Biology subjects

Christensen, J. R.

Publications and source records attributed to Christensen, J. R..

8 recordsLinked to original sources

bioRxiv2025

Acyl-coA binding protein AcbdA regulates peroxisome hitchhiking on early endosomes

Motor-driven transport on microtubules is critical for distributing organelles throughout the cell. Most commonly, organelle movement is mediated by cargo adaptors, proteins on the surface of an organelle that directly recruit microtubule-based motors. An alternative mechanism called hitchhiking was recently discovered: some organelles move, not by recruiting the motors directly, but instead by using membrane contact sites to attach to motor-driven vesicles and hitchhike along microtubules. Organelle hitchhiking is observed across fungi and animals. In filamentous fungi, nearly all peroxisomes move by hitchhiking on early endosomes (EEs). In the fungus Aspergillus nidulans, EE-associated linker proteins PxdA and DipA are critical for establishing EE-peroxisome membrane contact sites required for peroxisome movement. How peroxisomes recognize this subset of EEs and what peroxisome-membrane proteins exist that can interact with EEs is not known. Here, we undertook a forward mutagenesis screen to identify such proteins. We discovered an acyl-coA binding (ACB) domain-containing protein AcbdA/AN1062 that localizes to peroxisomes via its tail-anchored transmembrane domain (TMD). Deleting the AcbdA gene or only its N-terminal ACB domain perturbs the movement and distribution of peroxisomes. Importantly, AcbdA is not required for the movement of EEs or for the recruitment of PxdA and DipA on EEs. Fatty acid (FA)-induced increases in peroxisome movement require AcbdA, suggesting that peroxisome hitchhiking on EEs is coupled to FA metabolism. Mutating a conserved FFAT motif, predicted to interact with the endoplasmic reticulum (ER), has no effect on peroxisome movement. Taken together, our data indicate that AcbdA is a peroxisome-membrane protein required to tether peroxisomes to EEs during hitchhiking. AcbdAs involvement in peroxisome-EE contact site formation represents a divergence from known functions of Acbd4/5 proteins and adds layers to our understanding of the functionality of the Acbd4/5 family of proteins.

cell biology↗

bioRxiv2023

Woronin bodies move dynamically and bidirectionally by hitchhiking on early endosomes in Aspergillus nidulans

The proper functioning of organelles depends on their intracellular localization, mediated by motor protein-dependent transport on cytoskeletal tracks. Rather than directly associating with a motor protein, peroxisomes move by hitchhiking on motile early endosomes in the filamentous fungus Aspergillus nidulans. However, the cellular function of peroxisome hitchhiking is unclear. Peroxisome hitchhiking requires the protein PxdA, which is conserved within the fungal subphylum Pezizomycotina, but absent from other fungal clades. Woronin bodies are specialized peroxisomes that are also unique to the Pezizomycotina. In these fungi, multinucleate hyphal segments are separated by incomplete cell walls called septa that possess a central pore enabling cytoplasmic exchange. Upon damage to a hyphal segment, Woronin bodies plug septal pores to prevent wide-spread leakage. Here, we tested if peroxisome hitchhiking is important for Woronin body motility, distribution, and function in A. nidulans. We show that Woronin body proteins are present within all motile peroxisomes and hitchhike on PxdA-labeled early endosomes during bidirectional, long-distance movements. Loss of peroxisome hitchhiking by knocking out pxdA significantly affected Woronin body distribution and motility in the cytoplasm, but Woronin body hitchhiking is ultimately dispensable for septal localization and plugging.

cell biology↗

bioRxiv2021

Cytoplasmic dynein-1 cargo diversity is mediated by the combinatorial assembly of FTS-Hook-FHIP complexes

