Fibroblast growth factor-induced lens fiber cell elongation is driven by the stepwise activity of Rho and Rac.

The spheroidal shape of the eye lens is crucial for precise light focusing onto the retina. This shape is determined by concentrically aligned, convexly elongated lens fiber cells along the anterior and posterior axis of the lens. Upon differentiation at the lens equator, the fiber cells increase in height as their apical and basal tips migrate towards the anterior and posterior poles, respectively. The forces driving this elongation and migration remain unclear. We found that, in the mouse lens, membrane protrusions or lamellipodia are observed only in the maturing fibers undergoing cell curve conversion, indicating that lamellipodium formation is not the primary driver of earlier fiber migration. We demonstrated that elevated levels of fibroblast growth factor (FGF) suppressed the extension of Rac-dependent protrusions, suggesting changes in the activity of FGF controlling Rac activity, switching to lamellipodium-driven migration. Inhibitors of ROCK, myosin and actin reduced the height of both early and later fibers, indicating that elongation of these fibers relies on actomyosin contractility. Consistent with this, active RhoA was detected throughout these fibers. Given that FGF promotes fiber elongation, we propose that it does so through regulation of Rho activity.

Early Scanning of Nascent Polypeptides inside the Ribosomal Tunnel by NAC.

Cotranslational processing of newly synthesized proteins is fundamental for correct protein maturation. Protein biogenesis factors are thought to bind nascent polypeptides not before they exit the ribosomal tunnel. Here, we identify a nascent chain recognition mechanism deep inside the ribosomal tunnel by an essential eukaryotic cytosolic chaperone. The nascent polypeptide-associated complex (NAC) inserts the N-terminal tail of its ß subunit (N-ßNAC) into the ribosomal tunnel to sense substrates directly upon synthesis close to the peptidyl-transferase center. N-ßNAC escorts the growing polypeptide to the cytosol and relocates to an alternate binding site on the ribosomal surface. Using C. elegans as an in vivo model, we demonstrate that the tunnel-probing activity of NAC is essential for organismal viability and critical to regulate endoplasmic reticulum (ER) protein transport by controlling ribosome-Sec61 translocon interactions. Thus, eukaryotic protein maturation relies on the early sampling of nascent chains inside the ribosomal tunnel.

Ontogeny and function of the circadian clock in intestinal organoids.

Circadian rhythms regulate diverse aspects of gastrointestinal physiology ranging from the composition of microbiota to motility. However, development of the intestinal circadian clock and detailed mechanisms regulating circadian physiology of the intestine remain largely unknown. In this report, we show that both pluripotent stem cell-derived human intestinal organoids engrafted into mice and patient-derived human intestinal enteroids possess circadian rhythms and demonstrate circadian phase-dependent necrotic cell death responses to Clostridium difficile toxin B (TcdB). Intriguingly, mouse and human enteroids demonstrate anti-phasic necrotic cell death responses to TcdB. RNA-Seq analysis shows that ~3-10% of the detectable transcripts are rhythmically expressed in mouse and human enteroids. Remarkably, we observe anti-phasic gene expression of Rac1, a small GTPase directly inactivated by TcdB, between mouse and human enteroids, and disruption of Rac1 abolishes clock-dependent necrotic cell death responses. Our findings uncover robust functions of circadian rhythms regulating clock-controlled genes in both mouse and human enteroids governing organism-specific, circadian phase-dependent necrotic cell death responses, and lay a foundation for human organ- and disease-specific investigation of clock functions using human organoids for translational applications.

Convergence of Ras- and Rac-regulated formin pathways is pivotal for phagosome formation and particle uptake in Dictyostelium.

