellulo readout of compact lattice regions, we identify widespread lattice-state sensitivity across diverse MT-associated and signaling proteins. Finally, we show that these principles extend to neurons, where somatic, but not axonal, tau exhibits sensitivity to lattice expansion despite the expanded lattice architecture of distal axonal microtubules, suggesting additional neuron-specific regulation of lattice accessibility. Together, our findings identify the MT lattice as a dynamic mechanochemical platform whose nanoscale structural states spatially organize protein recruitment and signaling in cells and neurons.

cell biology↗

bioRxiv2026

Mechanical tension expands the microtubule lattice stepwise and modulates kinesin-1 binding in an isoform-dependent manner

Recent work has shown that the microtubule lattice possesses remarkable structural plasticity, with its conformation modulated by microtubule-associated proteins and motor proteins. However, how this plasticity responds to mechanical forces remains poorly understood. Here, we developed optical tweezers and fluorescence microscopy assays to measure the effect of tensile forces on single microtubules. Quantum dot decoration enabled nanometre-precision measurement of lattice distortions of [~]0.33% under a change of mean tensile force of [<]{Delta}F[>] = 10.6 pN, within the range Fmin = 1.29 pN to Fmax = 22.3 pN -- comparable to forces from one to three kinesin-1 motors. Within this force range, the binding rate of kinesin-1 isoform KIF5B decreased reversibly within seconds by [~]20% and the dissociation rate increased by [~]10%, reducing mean run length, that in extreme cases decreased by up to 46%. Substantial heterogeneity was also observed along individual microtubules, where distinct lattice regions responded differently to applied force, implying that lattice expansion is not always uniform. Consistent heterogeneity was observed in cells, where MAPs with competing conformational preferences assembled in non-overlapping patches along the same microtubule. A cooperatively-switching lattice Ising model based on tubulin conformational bistability, supported by dynamics simulations, quantitatively reproduces these observations with a critical switching force Fc = 8.5 pN, similar to established mechanosensory proteins such as talin and E-catenin. Strikingly, no significant effects were observed for KIF5C, revealing a kinesin isoform-dependent mechanoresponse. Together, these findings establish microtubules as mechanochemical signal transducers, converting mechanical forces into biochemical signals with the speed, spatial precision and sensitivity required for rapid cellular responses. Significance StatementMicrotubules have been implicated as mechanotransducers in both mammalian and plant cells, yet a physical characterization of how mechanical forces are sensed and transduced into biochemical signals has been lacking. The present study demonstrates that modest tensile forces of less than 20 pN are sufficient to expand cooperatively the microtubule lattice by [~]0.3%, which in turn modulates its biochemical interactions with kinesin-1 in an isoform-dependent manner, selectively affecting KIF5B motor activity but not KIF5C. Strikingly, this mechanotransduction occurs on a timescale of seconds, implying that microtubules are highly efficient conduits for propagating mechanical information across the cell body. These findings establish microtubules as bona fide mechanochemical signal transducers with the speed and sensitivity required for rapid cellular responses.

biophysics↗

bioRxiv2026

Dopamine vesicles are specified by mechanisms overriding canonical synaptic vesicle size constraints

Dopaminergic neurons contain distinct synaptic vesicle (SV) populations that segregate dopamine from glutamate, but the molecular basis of this segregation remains unclear. Using an established in vitro SV reconstitution system in fibroblasts, we show that synaptogyrin family proteins--synaptogyrin 1, synaptogyrin 2 and synaptogyrin 3--but not synaptophysin, selectively associate with VMAT2-positive dopamine vesicles in the presence of synapsin. Despite the known role of synaptophysin and synaptogyrin family proteins in generating small SVs (40-50 nm), VMAT2 vesicles retain a larger size (50-80 nm) when co-expressed with synaptogyrins, while remaining segregated from synaptophysin-containing vesicles. SV2 family proteins (SV2A, SV2B, and SV2C) localize to both VMAT2- and synaptophysin-positive vesicle populations, with SV2C preferentially enriched on VMAT2-positive dopamine vesicles. Consistent with the selective dystrophy of SV2C/VMAT2/DAT-positive dopaminergic terminals in parkinsonism-associated SJ1RQKI mice, iPSC-derived dopaminergic neurons carrying the same mutation accumulate small and enlarged SV-like organelles within autophagosomes, indicating defective SV clearance. Together, our findings suggest that dopamine vesicles are regulated by distinct trafficking mechanisms that override canonical SV size control, providing insight into the selective vulnerability of dopaminergic terminals in Parkinsons disease.