In eukaryotic cells, intracellular components are organized by the microtubule motors cytoplasmic dynein-1 (dynein) and kinesins, which are linked to cargos via adaptor proteins. While [~]40 kinesins transport cargo toward the plus end of microtubules, a single dynein moves cargo in the opposite direction. How dynein transports a wide variety of cargos remains an open question. The FTS-Hook-FHIP ("FHF") cargo adaptor complex links dynein to cargo in mammals and fungi. As human cells have three Hooks and four FHIP proteins, we hypothesized that the combinatorial assembly of different Hook and FHIP proteins could underlie dynein cargo diversity. Using proteomic approaches, we determine the protein interactome of each FHIP protein. Live-cell imaging and biochemical approaches show that different FHF complexes associate with distinct motile cargos. These complexes also move with dynein and its cofactor dynactin in single-molecule in vitro reconstitution assays. Complexes composed of FTS, FHIP1B, and Hook1/Hook3 co-localize with Rab5-tagged early endosomes via a direct interaction between FHIP1B and GTP-bound Rab5. In contrast, complexes composed of FTS, FHIP2A and Hook2 colocalize with Rab1A-tagged ER-to-Golgi cargos and FHIP2A is involved in the motility of Rab1A tubules. Our findings suggest that combinatorial assembly of different FTS-Hook-FHIP complexes is one mechanism dynein uses to achieve cargo specificity.

cell biology↗

bioRxiv2021

Optimizing microtubule arrangements for rapid cargo capture

Cellular functions such as autophagy, cell signaling and vesicular trafficking involve the retrograde transport of motor-driven cargo along microtubules. Typically, newly formed cargo engages in slow diffusive movement from its point of origin before attaching to a microtubule. In some cell types, cargo destined for delivery to the perinuclear region relies on capture at dynein-enriched loading zones located near microtubule plus-ends. Such systems include extended cell regions of neurites and fungal hyphae, where the efficiency of the initial diffusive loading process depends on the axial distribution of microtubule plus-ends relative to the initial cargo position. We use analytic mean first passage time calculations and numerical simulations to model diffusive capture processes in tubular cells, exploring how the spatial arrangement of microtubule plus-ends affects the efficiency of retrograde cargo transport. Our model delineates the key features of optimal microtubule ar-rangements that minimize mean cargo capture times. Namely, we show that configurations with a single long microtubule and broad distribution of additional microtubule plus-ends allow for efficient capture in a variety of different scenarios for retrograde transport. Live-cell imaging of microtubule plus-ends in Aspergillus nidulans hyphae indicates that their distributions exhibit these optimal qualitative features. Our results highlight important coupling effects between microtubule length distribution and retrograde cargo transport, providing guiding principles for the spatial arrangement of microtubules within tubular cell regions.

biophysics↗

bioRxiv2020

Wnt-mediated interactions of tumor-initiating cells with a macrophage niche drive skin tumor formation

Skin tumor-initiating stem cells (tSCs) fuel skin squamous cell carcinoma (SCC) formation and development in a complex tumor microenvironment, but a role for immune cells in the tSC niche governing the process of tumor formation has remained elusive. Here, we define the existence of a tSC-macrophage niche, expressing high levels of Wnt ligands. Using conditional mouse genetic models to abrogate the secretion of Wnts, we show that both hair follicle SC- and macrophage-derived Wnts are essential for driving skin tumorigenesis, tSCs maintenance, and counteracting tumor regression. Loss of Wnts in either population uncouples the tSC-macrophage association. The proteomic signature of SCC cells reveals CD99 as a Wnt-dependent receptor for the macrophagecancer cell interaction. These results establish a role for a macrophage-tSC niche in governing SCC initiation and maintenance, uncovering potential candidates for immunoprevention against SCC. One Sentence SummaryA macrophage-skin tumor-initiating cells bonding Wnt loop drives tumorigenesis.

cancer biology↗

bioRxiv2020

Regulation of peroxisome and lipid droplet hitchhiking by PxdA and the DipA phosphatase

