13 Plus 1: A 30-Year Perspective on Microtubule-Based Motility in Dictyostelium

Koonce, Michael P.

doi:10.3390/cells9030528

Cited by 5 publications

(6 citation statements)

References 53 publications

(63 reference statements)

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“…Thus, a minus-directed motor is highly activated by addition of PA to SLBs. This motor is likely dynein because processive minus-directed kinesins are not found in the genome of Dictyostelium (28). In an earlier report, we have found that Dynein motors cluster into cholesterol-rich microdomains on mature phagosomes, yielding robust motion and frequent minus-directed stalls (9).…”

Section: Resultsmentioning

confidence: 99%

Phosphatidic Acid Dependent Recruitment of Microtubule Motors to Spherical Supported Lipid Bilayers forIn-vitroMotility Assays

Kumar,

Sanghavi,

Chaudhury

et al. 2023

Preprint

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A variety of membrane-bound vesicles are transported by the kinesin and dynein motor proteins respectively towards plus and minus ends of microtubules inside cells. How specific lipids, particularly rare lipids, bind and activate motors on the vesicle membrane and the biological consequences thereof are poorly understood. Here we preparesphericalsupported lipid bilayers (SLBs) consisting of a latex bead enclosed within a membrane that contains phosphatidic acid (PA). When incubated with motor-protein enriched fractions fromDictyosteliumor from Rat brain, PA-coated SLBs exhibit robust motion alongin-vitropolymerized microtubules. We quantify the activity of SLB-bound motors by measuring their force generation against an Optical trap on polarity-labelled microtubules. Such experiments reveal that PA recruits dynein-dynactin fromDictyostelium, but kinesin-1 from rat brain fractions to drive SLB motion. Kinesin-1 is able to dominate over dynein when dynactin, an activator of dynein, is deficient on SLBs. We demonstrate how inhibiting only kinesin can block both kinesin and dynein activity on a cargo, thus providing an explanation for the long-standing “paradox of co-dependence” in the field of intracellular transport. We also find that as the size of a motor-driven SLB increases, the SLB is able to reverse direction more easily along a microtubule. This observation provides clues as to how micron-sized cargoes are efficiently able to navigate the crowded cytoskeletal space inside cells.

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Section: Resultsmentioning

confidence: 99%

Phosphatidic Acid Dependent Recruitment of Microtubule Motors to Spherical Supported Lipid Bilayers forIn-vitroMotility Assays

Kumar,

Sanghavi,

Chaudhury

et al. 2023

Preprint

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“…During growth, the most significantly downregulated genes were those encoding heat shock proteins ( hspE-1, hspG3 , hspG4, hspG5, hspG6, hspG7, hspG8, hspG12, hspH, hspJ, hspM, dnaja1 ) and genes involved in protein ubiquitination ( ubqG, ubqH, ubqI, ubqJ, DDB_G0285907 ) ( Supplementary Table S1 ). Conversely, the most significantly upregulated genes were those associated with cell cycle progression, including but not limited to, subunits of the anaphase-promoting complex ( anapc3, anapc5, anapc6, anapc7, anapc10 ), and genes involved in mitosis, such as kinesin-related genes ( kif2, kif4, kif10, kif12, kif13 ) ( Castro et al, 2005 ; Nag et al, 2008 ; Tikhonenko et al, 2009 ; Koonce, 2020 ). During starvation, genes related to development ( cotC, cotD, cotE ) were significantly downregulated in cln5 - cells.…”

Section: Resultsmentioning

confidence: 99%

An altered transcriptome underlies cln5-deficiency phenotypes in Dictyostelium discoideum

Kim

Huber

2022

Front. Genet.

