Muscle cofilin alters neuromuscular junction postsynaptic development to strengthen functional neurotransmission.

Cofilin, an actin-severing protein, plays key roles in muscle sarcomere addition and maintenance. Our previous work found that Drosophila cofilin (DmCFL) knockdown in muscle causes progressive deterioration of muscle structure and function and produces features seen in nemaline myopathy caused by cofilin mutations. We hypothesized that disruption of actin cytoskeleton dynamics by DmCFL knockdown would impact other aspects of muscle development, and, thus, conducted an RNA-sequencing analysis that unexpectedly revealed upregulated expression of numerous neuromuscular junction (NMJ) genes. We found that DmCFL is enriched in the muscle postsynaptic compartment and that DmCFL muscle knockdown causes F-actin disorganization in this subcellular domain prior to the sarcomere defects observed later in development. Despite NMJ gene expression changes, we found no significant changes in gross presynaptic Bruchpilot active zones or total postsynaptic glutamate receptor levels. However, DmCFL knockdown resulted in mislocalization of GluRIIA class glutamate receptors in more deteriorated muscles and strongly impaired NMJ transmission strength. These findings expand our understanding of the roles of cofilin in muscle to include NMJ structural development and suggest that NMJ defects may contribute to the pathophysiology of nemaline myopathy.

Slingshot homolog-1 amplifies mitochondrial abnormalities by distinctly impairing health and clearance of mitochondria.

Accumulating toxic protein assemblies, including Aß and tau, and dysfunctional mitochondria are associated with synaptic and neuronal loss in Alzheimer's disease (AD). Such accumulations are thought to be owing to clearance defects in the autophagy-lysosome pathway. Mitochondrial dysfunction is evident in AD brains and animal models at multiple levels, such as mitochondrial genomic mutations, disrupted bioenergetics, deregulated mitochondrial dynamics and impaired clearance of damaged mitochondria (mitophagy). Slingshot homolog-1 (SSH1) is a phosphatase activated by oxidative stress, high intracellular levels of Ca2+ and Aß42 oligomers (Aß42O), known for its function to dephosphorylate/activate cofilin through the N-terminal region. SSH1-mediated cofilin dephosphorylation results in Ab42O-induced severing of F-actin and translocation of cofilin to mitochondria, which promotes mitochondria-mediated apoptosis, synaptic loss and synaptic deficits. On the other hand, SSH1-mediated dephosphorylation/deactivation of the autophagy-cargo receptor p62 (SQSTM1), through its C-terminal region, inhibits p62 autophagy flux. However, the interplay between these two different activities of SSH1 in Aß42O-induced mitochondrial toxicity remains unclear. In this study, we assessed the role of endogenous SSH1 and different regions of SSH1 in regulating mitochondrial health, mitochondrial respiration, clearance of damaged mitochondria and synaptic integrity in vitro and in vivo. Our results indicate that SSH1 suppresses mitochondrial health and respiration through the cofilin-binding N-terminal region, whereas SSH1 impairs mitophagy through a newly identified ~ 100 residue p62-binding domain in the C-terminal region. These results indicate that both N-terminal and C-terminal regions negatively impact mitochondria by distinct and independent modalities to amplify mitochondrial abnormalities, making SSH1 an excellent target to mitigate AD pathogenesis.

Phosphatidylinositol 4-Kinase III Alpha Governs Cytoskeletal Organization for Invasiveness of Liver Cancer Cells.

BACKGROUND & AIMS: High expression of phosphatidylinositol 4-kinase III alpha (PI4KIIIα) correlates with poor survival rates in patients with hepatocellular carcinoma. In addition, hepatitis C virus (HCV) infections activate PI4KIIIα and contribute to hepatocellular carcinoma progression. We aimed at mechanistically understanding the impact of PI4KIIIα on the progression of liver cancer and the potential contribution of HCV in this process. METHODS: Several hepatic cell culture and mouse models were used to study the functional importance of PI4KIIIα on liver pathogenesis. Antibody arrays, gene silencing, and PI4KIIIα-specific inhibitor were applied to identify the involved signaling pathways. The contribution of HCV was examined by using HCV infection or overexpression of its nonstructural protein. RESULTS: High PI4KIIIα expression and/or activity induced cytoskeletal rearrangements via increased phosphorylation of paxillin and cofilin. This led to morphologic alterations and higher migratory and invasive properties of liver cancer cells. We further identified the liver-specific lipid kinase phosphatidylinositol 3-kinase C2 domain-containing subunit gamma (PIK3C2γ) working downstream of PI4KIIIα in regulation of the cytoskeleton. PIK3C2γ generates plasma membrane phosphatidylinositol 3,4-bisphosphate-enriched, invadopodia-like structures that regulate cytoskeletal reorganization by promoting Akt2 phosphorylation. CONCLUSIONS: PI4KIIIα regulates cytoskeleton organization via PIK3C2γ/Akt2/paxillin-cofilin to favor migration and invasion of liver cancer cells. These findings provide mechanistic insight into the contribution of PI4KIIIα and HCV to the progression of liver cancer and identify promising targets for therapeutic intervention.

