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Search indexed bioRxiv preprints in genomics, neuroscience, cell biology and bioinformatics. Read source abstracts and check manuscript versions; preprints are not peer reviewed.

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bioRxiv2018

Prolonged culture of primary human keratinocytes isolated from suction blisters

Keratinocytes are the most abundant cell type in the epidermis. They prevent desiccation and provide immunological and barrier defense against potential pathogens such as Staphylococcus aureus and Candida albicans. The study of this first line of immune defense is hindered by invasive isolation methods and insufficient techniques for long-term passage of primary keratinocytes in vitro. Primary keratinocytes have been successfully isolated from blister roofs induced by negative pressure, which separates the epidermis from the dermis in vivo in human subjects. This method allows collection of pure epidermal cells without dermal contamination in a minimally invasive manner. However, the isolated keratinocytes differentiate and senesce when cultured in vitro beyond five passages. Here, we present evidence that the Rho kinase (ROCK) inhibitor Y-27632 can be used to effectively increase the proliferative capabilities of keratinocytes isolated using the suction blister method, similar to what has been previously reported for primary keratinocytes isolated using alternative methods. We show that the increase in passage number is directly correlated to delayed differentiation, and that cells passaged long term with the inhibitor retain their ability to stratify in organotypic raft cultures and respond to cytokine treatment; additionally, the late passage cells have a heterogeneous mix of differentiated and non-differentiated cells which may be predicted by a ratio of select differentiation markers. The described method presents a minimally invasive procedure for keratinocyte isolation and prolonged culture that allows analysis of keratinocyte function in both healthy volunteers and patients with dermatologic diseases.

cell biology↗

bioRxiv2018

WNT4 and WNT3A activate cell autonomous Wnt signaling independent of secretion

The enzyme PORCN is considered essential for Wnt secretion and signaling, however, we observed PORCN inhibition did not phenocopy the effects of WNT4 knockdown in WNT4-dependent breast cancer cells. This suggests a unique relationship between PORCN and WNT4 signaling. To examine the role of PORCN in WNT4 signaling, WNT4 or WNT3A were over-expressed in breast and ovarian cancer, and fibrosarcoma cell lines. Conditioned medium from these lines, and co-culture systems, were used to assess the dependence of Wnt secretion and activity on critical Wnt secretion proteins PORCN and WLS. We observed that WLS was universally required for Wnt secretion and paracrine signaling. In contrast, the dependence of WNT3A secretion and activity on PORCN varied across cell lines, and WNT4 secretion was PORCN-independent in all models. Surprisingly, WNT4 did not present paracrine activity in any tested context. Absent the expected paracrine activity of secreted WNT4, we identified cell autonomous Wnt signaling activation by WNT4 and WNT3A, independent of PORCN or secretion. The PORCN-independent, cell-autonomous Wnt signaling demonstrated herein may be critical in WNT4-driven cellular contexts, or those that are otherwise considered to have dysfunctional Wnt signaling.\n\nSummary StatementWnt proteins can mediate an atypical mode of cell-autonomous signaling, distinct from paracrine signaling, that is independent of both palmitoylation by PORCN and Wnt secretion.

cell biology↗

bioRxiv2018

Extracellular Degradation into Adenosine and the Activities of Adenosine Kinase and AMPK Mediate Extracellular NAD+-produced increases in the Adenylate Pool of BV2 Microglia under Basal Conditions

Cumulating evidence has indicated NAD+ deficiency as a common central pathological factor of multiple diseases and aging. NAD+ supplement is highly protective in various disease and aging models, while two key questions remain unanswered: 1) Does extracellular NAD+ also produce its effects through its degradation product adenosine? 2) Does extracellular NAD+ produce the protective effects by affecting cells under pathological insults only, or by affecting both normal cell and cells under pathological insults? Since extracellular NAD+ can be degraded into adenosine, and endogenous adenosine levels are in the nanomolar range under physiological conditions, extracellular NAD+ may produce its effects through its degradation into adenosine. In this study we used BV2 microglia as a cellular model to test our hypothesis that NAD+ treatment can increase the intracellular adenylate pool under basal conditions through its extracellular degradation into adenosine. Our study has shown that extracellular NAD+ increases the adenylate pool of BV2 microglia under basal conditions through its degradation into adenosine that enters the cells through equilibrative nucleoside transporters. The intracellular adenosine is converted to AMP by adenosine kinase, which increases intracellular ATP by both activating AMPK and increasing ADP that drives mitochondrial FoF1-ATP synthase. Collectively, our study has suggested that extracellular NAD+ can enhance defensive capacity of normal cells through a novel pathway, which includes extracellular NAD+ degradation into adenosine and the activities of adenosine kinase and AMPK. Our findings have also suggested that NAD+ administration in various disease and aging models may significantly affect the microglia under basal conditions.

