MAP1B Regulates Axonal Development by Modulating Rho-GTPase Rac1 Activity

This article shows a novel function for the MAP1B protein, related to the control of actin dynamics through interaction with Tiam1

Strategic directions in constraint programming

An abstract is not available

A reference human induced pluripotent stem cell line for large-scale collaborative studies

Human induced pluripotent stem cell (iPSC) lines are a powerful tool for studying development and disease, but the considerable phenotypic variation between lines makes it challenging to replicate key findings and integrate data across research groups. To address this issue, we sub-cloned candidate human iPSC lines and deeply characterized their genetic properties using whole genome sequencing, their genomic stability upon CRISPR-Cas9-based gene editing, and their phenotypic properties including differentiation to commonly used cell types. These studies identified KOLF2.1J as an all-around well-performing iPSC line. We then shared KOLF2.1J with groups around the world who tested its performance in head-to-head comparisons with their own preferred iPSC lines across a diverse range of differentiation protocols and functional assays. On the strength of these findings, we have made KOLF2.1J and its gene-edited derivative clones readily accessible to promote the standardization required for large-scale collaborative science in the stem cell field

Dysfunction of Heterotrimeric Kinesin-2 in Rod Photoreceptor Cells and the Role of Opsin Mislocalization in Rapid Cell Death

Loss of kinesin-2 function causes rapid death of rod photoreceptors. The cell death is dependent on the expression of opsin, which first accumulates along the route to the outer segment, but not on signaling by opsin-arrestin complexes or by light activation; the key element appears to be the accumulation of excessive protein in the wrong place

Dynactin Subunit p150^Glued Is a Neuron-Specific Anti-Catastrophe Factor

<div><p>Regulation of microtubule dynamics in neurons is critical, as defects in the microtubule-based transport of axonal organelles lead to neurodegenerative disease. The microtubule motor cytoplasmic dynein and its partner complex dynactin drive retrograde transport from the distal axon. We have recently shown that the p150^Glued subunit of dynactin promotes the initiation of dynein-driven cargo motility from the microtubule plus-end. Because plus end-localized microtubule-associated proteins like p150^Glued may also modulate the dynamics of microtubules, we hypothesized that p150^Glued might promote cargo initiation by stabilizing the microtubule track. Here, we demonstrate <i>in vitro</i> using assembly assays and TIRF microscopy, and in primary neurons using live-cell imaging, that p150^Glued is a potent anti-catastrophe factor for microtubules. p150^Glued alters microtubule dynamics by binding both to microtubules and to tubulin dimers; both the N-terminal CAP-Gly and basic domains of p150^Glued are required in tandem for this activity. p150^Glued is alternatively spliced <i>in vivo</i>, with the full-length isoform including these two domains expressed primarily in neurons. Accordingly, we find that RNAi of p150^Glued in nonpolarized cells does not alter microtubule dynamics, while depletion of p150^Glued in neurons leads to a dramatic increase in microtubule catastrophe. Strikingly, a mutation in p150^Glued causal for the lethal neurodegenerative disorder Perry syndrome abrogates this anti-catastrophe activity. Thus, we find that dynactin has multiple functions in neurons, both activating dynein-mediated retrograde axonal transport and enhancing microtubule stability through a novel anti-catastrophe mechanism regulated by tissue-specific isoform expression; disruption of either or both of these functions may contribute to neurodegenerative disease.</p></div

p150^Glued promotes microtubule formation.

<p>(A) Schematic depicting endogenous full-length p150^Glued and dimeric and monomeric constructs. (B) Light scattering traces for recombinant polypeptides or buffer control incubated with tubulin show that the dimeric p150 Nt-GCN4 and neuronal MAPs tau23 and DCX-GFP robustly promote microtubule assembly, while monomeric p150 Nt and rKin430-GFP do not. (C) TIRF elongation assay. As shown in the schematic at top, rhodamine-labeled tubulin is polymerized from biotinylated Alexa488-labeled GMPCPP microtubule seeds. Montage below shows free tubulin assembly (red) from a stabilized seed (green) in the absence of p150. Arrowhead identifies the microtubule plus-end. (D) Kymograph plots showing representative examples with buffer, 200 nM p150 Nt, or p150 Nt-GCN4. (E) Polymerization rates and (F) catastrophe frequencies from seeded assembly shows that p150 Nt-GCN4 promotes polymerization and inhibits catastrophe. All error bars represent SEM of three or more independent experiments. Statistical testing was performed with a two-tailed <i>t</i> test. * <i>p</i><0.05; ** <i>p</i><0.01. For (E), <i>p</i><0.01 for all conditions compared to control.</p