Forgetting is regulated through Rac activity in Drosophila.

Initially acquired memory dissipates rapidly if not consolidated. Such memory decay is thought to result either from the inherently labile nature of newly acquired memories or from interference by subsequently attained information. Here we report that a small G protein Rac-dependent forgetting mechanism contributes to both passive memory decay and interference-induced forgetting in Drosophila. Inhibition of Rac activity leads to slower decay of early memory, extending it from a few hours to more than one day, and to blockade of interference-induced forgetting. Conversely, elevated Rac activity in mushroom body neurons accelerates memory decay. This forgetting mechanism does not affect memory acquisition and is independent of Rutabaga adenylyl cyclase-mediated memory formation mechanisms. Endogenous Rac activation is evoked on different time scales during gradual memory loss in passive decay and during acute memory removal in reversal learning. We suggest that Rac's role in actin cytoskeleton remodeling may contribute to memory erasure.

Cyclase-associated protein interacts with actin filament barbed ends to promote depolymerization and formin displacement.

Cyclase-associated protein (CAP) has emerged as a central player in cellular actin turnover, but its molecular mechanisms of action are not yet fully understood. Recent studies revealed that the N terminus of CAP interacts with the pointed ends of actin filaments to accelerate depolymerization in conjunction with cofilin. Here, we use in vitro microfluidics-assisted TIRF microscopy to show that the C terminus of CAP promotes depolymerization at the opposite (barbed) ends of actin filaments. In the absence of actin monomers, full-length mouse CAP1 and C-terminal halves of CAP1 (C-CAP1) and CAP2 (C-CAP2) accelerate barbed end depolymerization. Using mutagenesis and structural modeling, we show that these activities are mediated by the WH2 and CARP domains of CAP. In addition, we observe that CAP collaborates with profilin to accelerate barbed end depolymerization and that these effects depend on their direct interaction, providing the first known example of CAP-profilin collaborative effects in regulating actin. In the presence of actin monomers, CAP1 attenuates barbed end growth and promotes formin dissociation. Overall, these findings demonstrate that CAP uses distinct domains and mechanisms to interact with opposite ends of actin filaments and drive turnover. Further, they contribute to the emerging view of actin barbed ends as sites of dynamic molecular regulation, where numerous proteins compete and cooperate with each other to tune polymer dynamics, similar to the rich complexity seen at microtubule ends.

Cofilin regulates actin network homeostasis and microvilli length in mouse oocytes.

How multiple actin networks coexist in a common cytoplasm while competing for a shared pool of monomers is still an ongoing question. This is exemplified by meiotic maturation in the mouse oocyte, which relies on the dynamic remodeling of distinct cortical and cytoplasmic F-actin networks. Here, we show that the conserved actin-depolymerizing factor cofilin is activated in a switch-like manner upon meiosis resumption from prophase arrest. Interfering with cofilin activation during maturation resulted in widespread elongation of microvilli, while cytoplasmic F-actin was depleted, leading to defects in spindle migration and polar body extrusion. In contrast, cofilin inactivation in metaphase II-arrested oocytes resulted in a shutdown of F-actin dynamics, along with a dramatic overgrowth of the polarized actin cap. However, inhibition of the Arp2/3 complex to promote actin cap disassembly elicited ectopic microvilli outgrowth in the polarized cortex. These data establish cofilin as a key player in actin network homeostasis in oocytes and reveal that microvilli can act as a sink for monomers upon disassembly of a competing network.

Insights into the evolution of regulated actin dynamics via characterization of primitive gelsolin/cofilin proteins from Asgard archaea.