cell biology↗

bioRxiv2018

The in situ structures of mono-, di-, and tri-nucleosomes in human heterochromatin

The in situ 3-D organization of chromatin at the nucleosome and oligonucleosome levels is unknown. Here we use cryo-electron tomography (cryo-ET) to determine the in situ structures of HeLa nucleosomes, which have canonical core structures and asymmetric, flexible linker DNA. Subtomogram remapping suggests that sequential nucleosomes in heterochromatin follow irregular paths at the oligonucleosome level. This basic principle of higher-order repressive chromatin folding is compatible with the conformational variability of the two linker DNAs at the single-nucleosome level.

cell biology↗

bioRxiv2018

Gamete fusion rapidly reconstitutes a bi-partite transcription factor to block re-fertilization

The ploidy cycle, integral to sexual reproduction, requires not only meiosis to halve the number of chromosomes, but also mechanisms that ensure zygotes are formed by exactly two partners1-5. During sexual reproduction of the fungal model organism Schizosaccharomyces pombe, haploid P- and M-cells normally fuse to form a diploid zygote that immediately enters meiosis6. Here, we reveal that fast post-fusion reconstitution of a bi-partite transcription factor actively blocks re-fertilization. We first identify mutants that undergo transient cell fusion involving cytosol exchange but not karyogamy, and show this drives distinct cell fates in the two gametes: The P-partner undergoes lethal, haploid meiosis while the M-cell persists in mating. Consistently, we find that the zygotic transcription that drives meiosis is initiated rapidly only from the P-parental genome, even in wild type cells. This asymmetric gene expression depends on a bi-partite complex formed post-fusion between the nuclear P-cell-specific homeobox protein Pi and a cytosolic M-specific peptide Mi7,8, which is captured by Pi in the P-nucleus. Zygotic transcription is thus poised to initiate in the P-nucleus as fast as Mi reaches it. The asymmetric nuclear accumulation is inherent to the transcription factor design, and is reconstituted by a pair of synthetic interactors, one localized to the nucleus of one gamete and the other in the cytosol of its partner. Strikingly, imposing a delay in zygotic transcription, by postponing Mi expression or deleting its transcriptional target in the P-genome, leads to zygotes fusing with additional gametes, thus forming polyploids and eventually aneuploid progeny. We further show that the signaling cascade to block re-fertilization shares components with, but bifurcates from, meiotic induction9-11. Thus, cytoplasmic connection upon gamete fusion leads to rapid reconstitution of a bi-partite transcription factor in one partner to block re-fertilization and induce meiosis, thus ensuring genome maintenance during sexual reproduction.

cell biology↗

bioRxiv2018

Transmembrane Protein 135 (TMEM135) is a Liver X Receptor Target Gene that Mediates an Auxiliary Peroxisome Matrix Protein Import Pathway

The liver x receptors (LXRs) are key regulators of systemic lipid metabolism. We determined whether transmembrane protein 135 (TMEM135) is an LXR target gene and its physiologic function. An LXR agonist increased TMEM135 mRNA and protein in human hepatocyte and macrophage cell lines, which was prevented by LXR knockdown. The human TMEM135 promoter contains an LXR response element that bound the LXRs via EMSA and ChIP, and mediated LXR-induced transcription in reporter assays. Knockdown of TMEM135 in HepG2 cells caused triglyceride accumulation despite reduced lipogenic gene expression, indicating a potential role in {beta}-oxidation. To determine physiologic importance, TMEM135 was knocked-down via siRNA in livers of fed and fasted C57BL/6 mice. Fasting increased hepatic fatty acid and NADH concentrations in control mice, consistent with increased fatty acid uptake and {beta}-oxidation. However, in fasted TMEM135 knockdown mice, there was a further significant increase in hepatic fatty acid concentrations and a significant decrease in NADH, indicating an impairment in {beta}-oxidation by peroxisomes and/or mitochondria. Conversely, hepatic ketones tended to increase in fasted TMEM135 knockdown compared to control mice, and because ketogenesis is exclusively dependent on mitochondrial {beta}-oxidation, this indicates peroxisomal {beta}-oxidation was impaired in knockdown mice. Localization studies demonstrated that TMEM135 co-localized with peroxisomes but not mitochondria. Mechanistically, proteomic and Western blot analyses indicated that TMEM135 regulates concentrations of matrix enzymes within peroxisomes. In conclusion, TMEM135 is a novel LXR target gene in humans that mediates peroxisomal metabolism, and thus TMEM135 may be a therapeutic target for metabolic disorders associated with peroxisome dysfunction.

