Microtubule Motor Research Articles

MRNA 5-methylcytosine in Eimeria tenella oocysts: An abundant post-transcriptional modification associated with broad-ranging biological processes

Eimeria tenella is the major causative agent of chicken coccidiosis. 5-Methylcytosine (m5C) is a type of RNA chemical modifications reported to regulate diverse biological processes. However, the distribution and biological functions of m5C in E. tenella mRNAs are yet to be known. Herein, we report transcriptome-wide profiling of mRNA m5C in E. tenella by employing m5C RNA immunoprecipitation followed by a deep-sequencing approach (m5C-RIP-seq). Our data showed that m5C peaks were distributed across the whole mRNA body. Compared with unsporulated oocysts, there were 2813 hypermethylated and 1850 hypomethylated m5C peaks in sporulated oocysts. Generally, a positive correlation between m5C modification and gene expression levels was observed. The mRNA sequencing (RNA-seq) and m5C-RIP-seq data were consistent with the results of the quantitative reverse transcription PCR (RT-qPCR) and methylated RNA immunoprecipitation-qPCR (MeRIP-qPCR), respectively. Gene Ontology (GO) and pathway enrichment analysis predicated diverse biological functions and pathways, including microtubule motor activity, helicase activity, cGMP-PKG signaling pathway, aminoacyl-tRNA biosynthesis, glycolysis/gluconeogenesis, and spliceosome. Meanwhile, stage-specific gene expression signatures of m5C-related regulators were observed. Altogether, our findings reveal the transcriptional significance of m5C modification in E. tenella oocysts, providing resources and clues for further in-depth research.

"Mitotic" kinesin-5 is a dynamic brake for axonal growth.

During neuronal development, neurons undergo significant microtubule reorganization to shape axons and dendrites, establishing the framework for efficient wiring of the nervous system. Previous studies from our laboratory demonstrated the key role of kinesin-1 in driving microtubule-microtubule sliding, which provides the mechanical forces necessary for early axon outgrowth and regeneration in Drosophila melanogaster. In this study, we reveal the critical role of kinesin-5, a mitotic motor, in modulating the development of postmitotic neurons. Kinesin-5, a conserved homotetrameric motor, typically functions in mitosis by sliding antiparallel microtubules apart in the spindle. Here, we demonstrate that the Drosophila kinesin-5 homolog, Klp61F, is expressed in larval brain neurons, with high levels in ventral nerve cord (VNC) neurons. Knockdown of Klp61F using a pan-neuronal driver leads to severe locomotion defects and complete lethality in adult flies, mainly due to the absence of kinesin-5 in VNC motor neurons during early larval development. Klp61F depletion results in significant axon growth defects, both in cultured and in vivo neurons. By imaging individual microtubules, we observe a significant increase in microtubule motility, and excessive penetration of microtubules into the axon growth cone in Klp61F-depleted neurons. Adult lethality and axon growth defects are fully rescued by a chimeric human-Drosophila kinesin-5 motor, which accumulates at the axon tips, suggesting a conserved role of kinesin-5 in neuronal development. Altogether, our findings show that at the growth cone, kinesin-5 acts as a brake on kinesin-1-driven microtubule sliding, preventing premature microtubule entry into the growth cone. This regulatory role of kinesin-5 is essential for precise axon pathfinding during nervous system development.

Surface Passivation of Norland Optical Adhesive Improves the Guiding Efficiency of Gliding Microtubules in Microfluidic Devices.

The microtubule-kinesin biomolecular motor system, which is vital for cellular function, holds significant promise for nanotechnological applications. In vitro gliding assays have demonstrated the ability to transport microcargo by propelling microtubules across kinesin-coated surfaces. However, the uncontrolled directional motion of microtubules has posed significant challenges, limiting the system's application for precise cargo delivery. Microfluidic devices provide a means to direct microtubule movement through their geometric features. Norland Optical Adhesive (NOA) is valued for its mold-free application in microfluidic device fabrication; however, microtubules often climb up channel walls, limiting controlled movement. In this study, a surface passivation method for NOA is introduced, using polyethylene glycol via a thiol-ene click reaction. This technique significantly improved the directional control and concentration of microtubules within NOA microchannels. This approach presents new possibilities for the precise application of biomolecular motors in nanotechnology, enabling advancements in the design of microfluidic systems for complex biomolecular manipulations.

