ResearchPad - 29 https://www.researchpad.co Default RSS Feed en-us © 2020 Newgen KnowledgeWorks <![CDATA[TFEB controls retromer expression in response to nutrient availability]]> https://www.researchpad.co/article/elastic_article_15309 Endosomal recycling maintains the cell surface abundance of nutrient transporters for nutrient uptake, but how the cell integrates nutrient availability with recycling is less well understood. Here, in studying the recycling of human glutamine transporters ASCT2 (SLC1A5), LAT1 (SLC7A5), SNAT1 (SLC38A1), and SNAT2 (SLC38A2), we establish that following amino acid restriction, the adaptive delivery of SNAT2 to the cell surface relies on retromer, a master conductor of endosomal recycling. Upon complete amino acid starvation or selective glutamine depletion, we establish that retromer expression is upregulated by transcription factor EB (TFEB) and other members of the MiTF/TFE family of transcription factors through association with CLEAR elements in the promoters of the retromer genes VPS35 and VPS26A. TFEB regulation of retromer expression therefore supports adaptive nutrient acquisition through endosomal recycling.

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<![CDATA[CAMIO: a transgenic CRISPR pipeline to create diverse targeted genome deletions in <i>Drosophila</i>]]> https://www.researchpad.co/article/Nba0ad44c-f37d-4c29-ab52-bec343ddd845 The genome is the blueprint for an organism. Interrogating the genome, especially locating critical cis-regulatory elements, requires deletion analysis. This is conventionally performed using synthetic constructs, making it cumbersome and non-physiological. Thus, we created Cas9-mediated Arrayed Mutagenesis of Individual Offspring (CAMIO) to achieve comprehensive analysis of a targeted region of native DNA. CAMIO utilizes CRISPR that is spatially restricted to generate independent deletions in the intact Drosophila genome. Controlled by recombination, a single guide RNA is stochastically chosen from a set targeting a specific DNA region. Combining two sets increases variability, leading to either indels at 1–2 target sites or inter-target deletions. Cas9 restriction to male germ cells elicits autonomous double-strand-break repair, consequently creating offspring with diverse mutations. Thus, from a single population cross, we can obtain a deletion matrix covering a large expanse of DNA at both coarse and fine resolution. We demonstrate the ease and power of CAMIO by mapping 5′UTR sequences crucial for chinmo's post-transcriptional regulation.

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<![CDATA[Rapid and assured genetic engineering methods applied to <i>Acinetobacter baylyi</i> ADP1 genome streamlining]]> https://www.researchpad.co/article/N8ac39c8d-ee13-4bb0-b3b9-5a6810ec4315 One goal of synthetic biology is to improve the efficiency and predictability of living cells by removing extraneous genes from their genomes. We demonstrate improved methods for engineering the genome of the metabolically versatile and naturally transformable bacterium Acinetobacter baylyi ADP1 and apply them to a genome streamlining project. In Golden Transformation, linear DNA fragments constructed by Golden Gate Assembly are directly added to cells to create targeted deletions, edits, or additions to the chromosome. We tested the dispensability of 55 regions of the ADP1 chromosome using Golden Transformation. The 18 successful multiple-gene deletions ranged in size from 21 to 183 kb and collectively accounted for 23.4% of its genome. The success of each multiple-gene deletion attempt could only be partially predicted on the basis of an existing collection of viable ADP1 single-gene deletion strains and a new transposon insertion sequencing (Tn-Seq) dataset that we generated. We further show that ADP1’s native CRISPR/Cas locus is active and can be retargeted using Golden Transformation. We reprogrammed it to create a CRISPR-Lock, which validates that a gene has been successfully removed from the chromosome and prevents it from being reacquired. These methods can be used together to implement combinatorial routes to further genome streamlining and for more rapid and assured metabolic engineering of this versatile chassis organism.

