Phagocytosis of apoptotic cells by both professional and semi‐professional phagocytes is required for resolution of organ damage and maintenance of immune tolerance. KIM‐1/TIM‐1 is a ...phosphatidylserine receptor that is expressed on epithelial cells and can transform the cells into phagocytes. Here, we demonstrate that KIM‐1 phosphorylation and association with p85 results in encapsulation of phagosomes by lipidated LC3 in multi‐membrane organelles. KIM‐1‐mediated phagocytosis is not associated with increased ROS production, and NOX inhibition does not block LC3 lipidation. Autophagy gene expression is required for efficient clearance of apoptotic cells and phagosome maturation. KIM‐1‐mediated phagocytosis leads to pro‐tolerogenic antigen presentation, which suppresses CD4 T‐cell proliferation and increases the percentage of regulatory T cells in an autophagy gene‐dependent manner. Taken together, these data reveal a novel mechanism of epithelial biology linking phagocytosis, autophagy and antigen presentation to regulation of the inflammatory response.
Synopsis
This study reveals how KIM‐1/TIM‐1‐mediated phagocytosis down‐modulates inflammation through ATG5 and ULK1‐dependent autophagic processing of phagocytosed material to MHC I and MHC II.
Phagocytosis of apoptotic cells by KIM‐1, a non‐myeloid phagocytosis receptor, induces LC3 lipidation of resulting phagosomes.
Autophagy protein expression, ATG5, ULK1 and Beclin1, are required for LC3 targeting to phagosomes and efficient phagosome acidification and maturation.
KIM‐1 signaling via the PI3 kinase subunit p85 promotes LC3 lipidation and accumulation on phagosomes.
Phagocytosed material processed through autophagy is presented to MHC I and MHC II.
Enhanced antigen presentation resulting from autophagic processing results in a pro‐tolerogenic milieu, reducing the percentage of CD4 effector T cells while increasing the percentage of regulatory T cells.
Phosphatidylserine receptor KIM‐1/TIM‐1‐mediated phagocytosis and autophagic processing of apoptotic cells promote antigen presentation in epithelial cells to modulate inflammation.
Serial circulating tumor DNA (ctDNA) monitoring is emerging as a non-invasive strategy to predict and monitor immune checkpoint blockade (ICB) therapeutic efficacy across cancer types. Yet, limited ...data exist to show the relationship between ctDNA dynamics and tumor genome and immune microenvironment in patients receiving ICB. Here, we present an in-depth analysis of clinical, whole-exome, transcriptome, and ctDNA profiles of 73 patients with advanced solid tumors, across 30 cancer types, from a phase II basket clinical trial of pembrolizumab (NCT02644369) and report changes in genomic and immune landscapes (primary outcomes). Patients stratified by ctDNA and tumor burden dynamics correspond with survival and clinical benefit. High mutation burden, high expression of immune signatures, and mutations in BRCA2 are associated with pembrolizumab molecular sensitivity, while abundant copy-number alterations and B2M loss-of-heterozygosity corresponded with resistance. Upon treatment, induction of genes expressed by T cell, B cell, and myeloid cell populations are consistent with sensitivity and resistance. We identified the upregulated expression of PLA2G2D, an immune-regulating phospholipase, as a potential biomarker of adaptive resistance to ICB. Together, these findings provide insights into the diversity of immunogenomic mechanisms that underpin pembrolizumab outcomes.
Scribble (SCRIB) is a tumor-suppressor protein, playing critical roles in establishing and maintaining epithelial cell polarity. SCRIB is frequently amplified in human cancers but does not localize ...properly to cell-cell junctions, suggesting that mislocalization of SCRIB disrupts its tumor-suppressive activities. Using chemical reporters, here we showed that SCRIB localization was regulated by S-palmitoylation at conserved cysteine residues. Palmitoylation-deficient mutants of SCRIB were mislocalized, leading to disruption of cell polarity and loss of their tumor-suppressive activities to oncogenic YAP, MAPK and PI3K/AKT pathways. We further found that ZDHHC7 was the major palmitoyl acyltransferase regulating SCRIB. Knockout of ZDHHC7 led to SCRIB mislocalization and YAP activation, and disruption of SCRIB's suppressive activities in HRas(V12)-induced cell invasion. In summary, we demonstrated that ZDHHC7-mediated SCRIB palmitoylation is critical for SCRIB membrane targeting, cell polarity and tumor suppression, providing new mechanistic insights of how dynamic protein palmitoylation regulates cell polarity and tumorigenesis.
