Functional fluorescence labeling of actins in live-cell system using engineered split-GFP technology.

Actin filaments play essential roles in various cellular processes, and understanding their dynamics is crucial for studying cellular behaviors and actin-related diseases. However, conventional methods for visualizing actins often perturb its functionality or lack sufficient resolution for real-time imaging. In this study, we developed a method for functional fluorescence labeling of actin isoforms using split-GFP technology, specifically through insertion of a GFP11 tag into a flexible residue pair (T229/A230) of human β-actin (ACTB) or γ-actin (ACTG).

A mutual co-recognition mechanism ensures the proper assembly of heterotrimeric kinesin-2 for intraflagellar transport.

Heterotrimeric kinesin-2, composed of two distinct kinesin motors and a kinesin-associated protein (KAP), is essential for intraflagellar transport and ciliogenesis. KAP specifically recognizes the hetero-paired motor tails for the holoenzyme assembly, but the underlying mechanism remains unclear. Here, we determine the structure of KAP-1 in complex with the hetero-paired tails from kinesin-2 motors KLP-20 and KLP-11.

Functional residuomics - analyzing how missense mutations impact cellular systems.

Functional residuomics explores how individual amino acid residues influence protein function, interactions and cellular homeostasis, shifting the focus from gene-level mutations to residue-level alterations. Unlike gene-centric approaches, residuomics systematically examines missense mutations across the proteome, linking small changes in amino acid sequences to organelle dynamics and tissue phenotypes. By integrating mutagenesis with high-throughput phenotyping, this approach connects atomic-scale changes to larger biological systems, offering valuable insights for clinical diagnostics and therapeutic development.

Rapid memory shift between different synaptic ensembles promotes forgetting in Drosophila.

Forgetting is increasingly recognized as an active adaptive process, yet its mechanism remains elusive at the cellular and synaptic levels. Here, we show that the natural decay of hours-scale memory in Drosophila is promoted through shifting memory traces between distinct synaptic ensembles. Aversive conditioning produces an initial memory trace of presynaptic depression primarily confined to the γ1 compartment, one of five axonal compartments of γ Kenyon cells, that drives avoidance behavior.

Kinesin-2 autoinhibition requires elbow phosphorylation.

Kinesin motor proteins facilitate microtubule-based transport by converting chemical energy into mechanical forces, but this activity is autoinhibited until cargo is loaded. Regulatory mechanisms underlying this autoinhibitory conformation are not well understood. Here, we show that a NEver in mitosis Kinase NEKL-3 directly phosphorylates a flexible elbow region between two coiled-coil domains connecting the motor head and tail of an intraflagellar transport kinesin, OSM-3.

Phosphorylation-dependent regional motility of the ciliary kinesin OSM-3.

Kinesin motor proteins, vital for intracellular microtubule-based transport, display region-specific motility within cells, a phenomenon that remains molecularly enigmatic. This study focuses on the localized activation of OSM-3, an intraflagellar transport kinesin crucial for the assembly of ciliary distal segments in Caenorhabditis elegans sensory neurons. Fluorescence lifetime imaging microscopy unveiled an extended, active conformation of OSM-3 in the ciliary base and middle segments, where OSM-3 is conveyed as cargo by kinesin-II.

Alpha-tubulin tails regulate axoneme differentiation.

The tubulin tail is a key element for microtubule (MT) functionality, but the functional redundancy of tubulin genes complicates the genetic determination of their physiological functions. Here, we removed the C-terminal tail of five alpha- and four beta-tubulin genes in the genome. Sensory cilia typically exhibit an axoneme that longitudinally differentiates into a middle segment with doublet MTs and a distal segment with singlet MTs.

Multifaceted regulation of asymmetric cell division by the actin cytoskeleton.

Asymmetric cell division (ACD) is essential for generating cell diversity in multicellular eukaryotes, yet the underlying mechanisms remain largely unresolved. Well-established models of ACD, such as microtubule-based spindle displacement in Caenorhabditis elegans embryos and preprophase band assembly in plants provide valuable insights but fail to fully explain asymmetry establishment in others. In this article, we discuss how actin-dependent mechanisms govern ACD in model systems, highlighting emerging commonalities and differences.