Microtubule-associated protein, MAP1B, encodes functionally distinct polypeptides

Microtubule-associated protein, MAP1B, is crucial for neuronal morphogenesis and disruptions in MAP1B function are correlated with neurodevelopmental disorders. MAP1B encodes a single polypeptide that is processed into discrete proteins, a heavy chain (HC) and a light chain (LC); however, it is unclear if these two chains operate individually or as a complex within the cell. In vivo studies have characterized the contribution of MAP1B HC and LC to microtubule and actin-based processes, but their molecular mechanisms of action are unknown. Using in vitro reconstitution with purified proteins, we dissect the biophysical properties of the HC and LC and uncover distinct binding behaviors and functional roles for these MAPs. Our biochemical assays indicate that MAP1B HC and LC do not form a constitutive complex, supporting the hypothesis that these proteins operate independently within cells. Both HC and LC inhibit the microtubule motors, kinesin-3, kinesin-4, and dynein, and differentially affect the severing activity of spastin. Notably, MAP1B LC binds to actin filaments in vitro and can simultaneously bind and crosslink actin filaments and microtubules, a function not observed for MAP1B HC. Phosphorylation of MAP1B HC by DYRK1a negatively regulates its actin-binding activity without significantly affecting its microtubule-binding capacity, suggesting a dynamic contribution of MAP1B HC in cytoskeletal organization. Overall, our study provides new insights into the distinct functional properties of MAP1B HC and LC, underscoring their roles in coordinating cytoskeletal networks during neuronal development.

The Domestication of Wild Boar Could Result in a Relaxed Selection for Maintaining Olfactory Capacity.

Domesticated animals are artificially selected to exhibit desirable traits, however not all traits of domesticated animals are the result of deliberate selection. Loss of olfactory capacity in the domesticated pig (Sus scrofa domesticus) is one example. We used whole transcriptome analysis (RNA-Seq) to compare patterns of gene expression in the olfactory mucosa of the pig and two subspecies of wild boar (Sus scrofa), and investigate candidate genes that could be responsible for the loss of olfactory capacity. We identified hundreds of genes with reductions in transcript abundance in pig relative to wild boar as well as differences between the two subspecies of wild boar. These differences were detected mainly in genes involved in the formation and motility of villi, cilia and microtubules, functions associated with olfaction. In addition, differences were found in the abundances of transcripts of genes related to immune defenses, with the highest levels in continental wild boar subspecies. Overall, the loss of olfactory capacity in pigs appears to have been accompanied by reductions in the expression of candidate genes for olfaction. These changes could have resulted from unintentional selection for reduced olfactory capacity, relaxed selection for maintaining olfactory capacity, pleiotropic effects of genes under selection, or other non-selective processes. Our findings could be a cornerstone for future researches on wild boars, pigs, feral populations, and their evolutionary trajectories, aimed to provide tools to better calibrate species management as well as guidelines for breeders.

Nesprin-2 coordinates opposing microtubule motors during nuclear migration in neurons.

Nuclear migration is critical for the proper positioning of neurons in the developing brain. It is known that bidirectional microtubule motors are required for nuclear transport, yet the mechanism of the coordination of opposing motors is still under debate. Using mouse cerebellar granule cells, we demonstrate that Nesprin-2 serves as a nucleus-motor adaptor, coordinating the interplay of kinesin-1 and dynein. Nesprin-2 recruits dynein-dynactin-BicD2 independently of the nearby kinesin-binding LEWD motif. Both motor binding sites are required to rescue nuclear migration defects caused by the loss of function of Nesprin-2. In an intracellular cargo transport assay, the Nesprin-2 fragment encompassing the motor binding sites generates persistent movements toward both microtubule minus and plus ends. Nesprin-2 drives bidirectional cargo movements over a prolonged period along perinuclear microtubules, which advance during the migration of neurons. We propose that Nesprin-2 keeps the nucleus mobile by coordinating opposing motors, enabling continuous nuclear transport along advancing microtubules in migrating cells.