Asgard archaea genomes contain potential eukaryotic-like genes that provide intriguing insight for the evolution of eukaryotes. The eukaryotic actin polymerization/depolymerization cycle is critical for providing force and structure in many processes, including membrane remodeling. In general, Asgard genomes encode two classes of actin-regulating proteins from sequence analysis, profilins and gelsolins. Asgard profilins were demonstrated to regulate actin filament nucleation. Here, we identify actin filament severing, capping, annealing and bundling, and monomer sequestration activities by gelsolin proteins from Thorarchaeota (Thor), which complete a eukaryotic-like actin depolymerization cycle, and indicate complex actin cytoskeleton regulation in Asgard organisms. Thor gelsolins have homologs in other Asgard archaea and comprise one or two copies of the prototypical gelsolin domain. This appears to be a record of an initial preeukaryotic gene duplication event, since eukaryotic gelsolins are generally comprise three to six domains. X-ray structures of these proteins in complex with mammalian actin revealed similar interactions to the first domain of human gelsolin or cofilin with actin. Asgard two-domain, but not one-domain, gelsolins contain calcium-binding sites, which is manifested in calcium-controlled activities. Expression of two-domain gelsolins in mammalian cells enhanced actin filament disassembly on ionomycin-triggered calcium release. This functional demonstration, at the cellular level, provides evidence for a calcium-controlled Asgard actin cytoskeleton, indicating that the calcium-regulated actin cytoskeleton predates eukaryotes. In eukaryotes, dynamic bundled actin filaments are responsible for shaping filopodia and microvilli. By correlation, we hypothesize that the formation of the protrusions observed from Lokiarchaeota cell bodies may involve the gelsolin-regulated actin structures.

Actin Depolymerization Factor ADF1 Regulated by MYB30 Plays an Important Role in Plant Thermal Adaptation.

Actin filaments are essential for plant adaptation to high temperatures. However, the molecular mechanisms of actin filaments in plant thermal adaptation remain unclear. Here, we found that the expression of Arabidopsis actin depolymerization factor 1 (AtADF1) was repressed by high temperatures. Compared with wild-type seedlings (WT), the mutation of AtADF1 and the overexpression of AtADF1 led to promoted and inhibited plant growth under high temperature conditions, respectively. Further, high temperatures induced the stability of actin filaments in plants. Compared with WT, Atadf1-1 mutant seedlings showed more stability of actin filaments under normal and high temperature conditions, while the AtADF1 overexpression seedlings showed the opposite results. Additionally, AtMYB30 directly bound to the promoter of AtADF1 at a known AtMYB30 binding site, AACAAAC, and promoted the transcription of AtADF1 under high temperature treatments. Genetic analysis further indicated that AtMYB30 regulated AtADF1 under high temperature treatments. Chinese cabbage ADF1 (BrADF1) was highly homologous with AtADF1. The expression of BrADF1 was inhibited by high temperatures. BrADF1 overexpression inhibited plant growth and reduced the percentage of actin cable and the average length of actin filaments in Arabidopsis, which were similar to those of AtADF1 overexpression seedlings. AtADF1 and BrADF1 also affected the expression of some key heat response genes. In conclusion, our results indicate that ADF1 plays an important role in plant thermal adaptation by blocking the high-temperature-induced stability of actin filaments and is directly regulated by MYB30.

Rab23/Kif17 regulate meiotic progression in oocytes by modulating tubulin acetylation and actin dynamics.

Cytoskeletal dynamics are involved in multiple cellular processes during oocyte meiosis, including spindle organization, actin-based spindle migration and polar body extrusion. Here, we report that the vesicle trafficking protein Rab23, a GTPase, drives the motor protein Kif17, and that this is important for spindle organization and actin dynamics during mouse oocyte meiosis. GTP-bound Rab23 accumulated at the spindle and promoted migration of Kif17 to the spindle poles. Depletion of Rab23 or Kif17 caused polar body extrusion failure. Further analysis showed that depletion of Rab23/Kif17 perturbed spindle formation and chromosome alignment, possibly by affecting tubulin acetylation. Kif17 regulated tubulin acetylation by associating with αTAT and Sirt2, and depletion of Kif17 altered expression of these proteins. Moreover, depletion of Kif17 decreased the level of cytoplasmic actin, which abrogated spindle migration to the cortex. The tail domain of Kif17 associated with constituents of the RhoA-ROCK-LIMK-cofilin pathway to modulate assembly of actin filaments. Taken together, our results demonstrate that the Rab23-Kif17-cargo complex regulates tubulin acetylation for spindle organization and drives actin-mediated spindle migration during meiosis.