Macroendocytosis comprising phagocytosis and macropinocytosis is an actin-driven process regulated by small GTPases that depend on the dynamic reorganization of the membrane that protrudes and internalizes extracellular material by cup-shaped structures. To effectively capture, enwrap, and internalize their targets, these cups are arranged into a peripheral ring or ruffle of protruding actin sheets emerging from an actin-rich, nonprotrusive zone at its base. Despite extensive knowledge of the mechanism driving actin assembly of the branched network at the protrusive cup edge, which is initiated by the actin-related protein (Arp) 2/3 complex downstream of Rac signaling, our understanding of actin assembly in the base is still incomplete. In the Dictyostelium model system, the Ras-regulated formin ForG was previously shown to specifically contribute to actin assembly at the cup base. Loss of ForG is associated with a strongly impaired macroendocytosis and a 50% reduction in F-actin content at the base of phagocytic cups, in turn indicating the presence of additional factors that specifically contribute to actin formation at the base. Here, we show that ForG synergizes with the Rac-regulated formin ForB to form the bulk of linear filaments at the cup base. Consistently, combined loss of both formins virtually abolishes cup formation and leads to severe defects of macroendocytosis, emphasizing the relevance of converging Ras- and Rac-regulated formin pathways in assembly of linear filaments in the cup base, which apparently provide mechanical support to the entire structure. Remarkably, we finally show that active ForB, unlike ForG, additionally drives phagosome rocketing to aid particle internalization.

A TIAM1-TRIM28 complex mediates epigenetic silencing of protocadherins to promote migration of lung cancer cells.

Lung cancer is the leading cause of cancer deaths. Its high mortality is associated with high metastatic potential. Here, we show that the RAC1-selective guanine nucleotide exchange factor T cell invasion and metastasis-inducing protein 1 (TIAM1) promotes cell migration and invasion in the most common subtype of lung cancer, non-small-cell lung cancer (NSCLC), through an unexpected nuclear function. We show that TIAM1 interacts with TRIM28, a master regulator of gene expression, in the nucleus of NSCLC cells. We reveal that a TIAM1-TRIM28 complex promotes epithelial-to-mesenchymal transition, a phenotypic switch implicated in cell migration and invasion. This occurs through H3K9me3-induced silencing of protocadherins and by decreasing E-cadherin expression, thereby antagonizing cell-cell adhesion. Consistently, TIAM1 or TRIM28 depletion suppresses the migration of NSCLC cells, while migration is restored by the simultaneous depletion of protocadherins. Importantly, high nuclear TIAM1 in clinical specimens is associated with advanced-stage lung adenocarcinoma, decreased patient survival, and inversely correlates with E-cadherin expression.

Hyperactive Rac stimulates cannibalism of living target cells and enhances CAR-M-mediated cancer cell killing.

The 21kD GTPase Rac is an evolutionarily ancient regulator of cell shape and behavior. Rac2 is predominantly expressed in hematopoietic cells where it is essential for survival and motility. The hyperactivating mutation Rac2E62K also causes human immunodeficiency, although the mechanism remains unexplained. Here, we report that in Drosophila, hyperactivating Rac stimulates ovarian cells to cannibalize neighboring cells, destroying the tissue. We then show that hyperactive Rac2E62K stimulates human HL60-derived macrophage-like cells to engulf and kill living T cell leukemia cells. Primary mouse Rac2+/E62K bone-marrow-derived macrophages also cannibalize primary Rac2+/E62K T cells due to a combination of macrophage hyperactivity and T cell hypersensitivity to engulfment. Additionally, Rac2+/E62K macrophages non-autonomously stimulate wild-type macrophages to engulf T cells. Rac2E62K also enhances engulfment of target cancer cells by chimeric antigen receptor-expressing macrophages (CAR-M) in a CAR-dependent manner. We propose that Rac-mediated cell cannibalism may contribute to Rac2+/E62K human immunodeficiency and enhance CAR-M cancer immunotherapy.

Tiam1 methylation by NSD2 promotes Rac1 signaling activation and colon cancer metastasis.

Metastasis is a major cause of cancer therapy failure and mortality. However, targeting metastatic seeding and colonization remains a significant challenge. In this study, we identified NSD2, a histone methyltransferase responsible for dimethylating histone 3 at lysine 36, as being overexpressed in metastatic tumors. Our findings suggest that NSD2 overexpression enhances tumor metastasis both in vitro and in vivo. Further analysis revealed that NSD2 promotes tumor metastasis by activating Rac1 signaling. Mechanistically, NSD2 combines with and activates Tiam1 (T lymphoma invasion and metastasis 1) and promotes Rac1 signaling by methylating Tiam1 at K724. In vivo and in vitro studies revealed that Tiam1 K724 methylation could be a predictive factor for cancer prognosis and a potential target for metastasis inhibition. Furthermore, we have developed inhibitory peptide which was proved to inhibit tumor metastasis through blocking the interaction between NSD2 and Tiam1. Our results demonstrate that NSD2-methylated Tiam1 promotes Rac1 signaling and cancer metastasis. These results provide insights into the inhibition of tumor metastasis.

