Abstract The Chihuahuan Desert averages 247 mm of precipitation annually, 53% of which occurs between July and September. Our objective was to examine movement, activity, pasture use, and watering patterns of Angus x Hereford (AH) and Raramuri Criollo (RC) cows on days with precipitation events (PE, ≥ 1.3 mm rain recorded) vs. days with no precipitation (NP) at the Jornada Experimental Range in southern New Mexico during the summers of 2015, 2016, and 2017. Breeds grazed two adjacent pastures (12A = 1190 ha; 12C = 1165 ha) separately in a crossover design for 4 weeks each year. Seven to nine randomly selected cows per breed were collared with Lotek 3300-LR GPS set to log position every 10 min. An average of 7.6 PE occurred during the 4-week trials each year. On PE days, cows traveled farther (PE = 6.95 vs. NP = 6.43 km, P < 0.01), spent more time grazing (PE = 9.07 vs. NP = 8.76 h/day, P = 0.07) and less time resting (PE = 13.90 vs. NP = 14.38 h/day, P = 0.01), and traveled similar distances from watering points (PE = 0.84 vs. NP = 0.80 km, P = 0.41) compared to NP days. On PE days, cows spent less time within 200 m (PE = 1.32 vs. NP = 2.95 h, P < 0.01) and 100 m (PE = 0.79 vs. NP = 1.90 h, P < 0.01) of drinkers regardless of breed. We detected no rainfall x breed interaction (P > 0.05). Overall, RC cows traveled farther, at higher velocities, spent more time grazing, less time resting, and explored areas with greater radius than AH counterparts (P < 0.05). Precipitation appeared to relax environmental constraints on activity of both breeds, likely due to availability of ephemeral watering sources and a transient drop in temperature.
Abstract Systematic identification of signaling pathways required for the viability of cancer cells will facilitate the development of novel cancer therapies. We used gene essentiality measurements in 726 cancer cell lines to identify selective co-essentiality modules and found a functional ubiquitination cascade containing UBA6, BIRC6, KCMF1 and UBR4, which encode an E1, E2, and two heterodimeric E3 subunits, respectively, as a vulnerability in a subset of epithelial tumors. Suppressing BIRC6 in cancer cell lines that are dependent on this ubiquitination cascade led to a strong reduction in cell fitness in vitro, and to potent tumor regression and metastasis suppression in vivo. Mechanistically, BIRC6 suppression resulted in selective and robust activation of the integrated stress response (ISR) signaling via upregulation of the heme-regulated inhibitor (HRI). Using proteomic profiling, we found that HRI itself is a key degradation target of the UBA6/BIRC6/KCMF1/UBR4 cascade. These observations demonstrate a protein ubiquitination cascade regulating ISR and highlight the potential of this cascade as a novel therapeutic target for a subset of epithelial cancers. Citation Format: Lisa D. Cervia, Tsukasa Shibue, Benjamin Gaeta, Ashir Borah, Lisa Leung, Naomi Li, Nancy Dumont, Alfredo Gonzalez, Nolan Bick, Mariya Kazachkova, Joshua Dempster, John M. Krill-Burger, Namrata Udeshi, Meagan Olive, Steven A. Carr, David E. Root, Federica Piccioni, James M. McFarland, Francisca Vazquez, William C. Hahn. A ubiquitination cascade regulates the integrated stress response and epithelial cancer survival [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2021; 2021 Apr 10-15 and May 17-21. Philadelphia (PA): AACR; Cancer Res 2021;81(13_Suppl):Abstract nr 1950.
<div>Abstract<p>Systematic identification of signaling pathways required for the fitness of cancer cells will facilitate the development of new cancer therapies. We used gene essentiality measurements in 1,086 cancer cell lines to identify selective coessentiality modules and found that a ubiquitin ligase complex composed of <i>UBA6</i>, <i>BIRC6</i>, <i>KCMF1</i>, and <i>UBR4</i> is required for the survival of a subset of epithelial tumors that exhibit a high degree of aneuploidy. Suppressing BIRC6 in cell lines that are dependent on this complex led to a substantial reduction in cell fitness <i>in vitro</i> and potent tumor regression <i>in vivo</i>. Mechanistically, BIRC6 suppression resulted in selective activation of the integrated stress response (ISR) by stabilization of the heme-regulated inhibitor, a direct ubiquitination target of the UBA6/BIRC6/KCMF1/UBR4 complex. These observations uncover a novel ubiquitination cascade that regulates ISR and highlight the potential of ISR activation as a new therapeutic strategy.</p>Significance:<p>We describe the identification of a heretofore unrecognized ubiquitin ligase complex that prevents the aberrant activation of the ISR in a subset of cancer cells. This provides a novel insight on the regulation of ISR and exposes a therapeutic opportunity to selectively eliminate these cancer cells.</p><p><i><a href="https://aacrjournals.org/cancerdiscovery/article/doi/10.1158/2159-8290.CD-22-1440" target="_blank">See related commentary Leli and Koumenis, p. 535</a>.</i></p><p><a href="https://aacrjournals.org/cancerdiscovery/article/doi/10.1158/2159-8290.CD-13-3-ITI" target="_blank">This article is highlighted in the In This Issue feature, p. 517</a></p></div>