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Mutations in CLN5 cause a subtype of neuronal ceroid lipofuscinosis (NCL) called CLN5 disease. The NCLs, commonly referred to as Batten disease, are a family of neurodegenerative lysosomal storage diseases that affect all ages and ethnicities globally. Previous research showed that CLN5 participates in a variety of cellular processes. However, the precise function of CLN5 in the cell and the pathway(s) regulating its function are not well understood. In the model organism Dictyostelium discoideum, loss of the CLN5 homolog, cln5, impacts various cellular and developmental processes including cell proliferation, cytokinesis, aggregation, cell adhesion, and terminal differentiation. In this study, we used comparative transcriptomics to identify differentially expressed genes underlying cln5-deficiency phenotypes during growth and the early stages of multicellular development. During growth, genes associated with protein ubiquitination/deubiquitination, cell cycle progression, and proteasomal degradation were affected, while genes linked to protein and carbohydrate catabolism were affected during early development. We followed up this analysis by showing that loss of cln5 alters the intracellular and extracellular amounts of proliferation repressors during growth and increases the extracellular amount of conditioned medium factor, which regulates cAMP signalling during the early stages of development. Additionally, cln5- cells displayed increased intracellular and extracellular amounts of discoidin, which is involved in cell-substrate adhesion and migration. Previous work in mammalian models reported altered lysosomal enzyme activity due to mutation or loss of CLN5. Here, we detected altered intracellular activities of various carbohydrate enzymes and cathepsins during cln5- growth and starvation. Notably, cln5- cells displayed reduced β-hexosaminidase activity, which aligns with previous work showing that D. discoideum Cln5 and human CLN5 can cleave the substrate acted upon by β-hexosaminidase. Finally, consistent with the differential expression of genes associated with proteasomal degradation in cln5- cells, we also observed elevated amounts of a proteasome subunit and reduced proteasome 20S activity during cln5- growth and starvation. Overall, this study reveals the impact of cln5-deficiency on gene expression in D. discoideum, provides insight on the genes and proteins that play a role in regulating Cln5-dependent processes, and sheds light on the molecular mechanisms underlying CLN5 disease.

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“…DdKif10 is a 1238aa (139 kDa) polypeptide with an amino-terminal motor domain, a central region with two prominent predicted coiled-coil motifs, and a carboxy terminal tail that likely contains cargo-binding regions [17,18]. The motor/neck domain clearly groups to the Kinesin-8 family, with 45% identity over the first 425 residues to the human Kif18A protein.…”

Section: Resultsmentioning

confidence: 99%

“…Finally, our previous work suggests that DdKif10 collaborates with DdKif8 (Kinesin-4) to counterbalance dynein pulling activity during interphase in D. discoideum, acting in sum to support the radial MT array [7,18]. Laser cutting of the motile comet-like MT arrays induced by dominant-negative expression of dynein fragments indicates a pushing component that drives MT motility (MT plus-end-directed) [24], and we are unable to produce this distinctive MT behavior in either kinesin null cell background.…”

Section: Discussionmentioning

confidence: 99%

Bioenergetics of the Dictyostelium Kinesin-8 Motor Isoform

Koonce

Tikhonenko

2020

Biomolecules

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The functional organization of microtubules in eukaryotic cells requires a combination of their inherent dynamic properties, interactions with motor machineries, and interactions with accessory proteins to affect growth, shrinkage, stability, and architecture. In most organisms, the Kinesin-8 family of motors play an integral role in these organizations, well known for their mitotic activities in microtubule (MT) length control and kinetochore interactions. In Dictyostelium discoideum, the function of Kinesin-8 remains elusive. We present here some biochemical properties and localization data that indicate that this motor (DdKif10) shares some motility properties with other Kinesin-8s but also illustrates differences in microtubule localization and depolymerase action that highlight functional diversity.

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13 Plus 1: A 30-Year Perspective on Microtubule-Based Motility in Dictyostelium

Cited by 5 publications

References 53 publications

Phosphatidic Acid Dependent Recruitment of Microtubule Motors to Spherical Supported Lipid Bilayers forIn-vitroMotility Assays

Phosphatidic Acid Dependent Recruitment of Microtubule Motors to Spherical Supported Lipid Bilayers forIn-vitroMotility Assays

An altered transcriptome underlies cln5-deficiency phenotypes in Dictyostelium discoideum

Bioenergetics of the Dictyostelium Kinesin-8 Motor Isoform

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