Conditional Astrocyte Rac1KO Attenuates Hyperreflexia after Spinal Cord Injury.

Spasticity is a hyperexcitability disorder that adversely impacts functional recovery and rehabilitative efforts after spinal cord injury (SCI). The loss of evoked rate-dependent depression (RDD) of the monosynaptic H-reflex is indicative of hyperreflexia, a physiological sign of spasticity. Given the intimate relationship between astrocytes and neurons, that is, the tripartite synapse, we hypothesized that astrocytes might have a significant role in post-injury hyperreflexia and plasticity of neighboring neuronal synaptic dendritic spines. Here, we investigated the effect of selective Rac1KO in astrocytes (i.e., adult male and female mice, transgenic cre-flox system) on SCI-induced spasticity. Three weeks after a mild contusion SCI, control Rac1wt animals displayed a loss of H-reflex RDD, that is, hyperreflexia. In contrast, transgenic animals with astrocytic Rac1KO demonstrated near-normal H-reflex RDD similar to pre-injury levels. Reduced hyperreflexia in astrocytic Rac1KO animals was accompanied by a loss of thin-shaped dendritic spine density on α-motor neurons in the ventral horn. In SCI-Rac1wt animals, as expected, we observed the development of dendritic spine dysgenesis on α-motor neurons associated with spasticity. As compared with WT animals, SCI animals with astrocytic Rac1KO expressed increased levels of the glial-specific glutamate transporter, glutamate transporter-1 in the ventral spinal cord, potentially enhancing glutamate clearance from the synaptic cleft and reducing hyperreflexia in astrocytic Rac1KO animals. Taken together, our findings show for the first time that Rac1 activity in astrocytes can contribute to hyperreflexia underlying spasticity following SCI. These results reveal an opportunity to target cell-specific molecular regulators of H-reflex excitability to manage spasticity after SCI.Significance Statement Spinal cord injury leads to stretch reflex hyperexcitability, which underlies the clinical symptom of spasticity. This study shows for the first time that astrocytic Rac1 contributes to the development of hyperreflexia after SCI. Specifically, astrocytic Rac1KO reduced SCI-related H-reflex hyperexcitability, decreased dendritic spine dysgenesis on α-motor neurons, and elevated the expression of the astrocytic glutamate transporter-1 (GLT-1). Overall, this study supports a distinct role for astrocytic Rac1 signaling within the spinal reflex circuit and the development of SCI-related spasticity.

The Small G-Protein Rac1 in the Dorsomedial Striatum Promotes Alcohol-Dependent Structural Plasticity and Goal-Directed Learning in Mice.

The small G-protein Ras-related C3 botulinum toxin substrate 1 (Rac1) promotes the formation of filamentous actin (F-actin). Actin is a major component of dendritic spines, and we previously found that alcohol alters actin composition and dendritic spine structure in the nucleus accumbens (NAc) and the dorsomedial striatum (DMS). To examine if Rac1 contributes to these alcohol-mediated adaptations, we measured the level of GTP-bound active Rac1 in the striatum of mice following 7 weeks of intermittent access to 20% alcohol. We found that chronic alcohol intake activates Rac1 in the DMS of male mice. In contrast, Rac1 is not activated by alcohol in the NAc and DLS of male mice or in the DMS of female mice. Similarly, closely related small G-proteins are not activated by alcohol in the DMS, and Rac1 activity is not increased in the DMS by moderate alcohol or natural reward. To determine the consequences of alcohol-dependent Rac1 activation in the DMS of male mice, we inhibited endogenous Rac1 by infecting the DMS of mice with an adeno-associated virus (AAV) expressing a dominant negative form of the small G-protein (Rac1-DN). We found that overexpression of AAV-Rac1-DN in the DMS inhibits alcohol-mediated Rac1 signaling and attenuates alcohol-mediated F-actin polymerization, which corresponded with a decrease in dendritic arborization and spine maturation. Finally, we provide evidence to suggest that Rac1 in the DMS plays a role in alcohol-associated goal-directed learning. Together, our data suggest that Rac1 in the DMS plays an important role in alcohol-dependent structural plasticity and aberrant learning.

