APC15 drives the turnover of MCC-CDC20 to make the spindle assembly checkpoint responsive to kinetochore attachment

Faithful chromosome segregation during mitosis depends on the Spindle Assembly Checkpoint (SAC) that monitors kinetochore attachment to the mitotic spindle. Unattached kinetochores generate mitotic checkpoint proteins complexes (MCCs) that bind and inhibit the Anaphase Promoting Complex/Cyclosome (APC/C). How the SAC proficiently inhibits the APC/C but still allows its rapid activation when the last kinetochore attaches to the spindle is important to understand how cells maintain genomic stability. We show that the APC/C subunit APC15 is required for the turnover of the APC/C co-activator Cdc20 and release of MCCs during SAC signalling but not for APC/C activity per se. In the absence of APC15, MCCs and ubiquitylated Cdc20 remain ‘locked’ onto the APC/C, which prevents the ubiquitylation and degradation of Cyclin B1 when the SAC is satisfied. We conclude that APC15 mediates the constant turnover of Cdc20 and MCCs on the APC/C to allow the SAC to respond to the attachment state of kinetochores

GZMKhigh CD8+ T effector memory cells are associated with CD15high neutrophil abundance in non-metastatic colorectal tumors and predict poor clinical outcome.

CD8(+) T cells are a major prognostic determinant in solid tumors, including colorectal cancer (CRC). However, understanding how the interplay between different immune cells impacts on clinical outcome is still in its infancy. Here, we describe that the interaction of tumor infiltrating neutrophils expressing high levels of CD15 with CD8(+) T effector memory cells (T(EM)) correlates with tumor progression. Mechanistically, stromal cell-derived factor-1 (CXCL12/SDF-1) promotes the retention of neutrophils within tumors, increasing the crosstalk with CD8(+) T cells. As a consequence of the contact-mediated interaction with neutrophils, CD8(+) T cells are skewed to produce high levels of GZMK, which in turn decreases E-cadherin on the intestinal epithelium and favors tumor progression. Overall, our results highlight the emergence of GZMK(high) CD8(+) T(EM) in non-metastatic CRC tumors as a hallmark driven by the interaction with neutrophils, which could implement current patient stratification and be targeted by novel therapeutics

E-Cadherin Is Required for Centrosome and Spindle Orientation in Drosophila Male Germline Stem Cells

Many adult stem cells reside in a special microenvironment known as the niche, where they receive essential signals that specify stem cell identity. Cell-cell adhesion mediated by cadherin and integrin plays a crucial role in maintaining stem cells within the niche. In Drosophila melanogaster, male germline stem cells (GSCs) are attached to niche component cells (i.e., the hub) via adherens junctions. The GSC centrosomes and spindle are oriented toward the hub-GSC junction, where E-cadherin-based adherens junctions are highly concentrated. For this reason, adherens junctions are thought to provide a polarity cue for GSCs to enable proper orientation of centrosomes and spindles, a critical step toward asymmetric stem cell division. However, understanding the role of E-cadherin in GSC polarity has been challenging, since GSCs carrying E-cadherin mutations are not maintained in the niche. Here, we tested whether E-cadherin is required for GSC polarity by expressing a dominant-negative form of E-cadherin. We found that E-cadherin is indeed required for polarizing GSCs toward the hub cells, an effect that may be mediated by Apc2. We also demonstrated that E-cadherin is required for the GSC centrosome orientation checkpoint, which prevents mitosis when centrosomes are not correctly oriented. We propose that E-cadherin orchestrates multiple aspects of stem cell behavior, including polarization of stem cells toward the stem cell-niche interface and adhesion of stem cells to the niche supporting cells

Longitudinal gut microbiome changes in immune checkpoint blockade-treated advanced melanoma

Multiple clinical trials targeting the gut microbiome are being conducted to optimize treatment outcomes for immune checkpoint blockade (ICB). To improve the success of these interventions, understanding gut microbiome changes during ICB is urgently needed. Here through longitudinal microbiome profiling of 175 patients treated with ICB for advanced melanoma, we show that several microbial species-level genome bins (SGBs) and pathways exhibit distinct patterns from baseline in patients achieving progression-free survival (PFS) of 12 months or longer (PFS ≥12) versus patients with PFS shorter than 12 months (PFS <12). Out of 99 SGBs that could discriminate between these two groups, 20 were differentially abundant only at baseline, while 42 were differentially abundant only after treatment initiation. We identify five and four SGBs that had consistently higher abundances in patients with PFS ≥12 and <12 months, respectively. Constructing a log ratio of these SGBs, we find an association with overall survival. Finally, we find different microbial dynamics in different clinical contexts including the type of ICB regimen, development of immune-related adverse events and concomitant medication use. Insights into the longitudinal dynamics of the gut microbiome in association with host factors and treatment regimens will be critical for guiding rational microbiome-targeted therapies aimed at enhancing ICB efficacy