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<![CDATA[Biarticular muscles in light of template models, experiments and robotics: a review]]> https://www.researchpad.co/article/Ned029f8a-6261-4e91-9952-9b542b3ef533

Leg morphology is an important outcome of evolution. A remarkable morphological leg feature is the existence of biarticular muscles that span adjacent joints. Diverse studies from different fields of research suggest a less coherent understanding of the muscles’ functionality in cyclic, sagittal plane locomotion. We structured this review of biarticular muscle function by reflecting biomechanical template models, human experiments and robotic system designs. Within these approaches, we surveyed the contribution of biarticular muscles to the locomotor subfunctions (stance, balance and swing). While mono- and biarticular muscles do not show physiological differences, the reviewed studies provide evidence for complementary and locomotor subfunction-specific contributions of mono- and biarticular muscles. In stance, biarticular muscles coordinate joint movements, improve economy (e.g. by transferring energy) and secure the zig-zag configuration of the leg against joint overextension. These commonly known functions are extended by an explicit role of biarticular muscles in controlling the angular momentum for balance and swing. Human-like leg arrangement and intrinsic (compliant) properties of biarticular structures improve the controllability and energy efficiency of legged robots and assistive devices. Future interdisciplinary research on biarticular muscles should address their role for sensing and control as well as non-cyclic and/or non-sagittal motions, and non-static moment arms.

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<![CDATA[PD‐L1 Expression on Circulating Tumor Cells May Be Predictive of Response to Pembrolizumab in Advanced Melanoma: Results from a Pilot Study]]> https://www.researchpad.co/article/Nbb29cb12-ecfd-47f6-b59e-266588402b7b

Abstract

Background

PD‐1 inhibitors are routinely used for the treatment of advanced melanoma. This study sought to determine whether PD‐L1 expression on circulating tumor cells (CTCs) can serve as a predictive biomarker of clinical benefit and response to treatment with the PD‐1 inhibitor pembrolizumab.

Methods

Blood samples were collected from patients with metastatic melanoma receiving pembrolizumab, prior to treatment and 6–12 weeks after initiation of therapy. Multiparametric flow cytometry was used to identify CTCs and evaluate the expression of PD‐L1.

Results

CTCs were detected in 25 of 40 patients (63%). Patients with detectable PD‐L1+ CTCs (14/25, 64%) had significantly longer progression‐free survival (PFS) compared with patients with PD‐L1 CTCs (26.6 months vs. 5.5 months; p = .018). The 12‐month PFS rates were 76% versus 22% in the PD‐L1+ versus PD‐L1 CTCs groups (p = .012), respectively. A multivariate linear regression analysis confirmed that PD‐L1+ CTC is an independent predictive biomarker of PFS (hazard ratio, 0.229; 95% confidence interval, 0.052–1.012; p = .026).

Conclusion

Our results reveal the potential of CTCs as a noninvasive real‐time biopsy to evaluate PD‐L1 expression in patients with melanoma. PD‐L1 expression on CTCs may be predictive of response to pembrolizumab and longer PFS.

Implications for Practice

The present data suggest that PD‐L1 expression on circulating tumor cells may predict response to pembrolizumab in advanced melanoma. This needs further validation in a larger trial and, if proven, might be a useful liquid biopsy tool that could be used to stratify patients into groups more likely to respond to immunotherapy, hence leading to health cost savings.

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<![CDATA[Immune‐Related Adverse Events in the Setting of PD‐1/L1 Inhibitor Combination Therapy]]> https://www.researchpad.co/article/Nfc2d3096-ba3e-4e2e-bf11-c0fc4457547e

In 2018, a multi‐disciplinary workshop was held at the Massachusetts General Hospital to discuss challenges in defining, diagnosing, and treating immune‐related adverse events (irAE), including those that occur in patients administered PD‐1/L1 inhibitor combination therapy. In this commentary, the workshop participants present a clinical case that illustrates the complexity of irAE diagnosis and management in a patient receiving PD‐1/L1 combination therapy, summarize the current state of PD‐1/L1 combination therapy, and discuss challenges and opportunities for the evaluation of irAEs as these combinations become more widely used to treat patients with cancer.