Nucleolin regulates 14-3-3ζ mRNA and promotes cofilin phosphorylation to induce tunneling nanotube formation.

Tunneling nanotubes (TNTs) mediate intercellular communication between animal cells in health and disease, but the mechanisms of their biogenesis and function are poorly understood. Here we report that the RNA-binding protein (RBP) nucleolin, which interacts with the known TNT-inducing protein MSec, is essential for TNT formation in mammalian cells. Nucleolin, through its RNA-binding domains (RBDs), binds to and maintains the cytosolic levels of 14-3-3ζ mRNA, and is, therefore, required for TNT formation. A specific region of the 3'-untranslated region (UTR) of the 14-3-3ζ mRNA is likely to be involved in its regulation by nucleolin. Functional complementation experiments suggest that nucleolin and 14-3-3ζ form a linear signaling axis that promotes the phosphorylation and inactivation of the F-actin depolymerization factor cofilin to induce TNT formation. MSec also similarly inactivates cofilin, but potentiates TNT formation independent of the nucleolin-14-3-3ζ axis, despite biochemically interacting with both proteins. We show that 14-3-3ζ and nucleolin are required for the formation of TNTs between primary mouse neurons and astrocytes and in multiple other mammalian cell types. We also report that the Caenorhabditis elegans orthologs of 14-3-3ζ and MSec regulate the size and architecture of the TNT-like cellular protrusions of the distal tip cell (DTC), the germline stem cell niche in the gonad. Our study demonstrates a novel and potentially conserved mRNA-guided mechanism of TNT formation through the maintenance of cellular 14-3-3ζ mRNA levels by the RBP nucleolin.

Arabidopsis ADF1 is Regulated by MYB73 and is Involved in Response to Salt Stress Affecting Actin Filament Organization.

Actin cytoskeleton and transcription factors play key roles in plant response to salt stress; however, little is known about the link between the two regulators in response to salt stress. Actin-depolymerizing factors (ADFs) are conserved actin-binding proteins in eukaryotes. Here, we revealed that the expression level of ADF1 was induced by salt stress. The adf1 mutants showed significantly reduced survival rate, increased percentage of actin cable and reduced density of actin filaments, while ADF1 overexpression seedlings displayed the opposite results when compared with WT under the same condition. Furthermore, biochemical assays revealed that MYB73, a R2R3 MYB transcription factor, binds to the promoter of ADF1 and represses its expression via the MYB-binding site core motif ACCTAC. Taken together, our results indicate that ADF1 participates in salt stress by regulating actin organization and may also serve as a potential downstream target of MYB73, which is a negative regulator of salt stress.

Arabidopsis thaliana subclass I ACTIN DEPOLYMERIZING FACTORs and vegetative ACTIN2/8 are novel regulators of endoreplication.

Endoreplication is a type of cell cycle where genome replication occurs without mitosis. An increase of ploidy level by endoreplication is often associated with cell enlargement and an enhanced plant growth. Here we report Arabidopsis thaliana subclass I ACTIN DEPOLYMERIZING FACTORs (ADFs) and vegetative ACTIN2/8 as novel regulators of endoreplication. A. thaliana has 11 ADF members that are divided into 4 subclasses. Subclass I consists of four members, ADF1, -2, -3, and -4, all of which constitutively express in various tissues. We found that both adf4 knockout mutant and transgenic plants in which expressions of all of four subclass I ADFs are suppressed (ADF1-4Ri) showed an increased leaf area of mature first leaves, which was associated with a significant increase of epidermal pavement cell area. Ploidy analysis revealed that the ploidy level was significantly increased in mature leaves of ADF1-4Ri. The increased ploidy was also observed in roots of adf4 and ADF1-4Ri, as well as in dark-grown hypocotyls of adf4. Furthermore, double mutants of vegetative ACT2 and ACT8 (act2/8) exhibited an increase of leaf area and ploidy level in mature leaves. Therefore, actin-relating pathway could regulate endoreplication. The possible mechanisms that actin and ADFs regulate endoreplication are discussed.

Acidic microenvironment enhances MT1-MMP-mediated cancer cell motility through integrin ß1/cofilin/F-actin axis.