cell biology↗

bioRxiv2018

The cargo receptor SURF4 promotes the efficient cellular secretion of PCSK9

Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a secreted protein that plays an important role in regulating plasma cholesterol and cardiovascular disease risk. PCSK9 secretion uniquely depends on the cytoplasmic COPII protein SEC24A, suggesting the presence of a transmembrane ER cargo receptor mediating this interaction. Here, we report a novel approach that combines proximity-dependent biotinylation and proteomics together with genome-scale CRISPR screening to identify proteins that facilitate the efficient secretion of PCSK9 heterologously expressed in HEK293T cells. We first identified 35 candidate proteins that were labeled by BirA* fusions to PCSK9 and either COPII component SAR1A or SAR1B. We then performed genome-scale pooled CRISPR mutagenesis to identify genes whose perturbation resulted in intracellular accumulation of PCSK9-eGFP but not the control A1AT-mCherry. The 4 most enriched sgRNAs in this screen all targeted SURF4, a homologue of the yeast endoplasmic reticulum (ER) cargo receptor Erv29p and the only candidate also identified by proximity-dependent biotinylation. The functional contribution of SURF4 to PCSK9 secretion was confirmed with multiple independent SURF4-targeting sgRNAs, clonal SURF4-deficient cell lines, and functional rescue with SURF4 cDNA. Compatible with a function of SURF4 as a cargo receptor for PCSK9, fluorescence microscopy localized SURF4 to the early secretory pathway, coimmunoprecipitation revealed a physical interaction between SURF4 and PCSK9, and SURF4 deletion resulted in decreased extracellular secretion of PCSK9 and PCSK9 accumulation in the ER. Taken together, these findings support a model in which SURF4 functions as an ER cargo receptor for the efficient cellular secretion of PCSK9.

cell biology↗

bioRxiv2018

Chromomycin A2 potently inhibits glucose-stimulated insulin secretion from pancreatic beta cells.

Enhancers or inhibitors of insulin secretion could become therapeutics as well as lead to the identification of requisite {beta}-cell regulatory pathways and increase our understanding of pancreatic islet function. Toward this goal, we previously used an insulin-linked luciferase that is co-secreted with insulin in MIN6 {beta}-cells to perform a high-throughput natural product screen for chronic effects on glucose-stimulated insulin secretion. Using multiple phenotypic analyses, we identified that one of the top natural product hits, chromomycin A2 (CMA2), potently inhibited insulin secretion through at least three mechanisms: disruption of Wnt signaling, interfering with {beta}-cell gene expression, and suppression of triggering calcium (Ca2+) influx. Chronic treatment with CMA2 largely ablated glucose-stimulated insulin secretion even post-washout, but did not inhibit glucose-stimulated generation of ATP or Ca2+ influx. However, by using the KATP channel-opener diazoxide, we uncovered defects in depolarization-induced Ca2+ influx which may contribute to the suppressed secretory response. Glucose-responsive ERK1/2 and S6 phosphorylation were also disrupted by chronic CMA2 treatment. The FUSION bioinformatic database indicated that the phenotypic effects of CMA2 clustered with a number of Wnt/GSK3 pathway-related genes. Consistently, CMA2 decreased GSK3 phosphorylation and suppressed activation of a {beta}-catenin activity reporter. CMA2 and a related compound mithramycin are described to have DNA-interaction properties, possibly abrogating transcription factor binding to critical {beta}-cell gene promoters. We observed that CMA2, but not mithramycin, suppressed expression of PDX1 and UCN3. However, neither expression of INSI/II nor insulin content was affected by chronic CMA2. The mechanisms of CMA2-induced insulin secretion defects may involve components both proximal and distal to Ca2+ influx. Therefore, CMA2 is an example of a chemical that can simultaneously disrupt {beta}-cell function through both non-cytotoxic and cytotoxic mechanisms. Future applications of CMA2 and similar aureolic acid analogs for disease therapies should consider the potential impacts on pancreatic islet function.

cell biology↗

bioRxiv2018

Differential GAP requirement for Cdc42-GTP polarization during proliferation and sexual reproduction