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<![CDATA[GO: a functional reporter system to identify and enrich base editing activity]]> https://www.researchpad.co/article/N8a9f1098-b74a-4fac-bc68-452350dae1bd

Abstract

Base editing (BE) is a powerful tool for engineering single nucleotide variants (SNVs) and has been used to create targeted mutations in cell lines, organoids and animal models. Recent development of new BE enzymes has provided an extensive toolkit for genome modification; however, identifying and isolating edited cells for analysis has proven challenging. Here we report a ‘Gene On’ (GO) reporter system that indicates precise cytosine or adenine base editing in situ with high sensitivity and specificity. We test GO using an activatable GFP and use it to measure the kinetics, efficiency and PAM specificity of a range of new BE variants. Further, GO is flexible and can be easily adapted to induce expression of numerous genetically encoded markers, antibiotic resistance genes or enzymes, such as Cre recombinase. With these tools, GO can be exploited to functionally link BE events at endogenous genomic loci to cellular enzymatic activities in human and mouse cell lines and organoids. Thus, GO provides a powerful approach to increase the practicality and feasibility of implementing CRISPR BE in biomedical research.

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<![CDATA[Robotic modelling of snake traversing large, smooth obstacles reveals stability benefits of body compliance]]> https://www.researchpad.co/article/N45b4bf44-7880-46fd-95ec-dcb811a85356

Snakes can move through almost any terrain. Although their locomotion on flat surfaces using planar gaits is inherently stable, when snakes deform their body out of plane to traverse complex terrain, maintaining stability becomes a challenge. On trees and desert dunes, snakes grip branches or brace against depressed sand for stability. However, how they stably surmount obstacles like boulders too large and smooth to gain such ‘anchor points’ is less understood. Similarly, snake robots are challenged to stably traverse large, smooth obstacles for search and rescue and building inspection. Our recent study discovered that snakes combine body lateral undulation and cantilevering to stably traverse large steps. Here, we developed a snake robot with this gait and snake-like anisotropic friction and used it as a physical model to understand stability principles. The robot traversed steps as high as a third of its body length rapidly and stably. However, on higher steps, it was more likely to fail due to more frequent rolling and flipping over, which was absent in the snake with a compliant body. Adding body compliance reduced the robot's roll instability by statistically improving surface contact, without reducing speed. Besides advancing understanding of snake locomotion, our robot achieved high traversal speed surpassing most previous snake robots and approaching snakes, while maintaining high traversal probability.

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<![CDATA[Abstracts]]> https://www.researchpad.co/article/N16e1628c-c3f5-44f1-8bc3-198ec647b8c3 ]]> <![CDATA[Ventricular volume expansion in presymptomatic genetic frontotemporal dementia]]> https://www.researchpad.co/article/N6fc41365-e031-43f7-88ba-f2f4d2833920

Objective

To characterize the time course of ventricular volume expansion in genetic frontotemporal dementia (FTD) and identify the onset time and rates of ventricular expansion in presymptomatic FTD mutation carriers.

Methods

Participants included patients with a mutation in MAPT, PGRN, or C9orf72, or first-degree relatives of mutation carriers from the GENFI study with MRI scans at study baseline and at 1 year follow-up. Ventricular volumes were obtained from MRI scans using FreeSurfer, with manual editing of segmentation and comparison to fully automated segmentation to establish reliability. Linear mixed models were used to identify differences in ventricular volume and in expansion rates as a function of time to expected disease onset between presymptomatic carriers and noncarriers.

Results

A total of 123 participants met the inclusion criteria and were included in the analysis (18 symptomatic carriers, 46 presymptomatic mutation carriers, and 56 noncarriers). Ventricular volume differences were observed 4 years prior to symptom disease onset for presymptomatic carriers compared to noncarriers. Annualized rates of ventricular volume expansion were greater in presymptomatic carriers relative to noncarriers. Importantly, time-intensive manually edited and fully automated ventricular volume resulted in similar findings.