Tumor acidic microenvironment is the main feature of many solid tumors. As a part of the tumor microenvironment, it has a profound impact on the occurrence and development of tumors. However, the research on how tumor cells sense the changes of the external microenvironment and how the intracellular subcellular structures transmit the signals from extracellular to intracellular is unclear. In this study, we identify that the acidic microenvironment enhances cancer cell motility, and the expression of membrane-anchored membrane type 1-matrix metalloproteinase is also associated with cell motility, which indicates more degradation of the ECM under the acidic microenvironment. Moreover, the expression of cofilin is low in the acidic microenvironment, and the F-actin filaments are distributed more along the cells. The cytoskeletal F-actin changes are consistent with the potential of a high-invasive phenotype. Further study reveals the upstream control of the signal transductions from extracellular to intracellular, that is, the integrin ß1 functions to trigger the biological responses under the acidic microenvironment. Our results demonstrate that the acidic microenvironment enhances cancer cell motility through the integrin ß1/cofilin/F-actin signal axis. This study clearly shows the scheme of the signal transmissions from extracellular to intracellular and further reveals the cytoskeletal roles for the contributions of cancer cell motility under acidic microenvironment, which provides new targets for cancer intervention from the biochemical and biophysical perspectives.

Endoglin Is an Endothelial Housekeeper against Inflammation: Insight in ECFC-Related Permeability through LIMK/Cofilin Pathway.

Endoglin (Eng) is an endothelial cell (EC) transmembrane glycoprotein involved in adhesion and angiogenesis. Eng mutations result in vessel abnormalities as observed in hereditary hemorrhagic telangiectasia of type 1. The role of Eng was investigated in endothelial functions and permeability under inflammatory conditions, focusing on the actin dynamic signaling pathway. Endothelial Colony-Forming Cells (ECFC) from human cord blood and mouse lung/aortic EC (MLEC, MAEC) from Eng+/+ and Eng+/- mice were used. ECFC silenced for Eng with Eng-siRNA and ctr-siRNA were used to test tubulogenesis and permeability +/- TNFα and +/- LIM kinase inhibitors (LIMKi). In silico modeling of TNFα-Eng interactions was carried out from PDB IDs 5HZW and 5HZV. Calcium ions (Ca2+) flux was studied by Oregon Green 488 in epifluorescence microscopy. Levels of cofilin phosphorylation and tubulin post-translational modifications were evaluated by Western blot. F-actin and actin-tubulin distribution/co-localization were evaluated in cells by confocal microscopy. Eng silencing in ECFCs resulted in a decrease of cell sprouting by 50 ± 15% (p < 0.05) and an increase in pseudo-tube width (41 ± 4.5%; p < 0.001) compared to control. Upon TNFα stimulation, ECFC Eng-siRNA displayed a significant higher permeability compared to ctr-siRNA (p < 0.01), which is associated to a higher Ca2+ mobilization (p < 0.01). Computational analysis suggested that Eng mitigated TNFα activity. F-actin polymerization was significantly increased in ECFC Eng-siRNA, MAEC+/-, and MLEC+/- compared to controls (p < 0.001, p < 0.01, and p < 0.01, respectively) as well as actin/tubulin distribution (p < 0.01). Furthermore, the inactive form of cofilin (P-cofilin at Ser3) was significantly decreased by 36.7 ± 4.8% in ECFC Eng-siRNA compared to ctr-siRNA (p < 0.001). Interestingly, LIMKi reproduced the absence of Eng on TNFα-induced ECFC-increased permeability. Our data suggest that Eng plays a critical role in the homeostasis regulation of endothelial cells under inflammatory conditions (TNFα), and loss of Eng influences ECFC-related permeability through the LIMK/cofilin/actin rearrangement-signaling pathway.

Sarcomeres Morphology and Z-Line Arrangement Disarray Induced by Ventricular Premature Contractions through the Rac2/Cofilin Pathway.