We have identified longevity-associated genes in a long-lived Caenorhabditis elegans daf-2 (insulin/IGF receptor) mutant using serial analysis of gene expression (SAGE), a method that efficiently ...quantifies large numbers of mRNA transcripts by sequencing short tags. Reduction of daf-2 signaling in these mutant worms leads to a doubling in mean lifespan. We prepared C. elegans SAGE libraries from 1, 6, and 10-d-old adult daf-2 and from 1 and 6-d-old control adults. Differences in gene expression between daf-2 libraries representing different ages and between daf-2 versus control libraries identified not only single genes, but whole gene families that were differentially regulated. These gene families are part of major metabolic pathways including lipid, protein, and energy metabolism, stress response, and cell structure. Similar expression patterns of closely related family members emphasize the importance of these genes in aging-related processes. Global analysis of metabolism-associated genes showed hypometabolic features in mid-life daf-2 mutants that diminish with advanced age. Comparison of our results to recent microarray studies highlights sets of overlapping genes that are highly conserved throughout evolution and thus represent strong candidate genes that control aging and longevity.
Alternative splicing yields functionally distinctive gene products, and their balance plays critical roles in cell differentiation and development. We have previously shown that tumor-associated ...enhancer loss in coactivator gene CoAA leads to its altered alternative splicing. Here we identified two intergenic splicing variants, a zinc finger-containing coactivator CoAZ and a non-coding transcript ncCoAZ, between CoAA and its downstream corepressor gene RBM4. During stem/progenitor cell neural differentiation, we found that the switched alternative splicing and trans-splicing between CoAA and RBM4 transcripts result in lineage-specific expression of wild type CoAA, RBM4, and their variants. Stable expression of CoAA, RBM4, or their variants prevents the switch and disrupts the embryoid body formation. In addition, CoAA and RBM4 counter-regulate the target gene Tau at exon 10, and their splicing activities are subjected to the control by each splice variant. Further phylogenetic analysis showed that mammalian CoAA and RBM4 genes share common ancestry with the Drosophila melanogaster gene Lark, which is known to regulate early development and circadian rhythms. Thus, the trans-splicing between CoAA and RBM4 transcripts may represent a required regulation preserved during evolution. Our results demonstrate that a linked splicing control of transcriptional coactivator and corepressor is involved in stem/progenitor cell differentiation. The alternative splicing imbalance of CoAA and RBM4, because of loss of their common enhancer in cancer, may deregulate stem/progenitor cell differentiation.
Immune checkpoint inhibitors (ICIs) demonstrate unprecedented efficacy in multiple malignancies; however, the mechanisms of sensitivity and resistance are poorly understood and predictive biomarkers ...are scarce. INSPIRE is a phase 2 basket study to evaluate the genomic and immune landscapes of peripheral blood and tumors following pembrolizumab treatment.
Patients with incurable, locally advanced or metastatic solid tumors that have progressed on standard therapy, or for whom no standard therapy exists or standard therapy was not deemed appropriate, received 200 mg pembrolizumab intravenously every three weeks. Blood and tissue samples were collected at baseline, during treatment, and at progression. One core biopsy was used for immunohistochemistry and the remaining cores were pooled and divided for genomic and immune analyses. Univariable analysis of clinical, genomic, and immunophenotyping parameters was conducted to evaluate associations with treatment response in this exploratory analysis.
Eighty patients were enrolled from March 21, 2016 to June 1, 2017, and 129 tumor and 382 blood samples were collected. Immune biomarkers were significantly different between the blood and tissue. T cell PD-1 was blocked (≥98%) in the blood of all patients by the third week of treatment. In the tumor, 5/11 (45%) and 11/14 (79%) patients had T cell surface PD-1 occupance at weeks six and nine, respectively. The proportion of genome copy number alterations and abundance of intratumoral 4-1BB+ PD-1+ CD8 T cells at baseline (P < 0.05), and fold-expansion of intratumoral CD8 T cells from baseline to cycle 2-3 (P < 0.05) were associated with treatment response.
This study provides technical feasibility data for correlative studies. Tissue biopsies provide distinct data from the blood and may predict response to pembrolizumab.