Kinesin-14 HSET and KlpA are non-processive microtubule motors with load-dependent power strokes

Accurate chromosome segregation during cell division relies on coordinated actions of microtubule (MT)-based motor proteins in the mitotic spindle. Kinesin-14 motors play vital roles in spindle assembly and maintenance by crosslinking antiparallel MTs at the spindle midzone and anchoring spindle MTs’ minus ends at the poles. In this study, we investigate the force generation and motility of the Kinesin-14 motors HSET and KlpA. Our findings reveal that both motors are non-processive, producing single load-dependent power strokes per MT encounter, with estimated load-free power strokes of ~30 and ~35 nm, respectively. Each homodimeric motor generates forces of ~0.5 pN, but when assembled in teams, they cooperate to generate forces of 1 pN or more. Notably, the cooperative activity among multiple motors leads to increased MT-sliding velocities. These results quantitatively elucidate the structure-function relationship of Kinesin-14 motors and underscore the significance of cooperative behavior in their cellular functions.

Structural transitions in kinesin minus-end directed microtubule motility.

Kinesin motor proteins hydrolyze ATP to produce force for spindle assembly and vesicle transport, performing essential functions in cell division and motility, but the structural changes required for force generation are uncertain. We now report high-resolution structures showing new transitions in the kinesin mechanochemical cycle, including power stroke fluctuations upon ATP binding and a post-hydrolysis state with bound ADP + free phosphate. We find that rate-limiting ADP release occurs upon microtubule binding, accompanied by central β-sheet twisting, which triggers the power stroke - stalk rotation and neck mimic docking - upon ATP binding. Microtubule release occurs with β-strand-to-loop transitions, implying that β-strand refolding induces Pi release and the recovery stroke. The strained β-sheet during the power stroke and strand-to-loop transitions identify the β-sheet as the long-sought motor spring.

Herpes simplex virus type-1 cVAC formation in neuronal cells is mediated by dynein motor function and glycoprotein retrieval from the plasma membrane.

Herpesvirus assembly requires the cytoplasmic association of large macromolecular and membrane structures that derive from both the nucleus and cytoplasmic membrane systems. Results from the study of human cytomegalovirus (HCMV) in cells where it organizes a perinuclear cytoplasmic virus assembly compartment (cVAC) show a clear requirement for the minus-end-directed microtubule motor, dynein, for virus assembly. In contrast, the assembly of herpes simplex virus -1 (HSV-1) in epithelial cells where it forms multiple dispersed, peripheral assembly sites is only mildly inhibited by the microtubule-depolymerizing agent, nocodazole. Here, we make use of a neuronal cell line system in which HSV-1 forms a single cVAC and show that dynein and its co-factor dynactin localize to the cVAC, and dynactin is associated with membranes that contain the virion tegument protein pUL11. We also show that the virus membrane-associated structural proteins pUL51 and the viral envelope glycoprotein gE arrive at the cVAC by different routes. Specifically, gE arrives at the cVAC after retrieval from the plasma membrane, suggesting the need for an intact retrograde transport system. Finally, we demonstrate that inhibition of dynactin function profoundly inhibits cVAC formation and virus production during the cytoplasmic assembly phase of infection.IMPORTANCEMany viruses reorganize cytoplasmic membrane systems and macromolecular transport systems to promote the production of progeny virions. Clarifying the mechanisms by which they accomplish this may reveal novel therapeutic strategies and illustrate mechanisms that are critical for normal cellular organization. Here, we explore the mechanism by which HSV-1 moves macromolecular and membrane cargo to generate a virus assembly compartment in the infected cell. We find that the virus makes use of a well-characterized, microtubule-based transport system that is stabilized against drugs that disrupt microtubules.

RNaseH-based ribodepletion of total planarian RNA improves detection of longer and non-polyadenylated transcripts.