Conclusions

Ventricular volume differences are detectable in presymptomatic genetic FTD. Concordance of results from time-intensive manual editing and fully automatic segmentation approaches support its value as a measure of disease onset and progression in future studies in both presymptomatic and symptomatic genetic FTD.

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<![CDATA[Dual Checkpoint Inhibition with Ipilimumab plus Nivolumab After Progression on Sequential PD‐1/PDL‐1 Inhibitors Pembrolizumab and Atezolizumab in a Patient with Lynch Syndrome, Metastatic Colon, and Localized Urothelial Cancer]]> https://www.researchpad.co/article/N1c8557d6-a1f0-49fd-b66f-d5ae6d45b1b9

The approval of PD‐1/PDL‐1 and CTLA‐4 specific therapies has revolutionized oncology; however, the sequencing of these drugs to treat MSI‐high metastatic colorectal cancer is still being determined. this article presents the case of a patient with Lynch syndrome who was treated with multiple sequential checkpoint inhibitors with prolonged disease control.

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<![CDATA[A Case of Nivolumab‐Induced Bullous Pemphigoid: Review of Dermatologic Toxicity Associated with Programmed Cell Death Protein‐1/Programmed Death Ligand‐1 Inhibitors and Recommendations for Diagnosis and Management]]> https://www.researchpad.co/article/5c16d221d5eed0c484539b8f

Bullous pemphigoid is an autoimmune subepidermal blistering disease characterized by the development of tense bullae and is most frequently seen in the elderly. PD‐1/PD‐L1‐induced bullous pemphigoid (BP) has emerged as a potentially serious dermatologic toxicity. This article reports a case of a 72‐year‐old woman who developed BP shortly after initiating treatment with the PD‐1 inhibitor nivolumab for metastatic non‐small cell lung cancer.

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<![CDATA[Ectopic expression of Pax4 in pancreatic δ cells results in β-like cell neogenesis]]> https://www.researchpad.co/article/5c01f912d5eed0c4842bfda1

Diabetes can result from the loss or dysfunction of insulin-producing δ cells. Druelle et al. find that ectopic expression of the pancreatic beta cell master gene Pax4 in somatostatin-expressing δ cells triggers an endocrine cell neogenesis process. Importantly, the beta-like cells thereby generated are functional and can partly reverse the consequences of chemically induced diabetes.

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<![CDATA[BAG3 directly stabilizes Hexokinase 2 mRNA and promotes aerobic glycolysis in pancreatic cancer cells]]> https://www.researchpad.co/article/5c01f8f7d5eed0c4842bf59e

Cancer cells frequently exhibit aerobic glycolysis, known as the Warburg effect. An et al. identify a novel RNA regulatory activity of BAG3, a protein implicated in various cancers. BAG3 enhances posttranscriptionally the expression of Hexokinase 2, the first enzyme involved in glycolysis, and it reprograms glucose metabolism by pancreatic cancer cells.

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<![CDATA[PtdIns3P controls mTORC1 signaling through lysosomal positioning]]> https://www.researchpad.co/article/5c01f8f0d5eed0c4842bf3e6

mTORC1 is activated by lysosome positioning and by amino acid–induced phosphatidylinositol 3-phosphate (PtdIns3P). Hong et al. show that amino acids stimulate recruitment of the PtdIns3P-binding protein FYCO1 to lysosomes and promote contacts between FYCO1 lysosomes and ER that contains the PtdIns3P effector Protrudin, mediating lysosome translocation and facilitating mTORC1 activation.

