Mutation of the TP53 tumor suppressor gene is the most common genetic alteration in cancer, and almost 1000 alleles have been identified in human tumors. While virtually all TP53 mutations are ...thought to compromise wild type p53 activity, the prevalence and recurrence of missense TP53 alleles has motivated countless research studies aimed at understanding the function of the resulting mutant p53 protein. The data from these studies support three distinct, but perhaps not necessarily mutually exclusive, mechanisms for how different p53 mutants impact cancer: first, they lose the ability to execute wild type p53 functions to varying degrees; second, they act as a dominant negative (DN) inhibitor of wild type p53 tumor-suppressive programs; and third, they may gain oncogenic functions that go beyond mere p53 inactivation. Of these possibilities, the gain of function (GOF) hypothesis is the most controversial, in part due to the dizzying array of biological functions that have been attributed to different mutant p53 proteins. Herein we discuss the current state of understanding of TP53 allele variation in cancer and recent reports that both support and challenge the p53 GOF model. In these studies and others, researchers are turning to more systematic approaches to profile TP53 mutations, which may ultimately determine once and for all how different TP53 mutations act as cancer drivers and whether tumors harboring distinct mutations are phenotypically unique. From a clinical perspective, such information could lead to new therapeutic approaches targeting the effects of different TP53 alleles and/or better sub-stratification of patients harboring TP53 mutant cancers.
The primary cilium is an essential organizing center for signal transduction, and ciliary defects cause congenital disorders known collectively as ciliopathies.1–3 Primary cilia form by two pathways ...that are employed in a cell-type- and tissue-specific manner: an extracellular pathway in which the cilium grows out from the cell surface and an intracellular pathway in which the nascent cilium first forms inside the cell.4–8 After exposure to the external environment, cilia formed via the intracellular pathway may have distinct functional properties, as they often remain recessed within a ciliary pocket.9,10 However, the precise mechanism of intracellular ciliogenesis and its relatedness to extracellular ciliogenesis remain poorly understood. Here we show that Rab34, a poorly characterized GTPase recently linked to cilia,11–13 is a selective mediator of intracellular ciliogenesis. We find that Rab34 is required for formation of the ciliary vesicle at the mother centriole and that Rab34 marks the ciliary sheath, a unique sub-domain of assembling intracellular cilia. Rab34 activity is modulated by divergent residues within its GTPase domain, and ciliogenesis requires GTP binding and turnover by Rab34. Because Rab34 is found on assembly intermediates that are unique to intracellular ciliogenesis, we tested its role in the extracellular pathway used by polarized MDCK cells. Consistent with Rab34 acting specifically in the intracellular pathway, MDCK cells ciliate independently of Rab34 and its paralog Rab36. Together, these findings establish that different modes of ciliogenesis have distinct molecular requirements and reveal Rab34 as a new GTPase mediator of ciliary membrane biogenesis.
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•Rab34 is required for ciliary vesicle formation during ciliogenesis•Rab34 localizes to nascent ciliary membranes and marks the ciliary sheath•GTP binding and hydrolysis by Rab34 are required for intracellular ciliogenesis•Rab34 is dispensable for the extracellular ciliogenesis pathway used by MDCK cells
Different cell types form primary cilia through different assembly pathways. Ganga et al. show that the GTPase Rab34 is needed for ciliary membrane formation in the intracellular ciliogenesis pathway, but not the extracellular pathway. During intracellular ciliogenesis, Rab34 is recruited to nascent cilia and localizes to the ciliary sheath.
studies of cancer biology and assessment of therapeutic efficacy are critical to advancing cancer research and ultimately improving patient outcomes. Murine cancer models have proven to be an ...invaluable tool in pre-clinical studies. In this context, multi-parameter flow cytometry is a powerful method for elucidating the profile of immune cells within the tumor microenvironment and/or play a role in hematological diseases. However, designing an appropriate multi-parameter panel to comprehensively profile the increasing diversity of immune cells across different murine tissues can be extremely challenging.
To address this issue, we designed a panel with 13 fixed markers that define the major immune populations -referred to as the backbone panel- that can be profiled in different tissues but with the option to incorporate up to seven additional fluorochromes, including any marker specific to the study in question.
This backbone panel maintains its resolution across different spectral flow cytometers and organs, both hematopoietic and non-hematopoietic, as well as tumors with complex immune microenvironments.
Having a robust backbone that can be easily customized with pre-validated drop-in fluorochromes saves time and resources and brings consistency and standardization, making it a versatile solution for immuno-oncology researchers. In addition, the approach presented here can serve as a guide to develop similar types of customizable backbone panels for different research questions requiring high-parameter flow cytometry panels.
Although many tumors regress in response to neoadjuvant chemotherapy, residual tumor cells are detected in most cancer patients post-treatment. These residual tumor cells are thought to remain ...dormant for years before resuming growth, resulting in tumor recurrence. Considering that recurrent tumors are most often responsible for patient mortality, there exists an urgent need to study signaling pathways that drive tumor dormancy/recurrence. We have developed an in vitro model of tumor dormancy/recurrence. Short-term exposure of tumor cells (breast or prostate) to chemotherapy at clinically relevant doses enriches for a dormant tumor cell population. Several days after removing chemotherapy, dormant tumor cells regain proliferative ability and establish colonies, resembling tumor recurrence. Tumor cells from "recurrent" colonies exhibit increased chemotherapy resistance, similar to the therapy resistance of recurrent tumors in cancer patients. Previous studies using long-term chemotherapy selection models identified acquired mutations that drive tumor resistance. In contrast, our short term chemotherapy exposure model enriches for a slow-cycling, dormant, chemo-resistant tumor cell sub-population that can resume growth after drug removal. Studying unique signaling pathways in dormant tumor cells enriched by short-term chemotherapy treatment is expected to identify novel therapeutic targets for preventing tumor recurrence.