The overwhelming majority of RNA species isolated from cells or tissues using organic extraction are ribosomal RNAs (rRNA), whereas a relatively small percentage are messenger RNAs (mRNA). For studies that seek to detect mRNA transcripts and measure changes in their expression, this lopsided ratio of desired transcripts to undesired transcripts creates a significant challenge to obtaining sensitive and reproducible results. One method for improving mRNA detection is to selectively amplify polyadenylated (polyA) mRNA molecules when generating RNA-seq libraries, a strategy that is generally very successful in many species. However, this strategy is less effective when starting with total RNA from some species e.g., the planarian species Schmidtea mediterranea (S.med), as it generates libraries that still contain significant and variable amounts of rRNA reads. Further, commercially available ribodepletion kits do not efficiently deplete rRNAs from these samples because their sequences are divergent from mammalian rRNAs. Here we report a customized, optimized, and economical ribodepletion strategy than allows the generation of comprehensive RNA-seq libraries with less than one percent rRNA contamination. We show that this method improves transcript detection, particularly for those without polyA tails (e.g., core histones) and those that are relatively long (e.g., microtubule motor proteins). Using this custom ribodepletion approach, we also detected many transcripts that are not represented in the most recent set of S.med gene annotations, including a subset that are likely expressed transposable elements (TEs). To facilitate future differential expression analyses of these newly identified loci, we created both an annotation file of the new loci we identified and a bioinformatic pipeline for generating additional annotations from future libraries. As significant recent research shows that TE activation is regulated and functionally important, the resources provided here will provide a starting point for investigating such mechanisms in planarians and other species with less conserved rRNA sequences.

Distinct functions of microtubules and actin filaments in the transportation of the male germ unit in pollen

Flowering plants rely on the polarized growth of pollen tubes to deliver sperm cells (SCs) to the embryo sac for double fertilization. In pollen, the vegetative nucleus (VN) and two SCs form the male germ unit (MGU). However, the mechanism underlying directional transportation of MGU is not well understood. In this study, we provide the first full picture of the dynamic interplay among microtubules, actin filaments, and MGU during pollen germination and tube growth. Depolymerization of microtubules and inhibition of kinesin activity result in an increased velocity and magnified amplitude of VN’s forward and backward movement. Pharmacological washout experiments further suggest that microtubules participate in coordinating the directional movement of MGU. In contrast, suppression of the actomyosin system leads to a reduced velocity of VN mobility but without a moving pattern change. Moreover, detailed observation shows that the direction and velocity of VN’s movement are in close correlations with those of the actomyosin-driven cytoplasmic streaming surrounding VN. Therefore, we propose that while actomyosin-based cytoplasmic streaming influences on the oscillational movement of MGU, microtubules and kinesins avoid MGU drifting with the cytoplasmic streaming and act as the major regulator for fine-tuning the proper positioning and directional migration of MGU in pollen.

The combination of RNA-seq transcriptomics and data-independent acquisition proteomics reveal the mechanisms and function of different gooses testicular development at different stages of laying cycle

Egg production performance is an important economic trait in the poultry industry. In previous studies, attention has often been paid to the growth and development of the ovaries, while there has been less research on the testicular tissue of male goose. Due to various factors, there are usually significant differences in the process of testicular spermatogenesis among different goose breeds. The Jilin white goose (JL) is a high-production local goose species in China, domesticated from Anser cygnoides, which has a high egg-laying performance and the egg-laying period can last from February to July. In the production of goose within Jilin Province, the female goose of Jilin White goose is considered as an important maternal parent of synthetic lines, and ganders from Hungarian white goose (HU), Wanxi white goose (WX) and Jilin white goose are the main male parents. Each year, all 3 gander species begin to exhibit breeding capacity in February and reach the peak of reproductive capacity by April, marked by high fertilization rates. With the gradual increase in temperature, the testicular tissue of Hungarian and Wanxi goose gradually diminishes in its ability to produce sperm. the testicular tissue undergoes significant shrinkage by the end of June, resulting in a near loss of sperm production capability, thereby yielding low fertilization rates. However, the Jilin White goose demonstrates the ability to maintain a stable sperm production capacity. Individuals with low sperm motility contribute to increased seed production costs and pose constraints on the industrial development of livestock and poultry varieties. In this study, transcriptomics and proteomics data from gooses testicular of 3 different goose breeds inclouding Jilin white goose, Wanxi white gooseand Hungary white goose sampled in 2 stages, peak of laying cycle (PLC) and end of laying cycle (ELC). In a comparative analysis between PLC and ELC groups (ELC vs. PLC) of 3 breeds, we identified 401,340,6651 differentially expressed genes (DEGs) and 18,225,323 differentially expressed proteins (DEPs), respectively. Differentially expressed genes and proteins were significantly enriched in Gene Ontology (GO) terms such as phosphotransferase activity, cytoskeletal protein binding, microtubule motor activity, channel activity and carbohydrate metabolic process. The KEGG enrichment analysis of the DEGs in testicular showed that most differentially expressed mRNAs participate in the KEGG pathways, including ECM-receptor interaction, MAPK signaling pathway, carbon metabolism, Cell cycle, VEGF signaling pathway, Lipoic acid metabolism and p53 signaling pathway. The differential expression of 4 selected DEGs (SPAG6, NEK2, HSPA4L, SERF1A) was verified by qRT-PCR, and the results were consistent with RNA-seq data. In conclusion, this study reveals the differences in gene expression regulation in testicular tissues of different goose species, and screening candidate genes and proteins related to spermatogenesis.