RhoG facilitates a conformational transition in the guanine nucleotide exchange factor complex DOCK5/ELMO1 to an open state.

The dedicator of cytokinesis (DOCK)/engulfment and cell motility (ELMO) complex serves as a guanine nucleotide exchange factor (GEF) for the GTPase Rac. RhoG, another GTPase, activates the ELMO-DOCK-Rac pathway during engulfment and migration. Recent cryo-EM structures of the DOCK2/ELMO1 and DOCK2/ELMO1/Rac1 complexes have identified closed and open conformations that are key to understanding the autoinhibition mechanism. Nevertheless, the structural details of RhoG-mediated activation of the DOCK/ELMO complex remain elusive. Herein, we present cryo-EM structures of DOCK5/ELMO1 alone and in complex with RhoG and Rac1. The DOCK5/ELMO1 structure exhibits a closed conformation similar to that of DOCK2/ELMO1, suggesting a shared regulatory mechanism of the autoinhibitory state across DOCK-A/B subfamilies (DOCK1-5). Conversely, the RhoG/DOCK5/ELMO1/Rac1 complex adopts an open conformation that differs from that of the DOCK2/ELMO1/Rac1 complex, with RhoG binding to both ELMO1 and DOCK5. The alignment of the DOCK5 phosphatidylinositol (3,4,5)-trisphosphate binding site with the RhoG C-terminal lipidation site suggests simultaneous binding of RhoG and DOCK5/ELMO1 to the plasma membrane. Structural comparison of the apo and RhoG-bound states revealed that RhoG facilitates a closed-to-open state conformational change of DOCK5/ELMO1. Biochemical and surface plasmon resonance (SPR) assays confirm that RhoG enhances the Rac GEF activity of DOCK5/ELMO1 and increases its binding affinity for Rac1. Further analysis of structural variability underscored the conformational flexibility of the DOCK5/ELMO1/Rac1 complex core, potentially facilitating the proximity of the DOCK5 GEF domain to the plasma membrane. These findings elucidate the structural mechanism underlying the RhoG-induced allosteric activation and membrane binding of the DOCK/ELMO complex.

A genome-wide genetic screen identifies CYRI-B as a negative regulator of CEACAM3-mediated phagocytosis.

Opsonin-independent phagocytosis mediated by human carcinoembryonic antigen-related cell adhesion molecule 3 (CEACAM3) has evolved to control a subset of human-restricted bacterial pathogens. CEACAM3 engagement triggers rapid GTP-loading of the small GTPase Rac as a master regulator of cytoskeletal rearrangements and lamellipodia-driven internalization. To identify components of the CEACAM3-initiated signaling cascade, we performed a genome-wide CRISPR/Cas9-based screen in human myeloid cells. Following infection with fluorescently labeled bacteria, cells exhibiting elevated phagocytosis (gain-of-function) as well as cells showing reduced phagocytosis (loss-of-function) were sorted and enrichment of individual single-guide RNAs (sgRNAs) was determined by next generation sequencing. Concentrating on genes whose targeting by three distinct sgRNAs consistently resulted in a gain-of-function phenotype, we identified the Rac-GTP-sequestering protein CYRI-B as a negative regulator of CEACAM3-mediated phagocytosis. Clonal HL-60 cell lines with CYRI-B knockout showed enhanced CEACAM3-downstream signaling, such as Rac GTP loading and phosphorylation of PAK kinases, leading to increased phagocytosis of bacteria. Complementation of the CYRI-B knockout cells reverted the knockout phenotype. Our results unravel components of CEACAM3-initiated opsonin-independent phagocytosis on a genome-wide level and highlight CYRI-B as a negative regulator of CEACAM3-initiated signaling in myeloid cells.