In mammals, 10 TLRs recognize conserved pathogen-associated molecular patterns, resulting in the induction of inflammatory innate immune responses. One of these, TLR9, is activated intracellularly by ...bacterial DNA and synthetic oligodeoxynucleotides (ODN), containing unmethylated CpG dinucleotides. Following treatment with CpG ODN, TLR9 is found in lysosome-associated membrane protein type 1-positive lysosomes, and we asked which intracellular compartment contains TLR9 before CpG exposure. Surprisingly, we found by microscopy and supporting biochemical evidence that both transfected and endogenously expressed human TLR9 is retained in the endoplasmic reticulum. By contrast, human TLR4 trafficked to the cell surface, indicating that endoplasmic reticulum retention is not a property common to all TLRs. Because TLR9 is observed in endocytic vesicles following exposure to CpG ODN, our data indicate that a special mechanism must exist for translocating TLR9 to the signaling compartments that contain the CpG DNA.
You can continue to thoughtlessly pollute, Ruskin warns his readers, but in so doing you will destroy the earth and end your own existence. Six years earlier, in 1865, Ruskin coined the term ...“dis-ease” to denote a clear link between ill-being and environmental detachment. He yoked physical and mental health, elucidating “how literally that word Dis-Ease, the Negation and impossibility of Ease, expressed the entire moral state of our English Industry and its Amusements!” (“Of Kings’ Treasuries” 282). For Ruskin, nineteenth-century mills and factories, despite promising consumer satisfaction, made comfort impossible by endlessly producing frivolous, disposable goods, and thus waste. This needless consumption, a symptom of industry, produced an ignorance of true needs. Dis-ease, mental and bodily discomfort, resulted from alienation from the ecosystem, the networks of dependence between all species, and that estrangement blinded human beings to their actual role in the environment. While Ruskin focused on urban toxicity, the toxic ideological separation between humans and their environment impacted all spaces, a concern that several Victorian writers raised decades earlier than he did. This article traces the salutary cultural anxiety over improper sanitation and contaminants in two popular mid-nineteenth-century novels that demonstrate the effects of anthropogenic pollution in urban and rural environments, respectively. Published almost exactly one year apart, both Elizabeth Gaskell's Mary Barton (1848) and Charlotte Brontë’s Jane Eyre (1847) invoke what I call eco-consciousness in their description of urban and rural filth, portrayed as both visible and invisible toxins. Gaskell uncovers urban pollution in plain sight, going beyond smell to expose the causes of toxicity, while Brontë challenges the belief in the country as a safe haven from pollution, going beyond beauty to expose rural toxicity. Characters suffer physical disease and mental dis-ease resulting from a poor understanding of ecological relationships. Reading Jane Eyre alongside Mary Barton accentuates Brontë’s use of eco-consciousness to expose the hidden dangers of rural pollution that resulted from the very types of urban toxicity that Gaskell identifies.
There is increasing interest and evidence for the "flipped classroom" as a learner-centered approach to promote active engagement and enhance translation of knowledge into practice. However, there is ...little published work regarding application of the pedagogical approach to continuing education (CE) for advanced practice providers. The goal of CE is for providers to stay up-to-date with current best practices and skills, yet most CE activities employ teacher-centered, lecture-based methods focused on transmission of knowledge rather than application of knowledge. The purpose of this article is to describe successful use of the flipped classroom in a recent CE offering for emergency nurse practitioners, and to support further exploration of interactive approaches in CE.
Anticancer drug development campaigns often fail due to an incomplete understanding of the therapeutic index differentiating the efficacy of the agent against the cancer and its on-target toxicities ...to the host. To address this issue, we established a versatile preclinical platform in which genetically defined cancers are produced using somatic tissue engineering in transgenic mice harboring a doxycycline-inducible short hairpin RNA against the target of interest. In this system, target inhibition is achieved by the addition of doxycycline, enabling simultaneous assessment of efficacy and toxicity in the same animal. As proof of concept, we focused on CDK9—a cancer target whose clinical development has been hampered by compounds with poorly understood target specificity and unacceptable toxicities. We systematically compared phenotypes produced by genetic Cdk9 inhibition to those achieved using a recently developed highly specific small molecule CDK9 inhibitor and found that both perturbations led to robust antitumor responses. Remarkably, nontoxic levels of CDK9 inhibition could achieve significant treatment efficacy, and dose-dependent toxicities produced by prolonged CDK9 suppression were largely reversible upon Cdk9 restoration or drug withdrawal. Overall, these results establish a versatile in vivo target validation platform that can be employed for rapid triaging of therapeutic targets and lend support to efforts aimed at advancing CDK9 inhibitors for cancer therapy.
"Morbidly adherent placenta" is a term that describes the continuum of placenta accreta, increta, and percreta. The incidence of this type of abnormal placentation has increased significantly over ...recent decades. The reason is probably multifactorial but, partly, because of factors such as the increasing number of cesarean births. Women at greatest risk are those who have myometrial damage caused by a previous cesarean birth, with either anterior or posterior placenta previa overlying the uterine scar. This condition poses significant risks of morbidity and/or mortality to the pregnant woman and her fetus. A multidisciplinary approach to care throughout pregnancy is essential. This article describes the classification of morbidly adherent placenta, risk factors, methods of diagnosis, potential maternal and fetal complications, and intrapartum clinical management strategies to optimize outcomes.
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