Plant Kinesin Repertoires Expand with New Domain Architecture and Contract with the Loss of Flagella.

Kinesins are eukaryotic microtubule motor proteins subdivided into conserved families with distinct functional roles. While many kinesin families are widespreadin eukaryotes, each organismal lineage maintains a unique kinesin repertoire composed of many families with distinct numbers of genes. Previous genomic surveys indicated that land plant kinesin repertoires differ markedly from other eukaryotes. To determine when repertoires diverged during plant evolution, we performed robust phylogenomic analyses of kinesins in 24 representative plants, two algae, two animals, and one yeast. These analyses show that kinesin repertoires expand and contract coincident with major shifts in the biology of algae and land plants. One kinesin family and five subfamilies, eachdefined by unique domain architectures, emerged in the green algae. Four of those kinesin groups expanded in ancestors of modern land plants, while six other kinesin groups were lost in the ancestors of pollen-bearing plants. Expansions of different kinesin families and subfamilies occurred in moss and angiosperm lineages. Other kinesin families remained stable and did not expand throughout plant evolution. Collectively these data support a radiation of kinesin domain architectures in algae followed by differential positive and negative selection on kinesins families and subfamilies in different lineages of land plants.

Sarcopenia-related changes in serum GLP-1 level affect myogenic differentiation.

Sarcopenia, a group of muscle-related disorders, leads to the gradual decline and weakening of skeletal muscle over time. Recognizing the pivotal role of gastrointestinal conditions in maintaining metabolic homeostasis within skeletal muscle, we hypothesize that the effectiveness of the myogenic programme is influenced by the levels of gastrointestinal hormones in the bloodstream, and this connection is associated with the onset of sarcopenia. We first categorized 145 individuals from the Emergency Room of Taipei Veterans General Hospital into sarcopenia and non-sarcopenia groups, following the criteria established by the Asian Working Group for Sarcopenia. A thorough examination of specific gastrointestinal hormone levels in plasma was conducted to identify the one most closely associated with sarcopenia. Techniques, including immunofluorescence, western blotting, glucose uptake assays, seahorse real-time cell metabolic analysis, flow cytometry analysis, kinesin-1 activity assays and qPCR analysis, were applied to investigate its impacts and mechanisms on myogenic differentiation. Individuals in the sarcopenia group exhibited elevated plasma levels of glucagon-like peptide 1 (GLP-1) at 1021.5±313.5pg/mL, in contrast to non-sarcopenic individuals with levels at 351.1±39.0pg/mL (P<0.05). Although it is typical for GLP-1 levels to rise post-meal and subsequently drop naturally, detecting higher GLP-1 levels in starving individuals with sarcopenia raised the possibility of GLP-1 influencing myogenic differentiation in skeletal muscle. Further investigation using a cell model revealed that GLP-1 (1, 10 and 100ng/mL) dose-dependently suppressed the expression of the myogenic marker, impeding myocyte fusion and the formation of polarized myotubes during differentiation. GLP-1 significantly inhibited the activity of the microtubule motor kinesin-1, interfering with the translocation of glucose transporter 4 (GLUT4) to the cell membrane and the dispersion of mitochondria. These impairments subsequently led to a reduction in glucose uptake to 0.81±0.04 fold (P<0.01) and mitochondrial adenosine triphosphate (ATP) production from 25.24±1.57pmol/min to 18.83±1.11pmol/min (P<0.05). Continuous exposure to GLP-1, even under insulin induction, attenuated the elevated glucose uptake. The elevated GLP-1 levels observed in individuals with sarcopenia are associated with a reduction in myogenic differentiation. The impact of GLP-1 on both the membrane translocation of GLUT4 and the dispersion of mitochondria significantly hinders glucose uptake and the production of mitochondrial ATP necessary for the myogenic programme. These findings point us towards strategies to establish the muscle-gut axis, particularly in the context of sarcopenia. Additionally, these results present the potential of identifying relevant diagnostic biomarkers.

