Excerpt from HAMMERED

pages 78-9

Preschool music in the church school

https://place.asburyseminary.edu/ecommonsatsdissertations/2204/thumbnail.jp

Tropomyosin Promotes Lamellipodial Persistence by Collaborating with Arp2/3 at the Leading Edge

At the leading edge of migrating cells, protrusion of the lamellipodium is driven by Arp2/3-mediated polymerization of actin filaments [1]. This dense, branched actin network is promoted and stabilized by cortactin [2, 3]. In order to drive filament turnover, Arp2/3 networks are remodeled by proteins such as GMF, which blocks the actin-Arp2/3 interaction [4, 5], and coronin 1B, which acts by directing SSH1L to the lamellipodium where it activates the actin-severing protein cofilin [6, 7]. It has been shown in vitro that cofilin-mediated severing of Arp2/3 actin networks results in the generation of new pointed ends to which the actin-stabilizing protein tropomyosin (Tpm) can bind [8]. The presence of Tpm in lamellipodia, however, is disputed in the literature [9-19]. Here, we report that the Tpm isoforms 1.8/9 are enriched in the lamellipodium of fibroblasts as detected with a novel isoform-specific monoclonal antibody. RNAi-mediated silencing of Tpm1.8/9 led to an increase of Arp2/3 accumulation at the cell periphery and a decrease in the persistence of lamellipodia and cell motility, a phenotype consistent with cortactin- and coronin 1B-deficient cells [2, 7]. In the absence of coronin 1B or cofilin, Tpm1.8/9 protein levels are reduced while, conversely, inhibition of Arp2/3 with CK666 leads to an increase in Tpm1.8/9 protein. These findings establish a novel regulatory mechanism within the lamellipodium whereby Tpm collaborates with Arp2/3 to promote lamellipodial-based cell migration

All My Relations : Elders’ Teachings Grounding a Decolonial Bachelor of Nursing Program Philosophy

The aim of this paper is to share the process of developing a decolonial nursing program philosophical framework. Following cultural protocol, we asked two highly respected local Indigenous Knowledge holders and retired nurses to serve as Presiding Elders for the Bachelor of Nursing (BN) Curriculum Redesign Committee. Select committee members engaged with the Elders in a series of gatherings. Elders gifted ‘All My Relations’ as the unifying basis for their teachings of ‘The Four Realms’ and ‘Ani to Pisi -Spiderweb’ as the new BN program philosophical framework: local Indigenous teachings provide the standpoint for respectfully engaging with interconnected multiple worldviews to advance equity and restore wellness in nursing education. Collaboration with Elders provides an invaluable opportunity for decolonization in nursing workplaces and learning spaces. Résumé Le but de cet article est de partager le processus d’élaboration d’un cadre philosophique décolonial pour un programme de sciences infirmières. Conformément au protocole culturel, nous avons demandé à deux gardiens de connaissances autochtones locaux très respectés et à des infirmières à la retraite d’agir à titre d’Aînés Présidents du comité de refonte du curriculum du baccalauréat en sciences infirmières (BSC). Quelques membres du comité ont participé avec les aînés à une série de rencontres. Les aînés ont offert « All My Relations » comme base unificatrice de leurs enseignements sur « The Four Realms » (es quatre sphères) et « Ani to Pisi – Spiderweb » comme nouveau cadre philosophique du programme de baccalauréat en sciences infirmières : les enseignements autochtones locaux fournissent la perspective à partir de laquelle il est possible de s’engager respectueusement avec de multiples visions du monde interconnectées pour faire progresser l’équité et rétablir le bien-être dans la formation en sciences infirmières. La collaboration avec les aînés offre une occasion inestimable de décolonisation des lieux de travail et des espaces d’apprentissage en sciences infirmières

Genome-Wide Analysis of MEF2 Transcriptional Program Reveals Synaptic Target Genes and Neuronal Activity-Dependent Polyadenylation Site Selection

Although many transcription factors are known to control important aspects of neural development, the genome-wide programs that are directly regulated by these factors are not known. We have characterized the genetic program that is activated by MEF2, a key regulator of activity-dependent synapse development. These MEF2 target genes have diverse functions at synapses, revealing a broad role for MEF2 in synapse development. Several of the MEF2 targets are mutated in human neurological disorders including epilepsy and autism spectrum disorders, suggesting that these disorders may be caused by disruption of an activity-dependent gene program that controls synapse development. Our analyses also reveal that neuronal activity promotes alternative polyadenylation site usage at many of the MEF2 target genes, leading to the production of truncated mRNAs that may have different functions than their full-length counterparts. Taken together, these analyses suggest that the ubiquitously expressed transcription factor MEF2 regulates an intricate transcriptional program in neurons that controls synapse development