neuroscience↗

bioRxiv2023

Parkinsonism Sac domain mutation in Synaptojanin-1 affects ciliary properties in iPSC-derived dopaminergic neurons

Synaptojanin-1 (SJ1) is a major neuronal-enriched PI(4,5)P2 4- and 5-phosphatase implicated in the shedding of endocytic factors during endocytosis. A mutation (R258Q) that impairs selectively its 4-phosphatase activity causes Parkinsonism in humans and neurological defects in mice (SJ1RQKI mice). Studies of these mice showed, besides an abnormal assembly state of endocytic factors at synapses, the presence of dystrophic nerve terminals selectively in a subset of nigro-striatal dopamine (DA)-ergic axons, suggesting a special lability of DA neurons to the impairment of SJ1 function. Here we have further investigated the impact of SJ1 on DA neurons using iPSC-derived SJ1 KO and SJ1RQKI DA neurons and their isogenic controls. In addition to the expected enhanced clustering of endocytic factors in nerve terminals, we observed in both SJ1 mutant neuronal lines increased cilia length. Further analysis of cilia of SJ1RQDA neurons revealed abnormal accumulation of the Ca2+ channel Cav1.3 and of ubiquitin chains, suggesting an impaired clearing of proteins from cilia which may result from an endocytic defect at the ciliary base, where a focal concentration of SJ1 was observed. We suggest that SJ1 may contribute to the control of ciliary protein dynamics in DA neurons, with implications on cilia-mediated signaling.

neuroscience↗

bioRxiv2018

Forces and constraints controlling podosome assembly and disassembly

Podosomes are a singular category of integrin-mediated adhesions important in the processes of cell migration, matrix degradation, and cancer cell invasion. Despite a wealth of biochemical studies, the effects of mechanical forces on podosome integrity and dynamics are poorly understood. Here, we show that podosomes are highly sensitive to two groups of physical factors. First, we describe the process of podosome disassembly induced by activation of myosin-IIA filament assembly. Next, we find that podosome integrity and dynamics depends upon membrane tension and can be experimentally perturbed by osmotic swelling and deoxycholate treatment. We have also found that podosomes can be disrupted in a reversible manner by single or cyclic radial stretching of the substratum. We show that disruption of podosomes induced by osmotic swelling is independent of myosin-II filaments. Inhibition of the membrane sculpting protein, dynamin-II, but not clathrin, resulted in activation of myosin-IIA filament formation and disruption of podosomes. The effect of dynamin-II inhibition on podosomes was however independent of myosin-II filaments. Moreover, formation of organized arrays of podosomes in response to microtopographic cues (the ridges with triangular profile) was not accompanied by reorganization of myosin-II filaments. Thus, mechanical elements such as myosin-II filaments and factors affecting membrane tension/sculpting independently modulate podosome formation and dynamics, underlying a versatile response of these adhesion structures to intracellular and extracellular cues.

cell biology↗

bioRxiv2017

Microtubule regulation of integrin-based adhesions is mediated by myosin-IIA

The interrelationship between microtubules and the actin cytoskeleton in mechanoregulation of integrin-mediated adhesions is poorly understood. Here, we show that the effects of microtubules on two major types of cell-matrix adhesions, focal adhesions and podosomes, are mediated by KANK family proteins connecting the adhesion protein talin with microtubule tips. Both total microtubule disruption and microtubule uncoupling from adhesions by manipulations with KANKs trigger a massive assembly of myosin-IIA filaments. Myosin-IIA filaments, augmenting the focal adhesions and disrupting the podosomes, are indispensable effectors in the microtubule-dependent regulation of integrin-mediated adhesions. Myosin-IIA filament assembly depends on Rho activation by the RhoGEF, GEF-H1, which is trapped by microtubules when they are connected with integrin-mediated adhesions via KANK proteins but released after their disconnection. Thus, microtubule capturing by integrin-mediated adhesions modulates the GEF-H1-dependent effect of microtubules on the myosin-IIA filaments. Subsequent actomyosin reorganization then remodels the focal adhesions and podosomes, closing the regulatory loop.

cell biology↗