Xylooligosaccharides Enhance Lettuce Root Morphogenesis and Growth Dynamics.

Enhancing root development is pivotal for boosting crop yield and augmenting stress resilience. In this study, we explored the regulatory effects of xylooligosaccharides (XOSs) on lettuce root growth, comparing their impact with that of indole-3-butyric acid potassium salt (IBAP). Treatment with XOS led to a substantial increase in root dry weight (30.77%), total root length (29.40%), volume (21.58%), and surface area (25.44%) compared to the water-treated control. These enhancements were on par with those induced by IBAP. Comprehensive phytohormone profiling disclosed marked increases in indole-3-acetic acid (IAA), zeatin riboside (ZR), methyl jasmonate (JA-ME), and brassinosteroids (BRs) following XOS application. Through RNA sequencing, we identified 3807 differentially expressed genes (DEGs) in the roots of XOS-treated plants, which were significantly enriched in pathways associated with manganese ion homeostasis, microtubule motor activity, and carbohydrate metabolism. Intriguingly, approximately 62.7% of the DEGs responsive to XOS also responded to IBAP, underscoring common regulatory mechanisms. However, XOS uniquely influenced genes related to cutin, suberine, and wax biosynthesis, as well as plant hormone signal transduction, hinting at novel mechanisms of stress tolerance. Prominent up-regulation of genes encoding beta-glucosidase and beta-fructofuranosidase highlights enhanced carbohydrate metabolism as a key driver of XOS-induced root enhancement. Collectively, these results position XOS as a promising, sustainable option for agricultural biostimulation.

A de novo designed coiled coil-based switch regulates the microtubule motor kinesin-1

Many enzymes are allosterically regulated via conformational change; however, our ability to manipulate these structural changes and control function is limited. Here we install a conformational switch for allosteric activation into the kinesin-1 microtubule motor in vitro and in cells. Kinesin-1 is a heterotetramer that accesses open active and closed autoinhibited states. The equilibrium between these states centers on a flexible elbow within a complex coiled-coil architecture. We target the elbow to engineer a closed state that can be opened with a de novo designed peptide. The alternative states are modeled computationally and confirmed by biophysical measurements and electron microscopy. In cells, peptide-driven activation increases kinesin transport, demonstrating a primary role for conformational switching in regulating motor activity. The designs are enabled by our understanding of ubiquitous coiled-coil structures, opening possibilities for controlling other protein activities.

A novel role for KIFC1-MYH9 interaction in triple-negative breast cancer aggressiveness and racial disparity

African American (AA) women are twice as likely to develop triple-negative breast cancer (TNBC) as women of European descent. Additionally, AA women with TNBC present a much more aggressive disease course than their European American (EA) counterparts. Thus, there is an unmet clinical need to identify race-specific biomarkers and improve survival outcomes in AA patients with TNBC. The minus-end directed microtubule motor protein kinesin family member C1 (KIFC1) promotes centrosome clustering and chromosomal instability and is often overexpressed in TNBC. Previous findings suggest that KIFC1 plays a role in cell proliferation and migration in TNBC cells from AAs and that the levels of nuclear KIFC1 (nKIFC1) are particularly high in AA patients with TNBC. The nuclear localization of KIFC1 in interphase may underlie its previously unrecognized race-specific association. In this study, we found that in TNBC cells derived from AAs, nKIFC1 interacted with the tumor suppressor myosin heavy chain 9 (MYH9) over EA cells. Treatment of AA TNBC cells with commercial inhibitors of KIFC1 and MYH9 disrupted the interaction between KIFC1 and MYH9. To characterize the racial differences in the KIFC1-MYH9-MYC axis in TNBC, we established homozygous KIFC1 knockout (KO) TNBC cell lines. KIFC1 KO significantly inhibited proliferation, migration, and invasion in AA TNBC cells but not in EA TNBC cells. RNA sequencing analysis showed significant downregulation of genes involved in cell migration, invasion, and metastasis upon KIFC1 KO in TNBC cell lines from AAs compared to those from EAs. These data indicate that mechanistically, the role of nKIFC1 in driving TNBC progression and metastasis is stronger in AA patients than in EA patients, and that KIFC1 may be a critical therapeutic target for AA patients with TNBC.Graphical abstract