<p>Supplementary Figure S1 shows the strategy and result of co-essentiality module identificationfrom the DepMap CRISPR screen dataset and further characterization of the BIRC6 module. Supplementary Figure S2 shows the viability effect of BIRC6 depletion in BIRC6-dependent and -nondependent cancer cells lines as well asin nontransformed cells. Supplementary Figure S3 shows in vitro confirmations of the inducible BIRC6 depletion (by RNAi and CRISPR) systems as well as the effect of BIRC6 depletion on metastasis in xenograft models. Supplementary Figure S4 shows detailed results for the allele competition assay to evaluate the role of the BIR and UBC domains of BIRC6 and further evidence for the physical assembly of BIRC6 ubiquitin ligase complex. Supplementary Figure S5 shows evidence for the selective activation of the p-eIF2alpha/ATF4 arm of UPR (or ISR) upon BIRC6 depletion. Supplementary Figure S6 shows evidence supporting the importance of HRI-mediated ISR activation as a mechanism underlying the loss of viability caused by BIRC6 complex suppression. Supplementary Figure S7 shows evidence for the direct regulation of HRI ubiquitination and degradation by the BIRC6 complex Supplementary Figure S8 shows elevated expression of HRI mRNA in the tumor samples compared to the normal samples in large-scale datasets (TCGA, TARGET and GTEx). Supplementary Figure S9 shows the inefficiency of the gene expression and copy number of the functionally related genes in predicting BIRC6 dependency. Supplementary Table S1 lists the top 50 co-essentiality modules identified in the analysis explained in Supplementary Figure S1A. Supplementary Table S2 lists the key DNA and RNA sequences used in this study.</p>
Abstract High throughput drug screening is an established approach to investigate tumor biology and identify therapeutic leads. Traditional platforms use two-dimensional cultures which do not accurately reflect the biology of human tumors. More clinically relevant model systems such as three-dimensional tumor organoids can be difficult to scale and screen. Manually seeded organoids coupled to destructive endpoint assays allow for the characterization of treatment response, but do not capture transitory changes and intra-sample heterogeneity underlying clinically observed resistance to therapy. We present a pipeline to generate bioprinted tumor organoids linked to label-free, time-resolved imaging via high-speed live cell interferometry (HSLCI) and machine learning-based quantitation of individual organoids. Bioprinting cells gives rise to 3D structures with unaltered tumor histology and gene expression profiles. HSLCI imaging in tandem with machine learning-based segmentation and classification tools enables accurate, label-free parallel mass measurements for thousands of organoids. We demonstrate that this strategy identifies organoids transiently or persistently sensitive or resistant to specific therapies, information that could be used to guide rapid therapy selection.
Our objective was to describe key phenotypic characteristics of a population of Rarámuri Criollo (RC) cattle introduced from the Copper Canyon of Chihuahua, México into the Southwestern United States almost two decades ago. We recorded 26 phenotypic traits of 37 RC individuals including mature cows, first-calf heifers, and mature bulls raised at the USDA-ARS Jornada Experimental Range in southern New Mexico. This herd of RC cattle exhibited intermediate body sizes compared to the smaller Corriente and larger Texas Longhorn and Florida Cracker cattle. Coat colors were similar to those described for other Criollo biotypes but horn shape and size of RC appear to be different than that of other US-based Criollo breeds. Though smaller than commercial beef breeds, RC cattle appear to be well-matched to the Southwestern US environments as evidenced by previous studies that evaluated their grazing behavior, weight gains, and carcass quality. Rarámuri Criollo cattle are a genetic resource whose conservation could be critically important for climate change adaptation of ranches in the desert Southwest.
Abstract Host genetics is a key determinant of COVID-19 outcomes. Previously, the COVID-19 Host Genetics Initiative genome-wide association study used common variants to identify multiple loci associated with COVID-19 outcomes. However, variants with the largest impact on COVID-19 outcomes are expected to be rare in the population. Hence, studying rare variants may provide additional insights into disease susceptibility and pathogenesis, thereby informing therapeutics development. Here, we combined whole-exome and whole-genome sequencing from 21 cohorts across 12 countries and performed rare variant exome-wide burden analyses for COVID-19 outcomes. In an analysis of 5,085 severe disease cases and 571,737 controls, we observed that carrying a rare deleterious variant in the SARS-CoV-2 sensor toll-like receptor TLR7 (on chromosome X) was associated with a 5.3-fold increase in severe disease (95% CI: 2.75-10.05, p=5.41×10 −7 ). This association was consistent across sexes. These results further support TLR7 as a genetic determinant of severe disease and suggest that larger studies on rare variants influencing COVID-19 outcomes could provide additional insights. Author Summary COVID-19 clinical outcomes vary immensely, but a patient’s genetic make-up is an important determinant of how they will fare against the virus. While many genetic variants commonly found in the populations were previously found to be contributing to more severe disease by the COVID-19 Host Genetics Initiative, it isn’t clear if more rare variants found in less individuals could also play a role. This is important because genetic variants with the largest impact on COVID-19 severity are expected to be rarely found in the population, and these rare variants require different technologies to be studies (usually whole-exome or whole-genome sequencing). Here, we combined sequencing results from 21 cohorts across 12 countries to perform a rare variant association study. In an analysis comprising 5,085 participants with severe COVID-19 and 571,737 controls, we found that the gene for toll-like receptor 7 ( TLR7 ) on chromosome X was an important determinant of severe COVID-19. Importantly, despite being found on a sex chromosome, this observation was consistent across both sexes.