The most common ventricular premature contractions (VPCs) originate from the right ventricular outflow tract (RVOT), but the molecular mechanisms of altered cytoskeletons of VPC-induced cardiomyopathy remain unexplored. We created a RVOT bigeminy VPC pig model (n = 6 in each group). Echocardiography was performed. The histopathological alternations in the LV myocardium were analyzed, and next generation sequencing (NGS) and functional enrichment analyses were employed to identify the differentially expressed genes (DEGs) responsible for the histopathological alternations. Finally, a cell silencing model was used to confirm the key regulatory gene and pathway. VPC pigs had increased LV diameters in the 6-month follow-up period. A histological study showed more actin cytoskeleton disorganization and actin accumulation over intercalated disc, Z-line arrangement disarray, increased ß-catenin expression, and cardiomyocyte enlargement in the LV myocardium of the VPC pigs compared to the control pigs. The NGS study showed actin cytoskeleton signaling, RhoGDI signaling, and signaling by Rho Family GTPases and ILK Signaling presented z-scores with same activation states. The expressions of Rac family small GTPase 2 (Rac2), the p-cofilin/cofilin ratio, and the F-actin/G-actin ratio were downregulated in the VPC group compared to the control group. Moreover, the intensity and number of actin filaments per cardiomyocyte were significantly decreased by Rac2 siRNA in the cell silencing model. Therefore, the Rac2/cofilin pathway was found to play a crucial role in the sarcomere morphology and Z-line arrangement disarray induced by RVOT bigeminy VPCs.

Cofilin Signaling in the CNS Physiology and Neurodegeneration.

All eukaryotic cells are composed of the cytoskeleton, which plays crucial roles in coordinating diverse cellular functions such as cell division, morphology, migration, macromolecular stabilization, and protein trafficking. The cytoskeleton consists of microtubules, intermediate filaments, and actin filaments. Cofilin, an actin-depolymerizing protein, is indispensable for regulating actin dynamics in the central nervous system (CNS) development and function. Cofilin activities are spatiotemporally orchestrated by numerous extra- and intra-cellular factors. Phosphorylation at Ser-3 by kinases attenuate cofilin's actin-binding activity. In contrast, dephosphorylation at Ser-3 enhances cofilin-induced actin depolymerization. Cofilin functions are also modulated by various binding partners or reactive oxygen species. Although the mechanism of cofilin-mediated actin dynamics has been known for decades, recent research works are unveiling the profound impacts of cofilin dysregulation in neurodegenerative pathophysiology. For instance, oxidative stress-induced increase in cofilin dephosphorylation is linked to the accumulation of tau tangles and amyloid-beta plaques in Alzheimer's disease. In Parkinson's disease, cofilin activation by silencing its upstream kinases increases α-synuclein-fibril entry into the cell. This review describes the molecular mechanism of cofilin-mediated actin dynamics and provides an overview of cofilin's importance in CNS physiology and pathophysiology.

The Cofilin/Limk1 Pathway Controls the Growth Rate of Both Developing and Regenerating Motor Axons.

Regenerating axons often have to grow considerable distances to reestablish circuits, making functional recovery a lengthy process. One solution to this problem would be to co-opt the "temporal" guidance mechanisms that control the rate of axon growth during development to accelerate the rate at which nerves regenerate in adults. We have previously found that the loss of Limk1, a negative regulator of cofilin, accelerates the rate of spinal commissural axon growth. Here, we use mouse models to show that spinal motor axon outgrowth is similarly promoted by the loss of Limk1, suggesting that temporal guidance mechanisms are widely used during development. Furthermore, we find that the regulation of cofilin activity is an acute response to nerve injury in the peripheral nervous system. Within hours of a sciatic nerve injury, the level of phosphorylated cofilin dramatically increases at the lesion site, in a Limk1-dependent manner. This response may be a major constraint on the rate of peripheral nerve regeneration. Proof-of-principle experiments show that elevating cofilin activity, through the loss of Limk1, results in faster sciatic nerve growth, and improved recovery of some sensory and motor function.SIGNIFICANCE STATEMENT The studies shed light on an endogenous, shared mechanism that controls the rate at which developing and regenerating axons grow. An understanding of these mechanisms is key for developing therapies to reduce painful recovery times for nerve-injury patients, by accelerating the rate at which damaged nerves reconnect with their synaptic targets.

Endothelin type B receptor promotes cofilin rod formation and dendritic loss in neurons by inducing oxidative stress and cofilin activation.