Arbaclofen in fragile X syndrome: results of phase 3 trials

Background: Arbaclofen improved multiple abnormal phenotypes in animal models of fragile X syndrome (FXS) and showed promising results in a phase 2 clinical study. The objective of the study is to determine safety and efficacy of arbaclofen for social avoidance in FXS. Methods: Two phase 3 placebo-controlled trials were conducted, a flexible dose trial in subjects age 12-50 (209FX301, adolescent/adult study) and a fixed dose trial in subjects age 5-11 (209FX302, child study). The primary endpoint for both trials was the Social Avoidance subscale of the Aberrant Behavior Checklist-Community Edition, FXS-specific (ABC-C FX ). Secondary outcomes included other ABC-C FX subscale scores, Clinical Global Impression-Improvement (CGI-I), Clinical Global Impression-Severity (CGI-S), and Vineland Adaptive Behavior Scales, Second Edition (Vineland-II) Socialization domain score. Results: A total 119 of 125 randomized subjects completed the adolescent/adult study (n = 57 arbaclofen, 62 placebo) and 159/172 completed the child study (arbaclofen 5 BID n = 38; 10 BID n = 39; 10 TID n = 38; placebo n = 44). There were no serious adverse events (AEs); the most common AEs included somatic (headache, vomiting, nausea), neurobehavioral (irritability/agitation, anxiety, hyperactivity), decreased appetite, and infectious conditions, many of which were also common on placebo. In the combined studies, there were 13 discontinuations (n = 12 arbaclofen, 1 placebo) due to AEs (all neurobehavioral). The adolescent/adult study did not show benefit for arbaclofen over placebo for any measure. In the child study, the highest dose group showed benefit over placebo on the ABC-C FX Irritability subscale (p = 0.03) and Parenting Stress Index (PSI, p = 0.03) and trends toward benefit on the ABC-C FX Social Avoidance and Hyperactivity subscales (both p < 0.1) and CGI-I (p = 0.119). Effect size in the highest dose group was similar to effect sizes for FDA-approved serotonin reuptake inhibitors (SSRIs). Conclusions: Arbaclofen did not meet the primary outcome of improved social avoidance in FXS in either study. Data from secondary measures in the child study suggests younger patients may derive benefit, but additional studies with a larger cohort on higher doses would be required to confirm this finding. The reported studies illustrate the challenges but represent a significant step forward in translating targeted treatments from preclinical models to clinical trials in humans with FXS

GMF controls branched actin content and lamellipodial retraction in fibroblasts

The lamellipodium is an important structure for cell migration containing branched actin nucleated via the Arp2/3 complex. The formation of branched actin is relatively well studied, but less is known about its disassembly and how this influences migration. GMF is implicated in both Arp2/3 debranching and inhibition of Arp2/3 activation. Modulation of GMFβ, a ubiquitous GMF isoform, by depletion or overexpression resulted in changes in lamellipodial dynamics, branched actin content, and migration. Acute pharmacological inhibition of Arp2/3 by CK-666, coupled to quantitative live-cell imaging of the complex, showed that depletion of GMFβ decreased the rate of branched actin disassembly. These data, along with mutagenesis studies, suggest that debranching (not inhibition of Arp2/3 activation) is a primary activity of GMFβ in vivo. Furthermore, depletion or overexpression of GMFβ disrupted the ability of cells to directionally migrate to a gradient of fibronectin (haptotaxis). These data suggest that debranching by GMFβ plays an important role in branched actin regulation, lamellipodial dynamics, and directional migration

Negative Allosteric Modulation of mGluR5 Partially Corrects Pathophysiology in a Mouse Model of Rett Syndrome