The single crystal X-ray structure of the all-axial conformer of the (R,R,R,R ) enantiomer of the chiral donor tetramethyl-BEDT-TTF (TM-BEDT-TTF) was described and compared to the all-equatorial ...conformer. (S,S,S,S )-Tetramethyl-BEDT-TTF formed crystalline 1 : 1 complexes with TCNQ and TCNQ-F sub(4), as well as a THF solvate of the TCNQ complex. Donors bis((2S,4S)-pentane-2,4-dithio)te trathiafulvalene and (ethylenedithio)((2S,4S)-pentane- 2,4-dithio)tetrathiafulvalene, which contain seven-membered rings bearing chirally oriented methyl groups, only formed complexes with TCNQ-F sub(4). The TCNQ-F sub(4) complexes contain planar organosulfur systems, in contrast to the TCNQ complexes in which there is minimal charge transfer. A variety of crystal packing modes were observed. Electrocrystallization experiments with both enantiomers and the racemic form of tetramethyl-BEDT-TTF afforded mixed valence radical cation salts with the AsF sub(6) and SbF sub(6) anions formulated as (TM-BEDT-TTF) sub(2)XF sub(6) (X = As, Sb). Electrical conductivity was only found in one charge transfer complex, while the radical cation salts are all semiconducting.
Abstract
Phagocytosis of apoptotic cells by both professional and semi‐professional phagocytes is required for resolution of organ damage and maintenance of immune tolerance.
KIM
‐1/
TIM
‐1 is a ...phosphatidylserine receptor that is expressed on epithelial cells and can transform the cells into phagocytes. Here, we demonstrate that
KIM
‐1 phosphorylation and association with p85 results in encapsulation of phagosomes by lipidated
LC
3 in multi‐membrane organelles.
KIM
‐1‐mediated phagocytosis is not associated with increased
ROS
production, and
NOX
inhibition does not block
LC
3 lipidation. Autophagy gene expression is required for efficient clearance of apoptotic cells and phagosome maturation.
KIM
‐1‐mediated phagocytosis leads to pro‐tolerogenic antigen presentation, which suppresses
CD
4 T‐cell proliferation and increases the percentage of regulatory T cells in an autophagy gene‐dependent manner. Taken together, these data reveal a novel mechanism of epithelial biology linking phagocytosis, autophagy and antigen presentation to regulation of the inflammatory response.
Synopsis
image
This study reveals how
KIM
‐1/
TIM
‐1‐mediated phagocytosis down‐modulates inflammation through
ATG
5 and
ULK
1‐dependent autophagic processing of phagocytosed material to
MHC
I and
MHC II
.
Phagocytosis of apoptotic cells by
KIM
‐1, a non‐myeloid phagocytosis receptor, induces
LC
3 lipidation of resulting phagosomes.
Autophagy protein expression,
ATG
5,
ULK
1 and Beclin1, are required for
LC
3 targeting to phagosomes and efficient phagosome acidification and maturation.
KIM
‐1 signaling via the
PI
3 kinase subunit p85 promotes
LC
3 lipidation and accumulation on phagosomes.
Phagocytosed material processed through autophagy is presented to
MHC
I and
MHC II
.
Enhanced antigen presentation resulting from autophagic processing results in a pro‐tolerogenic milieu, reducing the percentage of
CD
4 effector T cells while increasing the percentage of regulatory T cells.
Three different approaches for modelling a semi-batch polymerization reactor using artificial neural networks (ANN) have been investigated. Based on the characteristics of the semi-batch reactor a ...multi-stage strategy is recommended. It divides the whole reaction process into two periods, semi-batch and batch, and further divides the semi-batch part into two sub-periods that are before and after the maximum temperature is reached. Different ANN architectures are used to model the three parts separately. The results demonstrate that the multi-stage approach proposed can be used to estimate difficult-to-measure polymer variables with acceptable accuracy for semi-batch processes. Concentrations of the monomer and the initiator in the reactor are estimated from reactor temperature, feed temperature and the concentration of the initiator in the feed.
TiN
x
(stoichiometric factor,
x=0.1–0.4) films were deposited at a substrate temperature typically of 480°C using an industrial-sized multi-target PVD coating machine. The stoichiometric factor,
x, ...depended on the manner of substrate rotation as well as the reactive gas flow during depositing. In parallel, multiphase compositions of αTi(N), εTi
2N and δTiN were found with hardness values varying from 1500 to 2300 Hk, and 2100 Hk for ε phase Ti
2N. The XRD spectral showed Bragg reflections associated with mixed phase compositions. The almost pure εTi
2N phase was found in the film with
x close to 0.34 and the XRD diffraction pattern of this film perfectly matched the ε phase documented as referred to in the JCPDS file 17-386. SEM and TEM cross-sections exhibited a very fine grain structure for films containing dominant εTi
2N phase. The microstructure of εTi
2N film was extremely homogeneous throughout the complete film growth. The surface of εTi
2N film was surprisingly smooth.
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