Endothelin-1 (ET-1) is a neuroactive peptide produced by neurons, reactive astrocytes, and endothelial cells in the brain. Elevated levels of ET-1 have been detected in the post-mortem brains of individuals with Alzheimer's disease (AD). We have previously demonstrated that overexpression of astrocytic ET-1 exacerbates memory deficits in aged mice or in APPK670/M671 mutant mice. However, the effects of ET-1 on neuronal dysfunction remain elusive. ET-1 has been reported to mediate superoxide formation in the vascular system via NADPH oxidase (NOX) and to regulate the actin cytoskeleton of cancer cell lines via the cofilin pathway. Interestingly, oxidative stress and cofilin activation were both reported to mediate one of the AD histopathologies, cofilin rod formation in neurons. This raises the possibility that ET-1 mediates neurodegeneration via oxidative stress- or cofilin activation-driven cofilin rod formation. Here, we demonstrate that exposure to 100 nm ET-1 or to a selective ET type B receptor (ETB) agonist (IRL1620) induces cofilin rod formation in dendrites of primary hippocampal neurons, accompanied by a loss of distal dendrites and a reduction in dendritic length. The 100 nm IRL1620 exposure induced superoxide formation and cofilin activation, which were abolished by pretreatment with a NOX inhibitor (5 µm VAS2870). Moreover, IRL1620-induced cofilin rod formation was partially abolished by pretreatment with a calcineurin inhibitor (100 nm FK506), which suppressed cofilin activation. In conclusion, our findings suggest a role for ETB in neurodegeneration by promoting cofilin rod formation and dendritic loss via NOX-driven superoxide formation and cofilin activation.

Linkage mapping and genome-wide association study reveals conservative QTL and candidate genes for Fusarium rot resistance in maize.

BACKGROUND: Fusarium ear rot (FER) caused by Fusarium verticillioides is a major disease of maize that reduces grain yield and quality globally. However, there have been few reports of major loci for FER were verified and cloned. RESULT: To gain a comprehensive understanding of the genetic basis of natural variation in FER resistance, a recombinant inbred lines (RIL) population and one panel of inbred lines were used to map quantitative trait loci (QTL) for resistance. As a result, a total of 10 QTL were identified by linkage mapping under four environments, which were located on six chromosomes and explained 1.0-7.1% of the phenotypic variation. Epistatic mapping detected four pairs of QTL that showed significant epistasis effects, explaining 2.1-3.0% of the phenotypic variation. Additionally, 18 single nucleotide polymorphisms (SNPs) were identified across the whole genome by genome-wide association study (GWAS) under five environments. Compared linkage and association mapping revealed five common intervals located on chromosomes 3, 4, and 5 associated with FER resistance, four of which were verified in different near-isogenic lines (NILs) populations. GWAS identified three candidate genes in these consistent intervals, which belonged to the Glutaredoxin protein family, actin-depolymerizing factors (ADFs), and AMP-binding proteins. In addition, two verified FER QTL regions were found consistent with Fusarium cob rot (FCR) and Fusarium seed rot (FSR). CONCLUSIONS: These results revealed that multi pathways were involved in FER resistance, which was a complex trait that was controlled by multiple genes with minor effects, and provided important QTL and genes, which could be used in molecular breeding for resistance.

NCAM regulates the proliferation, apoptosis, autophagy, EMT, and migration of human melanoma cells via the Src/Akt/mTOR/cofilin signaling pathway.

The neural cell adhesion molecule (NCAM) plays critical roles in multiple cellular processes in neural cells, mesenchymal stem cells, and various cancer cells. However, the effect and mechanism of NCAM in human melanoma cells are still unclear. In this study, we found that NCAM regulated the proliferation, apoptosis, autophagy, migration, and epithelial-to-mesenchymal transition of human melanoma cells by determining the biological behavior of NCAM knockdown A375 and M102 human melanoma cells. Further studies revealed that NCAM knockdown impaired the organization of actin cytoskeleton and reduced the phosphorylation of cofilin, an actin-cleaving protein. When cells were transfected with cofilin S3A (dephosphorylated cofilin), biological behavior similar to that of NCAM knockdown cells was observed. Research on the underlying molecular mechanism showed that NCAM knockdown suppressed activation of the Src/Akt/mTOR pathway. Specific inhibitors of Src and PI3K/Akt were employed to further verify the relationship between Src/Akt/mTOR signaling and cofilin, and the results showed that the phosphorylation level of cofilin decreased following inhibition of the Src/Akt/mTOR pathway. These results indicated that NCAM may regulate the proliferation, apoptosis, autophagy, migration, and epithelial-to-mesenchymal transition of human melanoma cells via the Src/Akt/mTOR/cofilin pathway-mediated dynamics of actin cytoskeleton.