Cytoplasmic mRNA Transport: Adaptors of mRNA Binding to Microtubule Motor Proteins

Cytoplasmic mRNA Transport: Adaptors of mRNA Binding to Microtubule Motor Proteins

FLN-2 functions in parallel to linker of nucleoskeleton and cytoskeleton complexes and CDC-42/actin pathways during P-cell nuclear migration through constricted spaces in Caenorhabditis elegans.

Nuclear migration through narrow constrictions is important for development, metastasis, and proinflammatory responses. Studies performed in tissue culture cells have implicated linker of nucleoskeleton and cytoskeleton (LINC) complexes, microtubule motors, the actin cytoskeleton, and nuclear envelope repair machinery as important mediators of nuclear movements through constricted spaces. However, little is understood about how these mechanisms operate to move nuclei in vivo. In Caenorhabditis elegans larvae, six pairs of hypodermal P cells migrate from lateral to ventral positions through a constricted space between the body wall muscles and the cuticle. P-cell nuclear migration is mediated in part by LINC complexes using a microtubule-based pathway and by an independent CDC-42/actin-based pathway. However, when both LINC complex and actin-based pathways are knocked out, many nuclei still migrate, suggesting the existence of additional pathways. Here, we show that FLN-2 functions in a third pathway to mediate P-cell nuclear migration. The predicted N-terminal actin-binding domain in FLN-2 that is found in canonical filamins is dispensable for FLN-2 function; this and structural predictions suggest that FLN-2 does not function as a filamin. The immunoglobulin-like repeats 4-8 of FLN-2 were necessary for P-cell nuclear migration. Furthermore, in the absence of the LINC complex component unc-84, fln-2 mutants had an increase in P-cell nuclear rupture. We conclude that FLN-2 functions to maintain the integrity of the nuclear envelope in parallel with the LINC complex and CDC-42/actin-based pathways to move P-cell nuclei through constricted spaces.

Active nuclear positioning and actomyosin contractility maintain leader cell integrity during gonadogenesis

Proper distribution of organelles can play an important role in a moving cell's performance. During C.elegans gonad morphogenesis, the nucleus of the leading distal tip cell (DTC) is always found at the front, yet the significance of this localization is unknown. Here, we identified the molecular mechanism that keeps the nucleus at the front, despite a frictional force that pushes it backward. The Klarsicht/ANC-1/Syne homology (KASH) domain protein UNC-83 links the nucleus to the motor protein kinesin-1 that moves along a polarized acentrosomal microtubule network. Interestingly, disrupting nuclear positioning on its own did not affect gonad morphogenesis. However, reducing actomyosin contractility on top of nuclear mispositioning led to a dramatic phenotype: DTC splitting and gonad bifurcation. Long-term live imaging of the double knockdown revealed that, while the gonad attempted to perform a planned U-turn, the DTC was stretched due to the lagging nucleus until it fragmented into a nucleated cell and an enucleated cytoplast, each leading an independent gonadal arm. Remarkably, the enucleated cytoplast had polarity and invaded, but it could only temporarily support germ cell proliferation. Based on a qualitative biophysical model, we conclude that the leader cell employs two complementary mechanical approaches to preserve its integrity and ensure proper organ morphogenesis while navigating through a complex 3D environment: active nuclear positioning by microtubule motors and actomyosin-driven cortical contractility.