Intraperitoneal drain placement and outcomes after elective colorectal surgery: international matched, prospective, cohort study

Despite current guidelines, intraperitoneal drain placement after elective colorectal surgery remains widespread. Drains were not associated with earlier detection of intraperitoneal collections, but were associated with prolonged hospital stay and increased risk of surgical-site infections.Background Many surgeons routinely place intraperitoneal drains after elective colorectal surgery. However, enhanced recovery after surgery guidelines recommend against their routine use owing to a lack of clear clinical benefit. This study aimed to describe international variation in intraperitoneal drain placement and the safety of this practice. Methods COMPASS (COMPlicAted intra-abdominal collectionS after colorectal Surgery) was a prospective, international, cohort study which enrolled consecutive adults undergoing elective colorectal surgery (February to March 2020). The primary outcome was the rate of intraperitoneal drain placement. Secondary outcomes included: rate and time to diagnosis of postoperative intraperitoneal collections; rate of surgical site infections (SSIs); time to discharge; and 30-day major postoperative complications (Clavien-Dindo grade at least III). After propensity score matching, multivariable logistic regression and Cox proportional hazards regression were used to estimate the independent association of the secondary outcomes with drain placement. Results Overall, 1805 patients from 22 countries were included (798 women, 44.2 per cent; median age 67.0 years). The drain insertion rate was 51.9 per cent (937 patients). After matching, drains were not associated with reduced rates (odds ratio (OR) 1.33, 95 per cent c.i. 0.79 to 2.23; P = 0.287) or earlier detection (hazard ratio (HR) 0.87, 0.33 to 2.31; P = 0.780) of collections. Although not associated with worse major postoperative complications (OR 1.09, 0.68 to 1.75; P = 0.709), drains were associated with delayed hospital discharge (HR 0.58, 0.52 to 0.66; P &lt; 0.001) and an increased risk of SSIs (OR 2.47, 1.50 to 4.05; P &lt; 0.001). Conclusion Intraperitoneal drain placement after elective colorectal surgery is not associated with earlier detection of postoperative collections, but prolongs hospital stay and increases SSI risk

Sister chromatid tension and the spindle assembly checkpoint

The spindle assembly checkpoint (SAC) is a feedback control system that monitors the state of kinetochore/microtubule attachment during mitosis and halts cell cycle progression until all chromosomes are properly aligned at the metaphase plate. The state of chromosome-microtubule attachment is implicated as a crucial factor in the checkpoint response. On the contrary, lack of tension in the centromere-kinetochore region of sister chromatids has been shown to regulate a pathway of correction of undesired chromosome-microtubule connections, while the presence of tension is believed to promote the stabilization of attachments. We discuss how tension-sensitive phenomena, such as attachment correction and stabilization, relate to the SAC and we speculate on the existence of a single pathway linking error correction and SAC activation

The role of proteases in fibrinonectin matrix remodeling in thyroid epithelial cell monolayer cultures

Fischer rat thyroid (FRT) cells organize a matrix of extracellular fibronectin (FN) fibrils, which undergoes extensive remodeling according to cell culture confluence. In non-confluent cells FN forms a fibrillar array associated with the ventral cell surface. However, basal FN is progressively removed in confluent cultures and substituted by non-fibrillar FN deposits at lateral cell domains in regions of cell-cell contacts. FRT cells secrete and expose on the plasma membrane the tissue-type plasminogen activator and, in serum-free cultures, plasminogen induces a rapid loss of FN fibrils. Incubation with plasmin inhibitors greatly reduces this effect. FRT cells also express annexin II, a plasminogen receptor, suggesting that plasmin activity is associated with the pericellular enviroment. This is in agreement with the observation that a great reduction in FN degradation is observed if the cells are pre-incubated with carboxypeptidase B, which prevents plasminogen binding to the cells. A gelatinolytic activity with a molecular weigth equivalent to MMP-2 has been demonstrated by zymography of culture media, and the presence of MMP-2 and MT1-MMP on the cell plasma membrane has been detected by immunofluorescence. These results indicate that in the FN remodeling process, occurring during FRT epithelium maturation, both plasmin-dependent (tPA activated) and plasmin-independent proteolytic activities are involved