Rac1 and Rac3 GTPases and TPC2 are required for axonal outgrowth and migration of cortical interneurons.

Rho GTPases, among them Rac1 and Rac3, are major transducers of extracellular signals and are involved in multiple cellular processes. In cortical interneurons, the neurons that control the balance between excitation and inhibition of cortical circuits, Rac1 and Rac3 are essential for their development. Ablation of both leads to a severe reduction in the numbers of mature interneurons found in the murine cortex, which is partially due to abnormal cell cycle progression of interneuron precursors and defective formation of growth cones in young neurons. Here, we present new evidence that upon Rac1 and Rac3 ablation, centrosome, Golgi complex and lysosome positioning is significantly perturbed, thus affecting both interneuron migration and axon growth. Moreover, for the first time, we provide evidence of altered expression and localization of the two-pore channel 2 (TPC2) voltage-gated ion channel that mediates Ca2+ release. Pharmacological inhibition of TPC2 negatively affected axonal growth and migration of interneurons. Our data, taken together, suggest that TPC2 contributes to the severe phenotype in axon growth initiation, extension and interneuron migration in the absence of Rac1 and Rac3.

A metazoan-specific C-terminal motif in EXC-4 and Gα-Rho/Rac signaling regulate cell outgrowth during tubulogenesis in C. elegans.

Chloride intracellular channels (CLICs) are conserved proteins for which the cellular and molecular functions remain mysterious. An important insight into CLIC function came from the discovery that Caenorhabditis elegans EXC-4/CLIC regulates morphogenesis of the excretory canal (ExCa) cell, a single-cell tube. Subsequent work showed that mammalian CLICs regulate vascular development and angiogenesis, and human CLIC1 can rescue exc-4 mutants, suggesting conserved function in biological tube formation (tubulogenesis) and maintenance. However, the cell behaviors and signaling pathways regulated by EXC-4/CLICs during tubulogenesis in vivo remain largely unknown. We report a new exc-4 mutation, affecting a C-terminal residue conserved in virtually all metazoan CLICs, that reveals a specific role for EXC-4 in ExCa outgrowth. Cell culture studies suggest a function for CLICs in heterotrimeric G protein (Gα/ß/γ)-Rho/Rac signaling, and Rho-family GTPases are common regulators of cell outgrowth. Using our new exc-4 mutant, we describe a previously unknown function for Gα-encoding genes (gpa-12/Gα12/13, gpa-7/Gαi, egl-30/Gαq and gsa-1/Gαs), ced-10/Rac and mig-2/RhoG in EXC-4-mediated ExCa outgrowth. Our results demonstrate that EXC-4/CLICs are primordial players in Gα-Rho/Rac-signaling, a pathway that is crucial for tubulogenesis in C. elegans and in vascular development.

RABV induces biphasic actin cytoskeletal rearrangement through Rac1 activity modulation.

Rabies virus (RABV) is highly lethal and triggers severe neurological symptoms. The neuropathogenic mechanism remains poorly understood. Ras-related C3 botulinum toxin substrate 1 (Rac1) is a Rho-GTPase that is involved in actin remodeling and has been reported to be closely associated with neuronal dysfunction. In this study, by means of a combination of pharmacological inhibitors, small interfering RNA, and specific dominant-negatives, we characterize the crucial roles of dynamic actin and the regulatory function of Rac1 in RABV infection, dominantly in the viral entry phase. The data show that the RABV phosphoprotein interacts with Rac1. RABV phosphoprotein suppress Rac1 activity and impedes downstream Pak1-Limk1-Cofilin1 signaling, leading to the disruption of F-actin-based structure formation. In early viral infection, the EGFR-Rac1-signaling pathway undergoes a biphasic change, which is first upregulated and subsequently downregulated, corresponding to the RABV entry-induced remodeling pattern of F-actin. Taken together, our findings demonstrate for the first time the role played by the Rac1 signaling pathway in RABV infection and may provide a clue for an explanation for the etiology of rabies neurological pathogenesis.IMPORTANCEThough neuronal dysfunction is predominant in fatal rabies, the detailed mechanism by which rabies virus (RABV) infection causes neurological symptoms remains in question. The actin cytoskeleton is involved in numerous viruses infection and plays a crucial role in maintaining neurological function. The cytoskeletal disruption is closely associated with abnormal nervous symptoms and induces neurogenic diseases. In this study, we show that RABV infection led to the rearrangement of the cytoskeleton as well as the biphasic kinetics of the Rac1 signal transduction. These results help elucidate the mechanism that causes the aberrant neuronal processes by RABV infection and may shed light on therapeutic development aimed at ameliorating neurological disorders.

