Biology subjects

McClintock, M. A.

Publications and source records attributed to McClintock, M. A..

5 recordsLinked to original sources

bioRxiv2026

FXR proteins and BICD2 cooperate to promote dynein-mediated RNP transport and stress granule assembly

Intracellular trafficking of ribonucleoprotein particles (RNPs) is a widespread strategy for regulating gene expression, yet how this process is orchestrated in mammalian systems remains unclear. Here, we show that Bicaudal D2 (BICD2), a partner of the microtubule motor dynein, is complexed with RNA-binding proteins (RBPs) in human cells. We also reveal that two of these RBPs - the Fragile X-related proteins FXR1 and FXR2 - bind directly to BICD2 and recruit mRNAs to dynein. Surprisingly, given its conventional designation as a cargo adaptor, BICD2 is not needed for dynein to interact with FXR1 and FXR2. Using in vitro reconstitution, we demonstrate that BICD2 instead activates dynein motility in concert with the FXR proteins. These data support a model in which BICD2 is recruited to pre-existing RNP-motor complexes, where it is licensed by the FXR proteins to activate processive movement. We further provide evidence that FXR proteins, BICD2 and dynein cooperate to promote rapid formation of stress granules, large RNPs that form in the cytoplasm in response to cellular insults. Collectively, our work reveals molecular mechanisms underlying RNP transport and stress granule assembly in human cells and indicates that cargo recruitment and motor activation are separable steps in dynein-mediated transport.

cell biology↗

bioRxiv2025

Cryo-EM structures of Egl-BicD-RNA complexes reveal how diverse mRNAs are selected for subcellular localization

Localization of mRNAs is a widespread mechanism for controlling where proteins operate in cells and underpins many fundamental processes, from embryonic patterning to synaptic plasticity. This spatial control is mediated by RNA-binding proteins (RBPs) that interact with localization signals within target mRNAs. However, these signals lack overt sequence or structural patterns, even when targeted by the same protein, raising the question of how specificity is achieved. Here, we investigate this question using the Drosophila RBP Egalitarian (Egl), which couples developmentally important mRNAs to microtubule-based transport via Bicaudal D (BicD) and the dynein motor protein. We present cryo-EM structures of Egl- BicD in complex with six different RNAs. Egl uses multiple non-canonical double-stranded RNA-binding domains to cooperatively form a recognition pocket around a localization signal. Despite substantial variation in length and sequence, each signal adopts a bent stem-loop conformation that, together with base-pair identities at two defined sites, drives Egl binding. We further demonstrate that Egl dimers couple RNA binding to transport initiation through coincident detection of two RNA elements within the same transcript. Collectively, we show that diverse localizing mRNAs are recognized through a combination of shape, positional sequence features, and number of structured RNA elements. This work provides a framework for understanding how other RBPs engage highly variable mRNAs.

cell biology↗

bioRxiv2023

A force-sensitive mutation reveals a spindle assembly checkpoint-independent role for dynein in anaphase progression

The cytoplasmic dynein-1 (dynein) motor organizes cells by shaping microtubule networks and moving a large variety of cargoes along them. However, dyneins diverse roles complicate in vivo studies of its functions significantly. To address this issue, we have used gene editing to generate a series of missense mutations in Drosophila Dynein heavy chain (Dhc). We find that mutations associated with human neurological disease cause a range of defects in larval and adult flies, including impaired cargo trafficking in neurons. We also describe a novel mutation in the microtubule-binding domain (MTBD) of Dhc that, remarkably, causes metaphase arrest of mitotic spindles in the embryo but does not impair other dynein-dependent processes. We demonstrate that the mitotic arrest is independent of dyneins well-established roles in silencing the spindle assembly checkpoint. In vitro reconstitution and optical trapping assays reveal that the mutation only impairs the performance of dynein under load. In silico all-atom molecular dynamics simulations show that this effect correlates with increased flexibility of the MTBD, as well as an altered orientation of the stalk domain, with respect to the microtubule. Collectively, our data point to a novel role of dynein in anaphase progression that depends on the motor operating in a specific load regime. More broadly, our work illustrates how cytoskeletal transport processes can be dissected in vivo by manipulating mechanical properties of motors.

cell biology↗

bioRxiv2022

Tropomyosin 1-I/C co-ordinates kinesin-1 and dynein motors during oskar mRNA transport

Dynein and kinesin motors mediate long-range intracellular transport, translocating towards microtubule minus and plus ends, respectively. Cargoes often undergo bidirectional transport by binding to both motors simultaneously. However, it is not known how motor activities are coordinated in such circumstances. In Drosophila, sequential activities of the dynein-dynactin-BicD-Egalitarian (DDBE) complex and of kinesin-1 deliver oskar mRNA from nurse cells to the oocyte, and within the oocyte to the posterior pole. Here, through in vitro reconstitution, we show that Tm1-I/C, a Tropomyosin-1 isoform, links kinesin-1 in an inactive state to DDBE-associated oskar mRNA. NMR spectroscopy, small-angle X-ray scattering and structural modeling indicate that Tm1-I/C suppresses kinesin-1 activity by stabilizing its autoinhibited conformation, thus preventing a tug-of-war between the opposite polarity motors until kinesin-1 is activated in the oocyte. Our work reveals a novel strategy ensuring sequential activity of microtubule motors.

biochemistry↗

bioRxiv2018

RNA-directed activation of cytoplasmic dynein-1 in reconstituted transport RNPs

Polarised mRNA transport is a prevalent mechanism for spatial control of protein synthesis. However, the composition of transported ribonucleoprotein particles (RNPs) and the regulation of their movement are poorly understood. We have reconstituted microtubule minus end-directed transport of mRNAs using purified components. A Bicaudal-D (BicD) adaptor protein and the RNA-binding protein Egalitarian (Egl) are sufficient for long-distance mRNA transport by the dynein motor and its accessory complex dynactin, thus defining a minimal transport-competent RNP. Unexpectedly, the RNA is required for robust activation of dynein motility. We show that a cis-acting RNA localisation signal stabilises the interaction of Egl with BicD, which licenses the latter protein to recruit dynein and dynactin. Our data support a model for BicD activation based on RNA-induced occupancy of two Egl-binding sites on the BicD dimer. Scaffolding of adaptor protein assemblies by cargoes is an attractive mechanism for regulating intracellular transport.

cell biology↗