Rett syndrome (RTT) is caused by mutations in the gene encoding methyl-CpG binding protein 2 (MECP2), an epigenetic regulator of mRNA transcription. Here, we report a test of the hypothesis of shared pathophysiology of RTT and fragile X, another monogenic cause of autism and intellectual disability. In fragile X, the loss of the mRNA translational repressor FMRP leads to exaggerated protein synthesis downstream of metabotropic glutamate receptor 5 (mGluR5). We found that mGluR5- and protein-synthesis-dependent synaptic plasticity were similarly altered in area CA1 of Mecp2 KO mice. CA1 pyramidal cell-type-specific, genome-wide profiling of ribosome-bound mRNAs was performed in wild-type and Mecp2 KO hippocampal CA1 neurons to reveal the MeCP2-regulated “translatome.” We found significant overlap between ribosome-bound transcripts overexpressed in the Mecp2 KO and FMRP mRNA targets. These tended to encode long genes that were functionally related to either cytoskeleton organization or the development of neuronal connectivity. In the Fmr1 KO mouse, chronic treatment with mGluR5-negative allosteric modulators (NAMs) has been shown to ameliorate many mutant phenotypes by correcting excessive protein synthesis. In Mecp2 KO mice, we found that mGluR5 NAM treatment significantly reduced the level of overexpressed ribosome-associated transcripts, particularly those that were also FMRP targets. Some Rett phenotypes were also ameliorated by treatment, most notably hippocampal cell size and lifespan. Together, these results suggest a potential mechanistic link between MeCP2-mediated transcription regulation and mGluR5/FMRP-mediated protein translation regulation through coregulation of a subset of genes relevant to synaptic functions. SIGNIFICANCE STATEMENT Altered regulation of synaptic protein synthesis has been hypothesized to contribute to the pathophysiology that underlies multiple forms of intellectual disability and autism spectrum disorder. Here, we show in a mouse model of Rett syndrome (Mecp2 KO) that metabotropic glutamate receptor 5 (mGluR5)- and protein-synthesis-dependent synaptic plasticity are abnormal in the hippocampus. We found that a subset of ribosome-bound mRNAs was aberrantly upregulated in hippocampal CA1 neurons of Mecp2 KO mice, that these significantly overlapped with FMRP direct targets and/or SFARI human autism genes, and that chronic treatment of Mecp2 KO mice with an mGluR5-negative allosteric modulator tunes down upregulated ribosome-bound mRNAs and partially improves mutant mice phenotypes.Howard Hughes Medical InstituteSimons Foundation (Grant SFARI 275050)Rett Syndrome Research Trus

If we build it they will come: targeting the immune response to breast cancer.

Historically, breast cancer tumors have been considered immunologically quiescent, with the majority of tumors demonstrating low lymphocyte infiltration, low mutational burden, and modest objective response rates to anti-PD-1/PD-L1 monotherapy. Tumor and immunologic profiling has shed light on potential mechanisms of immune evasion in breast cancer, as well as unique aspects of the tumor microenvironment (TME). These include elements associated with antigen processing and presentation as well as immunosuppressive elements, which may be targeted therapeutically. Examples of such therapeutic strategies include efforts to (1) expand effector T-cells, natural killer (NK) cells and immunostimulatory dendritic cells (DCs), (2) improve antigen presentation, and (3) decrease inhibitory cytokines, tumor-associated M2 macrophages, regulatory T- and B-cells and myeloid derived suppressor cells (MDSCs). The goal of these approaches is to alter the TME, thereby making breast tumors more responsive to immunotherapy. In this review, we summarize key developments in our understanding of antitumor immunity in breast cancer, as well as emerging therapeutic modalities that may leverage that understanding to overcome immunologic resistance

Lamellipodia are crucial for haptotactic sensing and response

Haptotaxis is the process by which cells respond to gradients of substrate-bound cues, such as extracellular matrix proteins (ECM); however, the cellular mechanism of this response remains poorly understood and has mainly been studied by comparing cell behavior on uniform ECMs with different concentrations of components. To study haptotaxis in response to gradients, we utilized microfluidic chambers to generate gradients of the ECM protein fibronectin, and imaged the cell migration response. Lamellipodia are fan-shaped protrusions that are common in migrating cells. Here, we define a new function for lamellipodia and the cellular mechanism required for haptotaxis – differential actin and lamellipodial protrusion dynamics lead to biased cell migration. Modest differences in lamellipodial dynamics occurring over time periods of seconds to minutes are summed over hours to produce differential whole cell movement towards higher concentrations of fibronectin. We identify a specific subset of lamellipodia regulators as being crucial for haptotaxis. Numerous studies have linked components of this pathway to cancer metastasis and, consistent with this, we find that expression of the oncogenic Rac1 P29S mutation abrogates haptotaxis. Finally, we show that haptotaxis also operates through this pathway in 3D environments