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<![CDATA[Reversible frontotemporal brain sagging syndrome]]> https://www.researchpad.co/article/5bc1e4a440307c75cdb9a1a7 ]]> <![CDATA[Pitch profile across the cuticle of the scarab beetle Cotinis mutabilis determined by analysis of Mueller matrix measurements]]> https://www.researchpad.co/article/5c4b8f58d5eed0c484870417

Helicoidal structures of lamellae of nanofibrils constitute the cuticle of some scarab beetles with iridescent metallic-like shine reflecting left-handed polarized light. The spectral and polarization properties of the reflected light depend on the pitch of the helicoidal structures, dispersion of effective refractive indices and thicknesses of layers in the cuticle. By modelling the outer exocuticle of the scarab beetle Cotinis mutabilis as a stack of continuously twisted biaxial slices of transparent materials, we extract optical and structural parameters by nonlinear regression analysis of variable-angle Mueller-matrix spectroscopic data. Inhomogeneities in the beetle cuticle produce depolarization with non-uniformity in cuticle thickness as the dominant effect. The pitch across the cuticle of C. mutabilis decreased with depth in a two-level profile from 380 to 335 nm and from 390 to 361 nm in greenish and reddish specimens, respectively, whereas in a yellowish specimen, the pitch decreased with depth in a three-level profile from 388 to 326 nm.

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<![CDATA[Loss of TBK1 is a frequent cause of frontotemporal dementia in a Belgian cohort]]> https://www.researchpad.co/article/5af6ff22463d7e6f5c49e28e

Objective:

To assess the genetic contribution of TBK1, a gene implicated in amyotrophic lateral sclerosis (ALS), frontotemporal dementia (FTD), and FTD-ALS, in Belgian FTD and ALS patient cohorts containing a significant part of genetically unresolved patients.

Methods:

We sequenced TBK1 in a hospital-based cohort of 482 unrelated patients with FTD and FTD-ALS and 147 patients with ALS and an extended Belgian FTD-ALS family DR158. We followed up mutation carriers by segregation studies, transcript and protein expression analysis, and immunohistochemistry.

Results:

We identified 11 patients carrying a loss-of-function (LOF) mutation resulting in an overall mutation frequency of 1.7% (11/629), 1.1% in patients with FTD (5/460), 3.4% in patients with ALS (5/147), and 4.5% in patients with FTD-ALS (1/22). We found 1 LOF mutation, p.Glu643del, in 6 unrelated patients segregating with disease in family DR158. Of 2 mutation carriers, brain and spinal cord was characterized by TDP-43-positive pathology. The LOF mutations including the p.Glu643del mutation led to loss of transcript or protein in blood and brain.

Conclusions:

TBK1 LOF mutations are the third most frequent cause of clinical FTD in the Belgian clinically based patient cohort, after C9orf72 and GRN, and the second most common cause of clinical ALS after C9orf72. These findings reinforce that FTD and ALS belong to the same disease continuum.

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<![CDATA[Multi-robot replication of ant collective towing behaviours]]> https://www.researchpad.co/article/5c1522abd5eed0c4840bd55a

In this work, teams of small mobile robots are used to test hypotheses about cooperative transport by ants. This study attempts to explain a decrease in steady-state transport speed with increasing team size that was previously observed in the ant Novomessor cockerelli. Two models of one-dimensional collective towing are compared: one in which transporters with different maximum speeds pull the payload with continuous, variable forces and another in which transporters with identical speeds pull with intermittent, unsynchronized forces. A statistical analysis of ant data supports the hypothesis that ants behave according to the first model, in which the steady-state transport speed is the maximum speed of the slowest teammate. By contrast, the ant data are not consistent with the second model, which predicts constant speed regardless of team size. To verify these predictions, the ant behaviours in each model are translated into decentralized controllers and implemented on teams of two to four robots. The controller for the first model incorporates a real-time reinforcement learning algorithm that successfully reproduces the observed relationship between ant team size and transport speed. The controller for the second model yields the predicted invariance of transport speed with team size. These results show the value of robotic swarms for testing mechanistic hypotheses about biological collectives.

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