In canonical microtubule-based transport, adaptor proteins link cargos to the molecular motors dynein and kinesin. Recently, an alternative mode of transport known as hitchhiking was discovered, in which a cargo achieves motility by hitching a ride on an already-motile cargo, rather than attaching to a motor protein. Hitchhiking has been best-studied in two filamentous fungi, Aspergillus nidulans and Ustilago maydis. In U. maydis, ribonucleoprotein complexes, peroxisomes, lipid droplets, and endoplasmic reticulum all hitchhike on early endosomes. In A. nidulans, peroxisomes hitchhike using a putative molecular linker, PxdA, that associates with early endosomes. However, whether other organelles use PxdA to hitchhike on early endosomes is unclear, as are the molecular mechanisms that regulate hitchhiking in A. nidulans. Here we find that the proper distribution of lipid droplets, mitochondria and autophagosomes do not require PxdA, suggesting that PxdA is a molecular linker specific to peroxisomes. We also identify two new pxdA alleles, including a point mutation (R2044P) that disrupts PxdAs ability to associate with early endosomes and reduces peroxisome movement. Finally, we identify a novel regulator of peroxisome hitchhiking, the phosphatase DipA. DipA co-localizes with early endosomes and its early endosome-association relies on PxdA.

cell biology↗

bioRxiv2019

Hitching a Ride: Mechanics of Organelle Transport Through Linker-Mediated Hitchhiking

In contrast to the canonical picture of transport by direct attachment to motor proteins, recent evidence shows that a number of intracellular cargos navigate the cytoplasm by hitchhiking on motor-driven carrier organelles. We describe a quantitative model of intracellular cargo transport via hitchhiking, examining the efficiency of hitchhiking initiation as a function of geometric and mechanical parameters. We focus specifically on the parameter regime relevant to the hitchhiking motion of peroxisome organelles in fungal hyphae. Our work predicts the dependence of transport initiation rates on the distribution of cytoskeletal tracks and carrier organelles, as well as the number, length and flexibility of the linker proteins that mediate contact between the carrier and the hitchhiking cargo. Furthermore, we demonstrate that attaching organelles to microtubules can result in a substantial enhancement of the hitchhiking initiation rate in tubular geometries such as those found in fungal hyphae. This enhancement is expected to increase the overall transport rate of hitchhiking organelles, and lead to greater efficiency in organelle dispersion. Our results leverage a quantitative physical model to highlight the importance of organelle encounter dynamics in non-canonical intracellular transport.\n\nSIGNIFICANCEA variety of cellular components are transported via hitchhiking by attaching to other motile organelles. Defects in the molecular machinery responsible for organelle hitchhiking may be linked with neurodegenerative disorders. To date, no comprehensive physical models of this non-canonical mode of transport have been developed. In particular, the connection between molecular- and organelle-scale properties of hitchhiking components and their effect on cellular-scale transport has remained unclear. Here, we investigate the mechanics of hitchhiking initiation and explore organelle interactions that can modulate the efficiency of this process.

biophysics↗

bioRxiv2016

Chlamydomonas reinhardtii formin and profilin are optimized for acute rapid actin filament assembly

The regulated assembly of multiple filamentous actin (F-actin) networks from an actin monomer pool is important for a variety of cellular processes. Chlamydomonas reinhardtii is a unicellular green alga expressing a conventional and divergent actin that is an emerging system for investigating the complex regulation of actin polymerization. One actin network that contains exclusively conventional F-actin in Chlamydomonas is the fertilization tubule, a mating structure at the apical cell surface in gametes. In addition to two actin genes, Chlamydomonas expresses a profilin (PRF1) and four formin genes (FOR1-4), one of which (FOR1) we have characterized for the first time. We found that unlike typical profilins, PRF1 prevents unwanted actin assembly by strongly inhibiting both F-actin nucleation and barbed end elongation at equimolar concentrations to actin. However, FOR1 stimulates the assembly of rapidly elongating actin filaments from PRF1-bound actin. PRF1 further favors FOR1-mediated actin assembly by potently inhibiting Arp2/3 complex-mediated actin assembly. Furthermore, for1 and prf1-1 mutants, as well as the small molecule formin inhibitor SMIFH2, prevent fertilization tubule formation in gametes, suggesting that polymerization of F-actin for fertilization tubule formation is a primary function of FOR1. Together, these findings indicate that FOR1 and PRF1 cooperate to selectively and rapidly assemble F-actin at the right time and place.\n\nSUMMARY STATEMENTThe Chlamydomonas reinhardtii formin FOR1 initiates rapid assembly of fertilization tubule actin filaments from monomers associated with the actin-assembly inhibitor profilin PRF1.

cell biology↗