FilGAP controls cell-extracellular matrix adhesion and process formation of kidney podocytes.

The function of kidney podocytes is closely associated with actin cytoskeleton regulated by Rho small GTPases. Loss of actin-driven cell adhesions and processes is connected to podocyte dysfunction, proteinuria, and kidney diseases. FilGAP, a GTPase-activating protein for Rho small GTPase Rac1, is abundantly expressed in kidney podocytes, and its gene is linked to diseases in a family with focal segmental glomerulosclerosis. In this study, we have studied the role of FilGAP in podocytes in vitro. Depletion of FilGAP in cultured podocytes induced loss of actin stress fibers and increased Rac1 activity. Conversely, forced expression of FilGAP increased stress fiber formation whereas Rac1 activation significantly reduced its formation. FilGAP localizes at the focal adhesion (FA), an integrin-based protein complex closely associated with stress fibers, that mediates cell-extracellular matrix (ECM) adhesion, and FilGAP depletion decreased FA formation and impaired attachment to the ECM. Moreover, in unique podocyte cell cultures capable of inducing the formation of highly organized processes including major processes and foot process-like projections, FilGAP depletion or Rac1 activation decreased the formation of these processes. The reduction of FAs and process formations in FilGAP-depleted podocyte cells was rescued by inhibition of Rac1 or P21-activated kinase 1 (PAK1), a downstream effector of Rac1, and PAK1 activation inhibited their formations. Thus, FilGAP contributes to both cell-ECM adhesion and process formation of podocytes by suppressing Rac1/PAK1 signaling.

The HACE1 E3 ligase mediates RAC1-dependent control of mTOR signaling complexes.

HACE1 is a HECT family E3 ubiquitin-protein ligase with broad but incompletely understood tumor suppressor activity. Here, we report a previously unrecognized link between HACE1 and signaling complexes containing mammalian target of rapamycin (mTOR). HACE1 blocks mTORC1 and mTORC2 activities by reducing mTOR stability in an E3 ligase-dependent manner. Mechanistically, HACE1 binds to and ubiquitylates Ras-related C3 botulinum toxin substrate 1 (RAC1) when RAC1 is associated with mTOR complexes, including at focal adhesions, leading to proteasomal degradation of RAC1. This in turn decreases the stability of mTOR to reduce mTORC1 and mTORC2 activity. HACE1 deficient cells show enhanced mTORC1/2 activity, which is reversed by chemical or genetic RAC1 inactivation but not in cells expressing the HACE1-insensitive mutant, RAC1K147R . In vivo, Rac1 deletion reverses enhanced mTOR expression in KRasG12D -driven lung tumors of Hace1-/- mice. HACE1 co-localizes with mTOR and RAC1, resulting in RAC1-dependent loss of mTOR protein stability. Together, our data demonstrate that HACE1 destabilizes mTOR by targeting RAC1 within mTOR-associated complexes, revealing a unique ubiquitin-dependent process to control the activity of mTOR signaling complexes.

Muscle stem cells get a new look: Dynamic cellular projections as sensors of the stem cell niche.

Cellular mechanisms whereby quiescent stem cells sense tissue injury and transition to an activated state are largely unknown. Quiescent skeletal muscle stem cells (MuSCs, also called satellite cells) have elaborate, heterogeneous projections that rapidly retract in response to muscle injury. They may therefore act as direct sensors of their niche environment. Retraction is driven by a Rac-to-Rho GTPase activity switch that promotes downstream MuSC activation events. These and other observations lead to several hypotheses: (1) projections are morphologically dynamic at quiescence, providing a surveillance function for muscle damage; (2) quiescent projection dynamics are regulated by the relative balance of Rac and Rho activities promoted by niche-derived cues; (3) projections, particularly their associated filopodia, sense tissue damage via changes to the biomechanical properties of the niche and/or detection of signaling cues released by damaged myofibers; and (4) the dynamic nature of projections results in a population of MuSCs with heterogeneous functional properties. These concepts may extend to other types of quiescent stem cells, as well as prove useful in translational research settings.