Conformational reorganization and phase separation drive hyper-editing of ADR-2-ADBP-1 complex.

Adenosine deaminase acting on RNA (ADAR) proteins, which mediate adenosine-to-inosine editing of double-stranded ribonucleic acid (dsRNA) substrates, play essential roles in balancing innate immunity. Using cryogenic electron microscopy, we solved the structure of the Caenorhabditis elegans ADR-2-ADBP-1 complex (stoichiometric ratio, 2:2), which is an asymmetric ADR-2 dimer with one editing site blocked by the other ADR-2. Unexpectedly, dsRNA recruitment triggered dissociation of the ADR-2 dimer, exposing more competent dsRNA editing sites.

Ephrin A1 functions as a ligand of EGFR to promote EMT and metastasis in gastric cancer.

Distant metastasis is the major cause of gastric cancer mortality, and epidermal growth factor receptor (EGFR) activation plays critical roles in gastric cancer dissemination. However, EGFR targeting therapies in gastric cancer show only marginal effects, and the molecular mechanisms of oncogenic EGFR signaling remain poorly defined. Here, we report Ephrin A1 as a novel ligand of EGFR in gastric cancer.

Distinct roles of two homologous kinesins in mammalian motile cilia.

How do the two kinesin-9 members Kif6 and Kif9 function in mammalian cilia? Ou and colleagues discuss new work from Fang et al. (https://doi.org/10.

Ephrin A1 Stimulates CCL2 Secretion to Facilitate Premetastatic Niche Formation and Promote Gastric Cancer Liver Metastasis.

The liver is a primary target for distal metastasis of gastric cancer. The hepatic premetastatic niche (PMN) facilitates crucial communications between primary tumor and liver, thereby playing an essential role in hepatic metastasis. Identification of the molecular mechanisms driving PMN formation in gastric cancer could facilitate development of strategies to prevent and treat liver metastasis.

A machine learning enhanced EMS mutagenesis probability map for efficient identification of causal mutations in Caenorhabditis elegans.

Chemical mutagenesis-driven forward genetic screens are pivotal in unveiling gene functions, yet identifying causal mutations behind phenotypes remains laborious, hindering their high-throughput application. Here, we reveal a non-uniform mutation rate caused by Ethyl Methane Sulfonate (EMS) mutagenesis in the C. elegans genome, indicating that mutation frequency is influenced by proximate sequence context and chromatin status.

AlphaFold2-guided engineering of split-GFP technology enables labeling of endogenous tubulins across species while preserving function.

Dynamic properties are essential for microtubule (MT) physiology. Current techniques for in vivo imaging of MTs present intrinsic limitations in elucidating the isotype-specific nuances of tubulins, which contribute to their versatile functions. Harnessing the power of the AlphaFold2 pipeline, we engineered a strategy for the minimally invasive fluorescence labeling of endogenous tubulin isotypes or those harboring missense mutations.

Cryo-electron tomography elucidates annular intraluminal configurations in Caenorhabditis elegans microtubules.

Background Information: Microtubules serve as integral components in cellular operations such as cell division, intracellular trafficking, and cellular architecture. Composed of tubulin protein subunits, these hollow tubular structures have been increasingly elucidated through advanced cryo-electron microscopy (Cryo-EM), which has unveiled the presence of microtubule inner proteins (MIPs) within the microtubular lumen.

Results: In the present investigation, we employ a synergistic approach incorporating high-pressure freezing, cryo-focused ion beam milling, and Cryo-electron tomography (Cryo-ET) to interrogate the in situ architecture of microtubules in Caenorhabditis elegans larvae.

Vacuolar H-ATPase determines daughter cell fates through asymmetric segregation of the nucleosome remodeling and deacetylase complex.