Social experiences switch states of memory engrams through regulating hippocampal Rac1 activity.

In pathological or artificial conditions, memory can be formed as silenced engrams that are unavailable for retrieval by presenting conditioned stimuli but can be artificially switched into the latent state so that natural recall is allowed. However, it remains unclear whether such different states of engrams bear any physiological significance and can be switched through physiological mechanisms. Here, we show that an acute social reward experience switches the silent memory engram into the latent state. Conversely, an acute social stress causes transient forgetting via turning a latent memory engram into a silent state. Such emotion-driven bidirectional switching between latent and silent states of engrams is mediated through regulation of Rac1 activity­dependent reversible forgetting in the hippocampus, as stress-activated Rac1 suppresses retrieval, while reward recovers silenced memory under amnesia by inhibiting Rac1. Thus, data presented reveal hippocampal Rac1 activity as the basis for emotion-mediated switching between latent and silent engrams to achieve emotion-driven behavioral flexibility.

Gain-of-function mutant p53 activates small GTPase Rac1 through SUMOylation to promote tumor progression.

Tumor suppressor p53 is frequently mutated in human cancer. Mutant p53 often promotes tumor progression through gain-of-function (GOF) mechanisms. However, the mechanisms underlying mutant p53 GOF are not well understood. In this study, we found that mutant p53 activates small GTPase Rac1 as a critical mechanism for mutant p53 GOF to promote tumor progression. Mechanistically, mutant p53 interacts with Rac1 and inhibits its interaction with SUMO-specific protease 1 (SENP1), which in turn inhibits SENP1-mediated de-SUMOylation of Rac1 to activate Rac1. Targeting Rac1 signaling by RNAi, expression of the dominant-negative Rac1 (Rac1 DN), or the specific Rac1 inhibitor NSC23766 greatly inhibits mutant p53 GOF in promoting tumor growth and metastasis. Furthermore, mutant p53 expression is associated with enhanced Rac1 activity in clinical tumor samples. These results uncover a new mechanism for Rac1 activation in tumors and, most importantly, reveal that activation of Rac1 is an unidentified and critical mechanism for mutant p53 GOF in tumorigenesis, which could be targeted for therapy in tumors containing mutant p53.

Unveiling the role of RAC3 in the growth and invasion of cisplatin-resistant bladder cancer cells.

Bladder cancer is one of the most prevalent cancers worldwide, and its morbidity and mortality rates have been increasing over the years. However, how RAC family small GTPase 3 (RAC3) affects the proliferation, migration and invasion of cisplatin-resistant bladder cancer cells remains unclear. Bioinformatics techniques were used to investigate the expression of RAC3 in bladder cancer tissues. Influences of RAC3 in the grade, stage, distant metastasis, and survival rate of bladder cancer were also examined. Analysis of the relationship between RAC3 expression and the immune microenvironment (TIME), genomic mutations, and stemness index. In normal bladder cancer cells (T24, 5637, and BIU-87) and cisplatin-resistant bladder cancer cells (BIU-87-DDP), the expression of RAC3 was detected separately with Western blotting. Plasmid transfection was used to overexpress or silence the expression of RAC3 in bladder cancer cells resistant to cisplatin (BIU-87-DDP). By adding activators and inhibitors, the activities of the JNK/MAPK signalling pathway were altered. Cell viability, invasion, and its level of apoptosis were measured in vitro using CCK-8, transwell, and flow cytometry. The bioinformatics analyses found RAC3 levels were elevated in bladder cancer tissues and were associated with a poor prognosis in bladder cancer. RAC3 in BIU-87-DDP cells expressed a higher level than normal bladder cancer cells. RAC3 overexpression promoted BIU-87-DDP proliferation. The growth of BIU-87-DDP cells slowed after the knockdown of RAC3, and RAC3 may have had an impact on the activation of the JNK/MAPK pathway.