Asymmetric cell divisions (ACDs) generate two daughter cells with identical genetic information but distinct cell fates through epigenetic mechanisms. However, the process of partitioning different epigenetic information into daughter cells remains unclear. Here, we demonstrate that the nucleosome remodeling and deacetylase (NuRD) complex is asymmetrically segregated into the surviving daughter cell rather than the apoptotic one during ACDs in .

Inhibition of a cyclic nucleotide-gated channel on neuronal cilia activates unfolded protein response in intestinal cells to promote longevity.

Ciliary defects are linked to ciliopathies, but impairments in the sensory cilia of neurons extend lifespan, a phenomenon with previously unclear mechanisms. Our study reveals that neuronal cilia defects trigger the unfolded protein response of the endoplasmic reticulum (UPR) within intestinal cells, a process dependent on the insulin/insulin-like growth factor 1 (IGF-1) signaling transcription factor and the release of neuronal signaling molecules. While inhibiting UPR doesn't alter the lifespan of wild-type worms, it normalizes the extended lifespan of ciliary mutants.

Transposase-assisted tagmentation: an economical and scalable strategy for single-worm whole-genome sequencing.

AlphaMissense identifies 23 million human missense variants as likely pathogenic, but only 0.1% have been clinically classified. To experimentally validate these predictions, chemical mutagenesis presents a rapid, cost-effective method to produce billions of mutations in model organisms.

Loss of cilia in chemosensory neurons inhibits pathogen avoidance in Caenorhabditis elegans.

Pathogen avoidance is a crucial and evolutionarily conserved behavior that enhances survival by preventing infection in diverse species, including Caenorhabditis elegans (C. elegans). This behavior relies on multiple chemosensory neurons equipped with cilia that are exposed to the external environment.

Neurons dispose of hyperactive kinesin into glial cells for clearance.

Microtubule-based kinesin motor proteins are crucial for intracellular transport, but their hyperactivation can be detrimental for cellular functions. This study investigated the impact of a constitutively active ciliary kinesin mutant, OSM-3CA, on sensory cilia in C. elegans.

KIF11 UFMylation Maintains Photoreceptor Cilium Integrity and Retinal Homeostasis.

The photoreceptor cilium is vital for maintaining the structure and function of the retina. However, the molecular mechanisms underlying the photoreceptor cilium integrity and retinal homeostasis are largely unknown. Herein, it is shown that kinesin family member 11 (KIF11) localizes at the transition zone (connecting cilium) of the photoreceptor and plays a crucial role in orchestrating the cilium integrity.

Utilizing the scale-invariant feature transform algorithm to align distance matrices facilitates systematic protein structure comparison.

Motivation: Protein structure comparison is pivotal for deriving homological relationships, elucidating protein functions, and understanding evolutionary developments. The burgeoning field of in-silico protein structure prediction now yields billions of models with near-experimental accuracy, necessitating sophisticated tools for discerning structural similarities among proteins, particularly when sequence similarity is limited.

Results: In this article, we have developed the align distance matrix with scale (ADAMS) pipeline, which synergizes the distance matrix alignment method with the scale-invariant feature transform algorithm, streamlining protein structure comparison on a proteomic scale.

A plant flavonol and genetic suppressors rescue a pathogenic mutation associated with kinesin in neurons.

KIF1A, a microtubule-based motor protein responsible for axonal transport, is linked to a group of neurological disorders known as KIF1A-associated neurological disorder (KAND). Current therapeutic options for KAND are limited. Here, we introduced the clinically relevant KIF1A(R11Q) variant into the homolog UNC-104, resulting in uncoordinated animal behaviors.

Inhibition of the chromatin remodeling factor NURF rescued sterility by a clinic variant of NuRD.

The nucleosome remodeling and deacetylase (NuRD) complex is essential for gene expression and cell fate determination, and missense mutations of NuRD caused neurodevelopmental diseases. However, the molecular pathogenesis of clinic NuRD variants is unknown. Here, we introduced a clinic CHD3 (L915F) variant into homologue LET-418, impairing germline and vulva development and ultimately causing animal sterility.