The formation of a local zone of Cdc42 GTPase activity, which governs cell polarization in many cell types, requires not only local activation but also switch-off mechanisms. Here we identify Rga3, a paralog of Rga4, as a novel Cdc42 GTPase activating protein (GAP) in the fission yeast S. pombe. Contrary to Rga4, Rga3 localizes with Cdc42-GTP to sites of polarity. Rga3 is dispensable for cell polarization during mitotic growth, but limits the lifetime of unstable Cdc42-GTP patches that underlie cell pairing during sexual reproduction, masking a partly compensatory patch wandering motion. In consequence, cells lacking rga3 hyperpolarize and loose out in mating competition. Rga3 synergizes with the Cdc42 GAPs Rga4 and Rga6 to restrict Cdc42-GTP zone sizes during mitotic growth. Surprisingly, triple mutant cells, which are almost fully round, retain pheromone-dependent dynamic polarization of Cdc42-GTP, extend a polarized projection and mate. Thus, the requirement for Cdc42-GTP hydrolysis by GTPase activating proteins is distinct during polarization by intrinsic or extrinsic cues.

cell biology↗

bioRxiv2018

Microtubule plus-tips act as signaling hubs for positioning the cleavage furrow during cytokinesis.

Cell division in animal cells culminates with the formation of a contractile ring that divides the cytosol through formation of a cleavage furrow. Microtubules (MTs) are essential for furrow positioning, but the molecular nature of MT-derived spatial signals is unresolved. In this study essential cytokinesis regulators (the centralspindlin complex, aurora B kinase (ABK), and polo kinase) were visualized in Drosophila melanogaster (Dm) cells and found localize to and track MT plus-ends during cytokinesis. The RhoA GEF Pebble (Dm ECT2) did not robustly tip-track but became enriched at MT plus-tips rapidly following cortical contact resulting in RhoA activation and enrichment of myosin-regulatory light chain. Abrogation of cytokinesis regulator tip-tracking by EB1 depletion or deletion of a novel EB1-interaction motif (hxxPTxh) in the centralspindlin component RacGAP50C resulted in higher incidences of cytokinesis failure. We propose that EB1-dependent, MT plus-tip-based signaling hubs recruit cortical Dm ECT2 upon contact to locally activate RhoA.

cell biology↗

bioRxiv2018

A long-non-coding RNA, LINC00473, confers the human adipose tissue thermogenic phenotype through enhanced cAMP responsiveness.

Specialized adipocytes localized in distinct depots mediate the many physiological functions of adipose tissue. In humans, paucity of thermogenic adipocytes correlates with high metabolic disease risk, raising much interest in the mechanisms by which these cells arise. Here we report molecular signatures associated with adipocyte development in different human depots and identify a long non-coding RNA, LINC00473, as the transcript most closely associated with enrichment of thermogenic adipocytes. LINC00473 expression is low in subjects with obesity or type-2 diabetes and is highly correlated with cAMP signaling and mitochondrial oxidative phosphorylation pathways. LINC00473 is localized in the nucleus and the cytoplasm, and its knockdown impairs induction of UCP1 and mitochondrial respiration. These results reveal that depot-enriched genes that modulate responsiveness to external stimuli, specifically LINC00473, are important determinants of the adipose tissue thermogenic phenotype, and potential targets for metabolic disease therapy.

cell biology↗

bioRxiv2018

mTOR-dependent phosphorylation controls TFEB nuclear export

The transcriptional activation of catabolic processes during starvation is induced by the nuclear translocation and consequent activation of transcription factor EB (TFEB), a master modulator of autophagy and lysosomal biogenesis. However, how TFEB is inactivated upon nutrient re-feeding is currently unknown. Here we show that TFEB subcellular localization is dynamically controlled by its continuous shuttling between the cytosol and the nucleus, with the nuclear export representing a limiting step. TFEB nuclear export is mediated by CRM1 and is modulated by nutrient availability via mTOR-dependent hierarchical multisite phosphorylation of serines S142 and S138, which are localized in proximity of a nuclear export signal (NES). Our data reveal that modulation of TFEB nuclear export via phosphorylation plays a major role in the modulation of TFEB localization and activity.

cell biology↗

bioRxiv2018

ER-to-Golgi trafficking of procollagen in the absence of large carriers.

Secretion and assembly of collagen is fundamental to the function of the extracellular matrix. Defects in the assembly of a collagen matrix lead to pathologies including fibrosis and osteogenesis imperfecta. Owing to the size of fibril-forming procollagen molecules it is assumed that they are transported from the endoplasmic reticulum to the Golgi in specialised large COPII-dependent carriers. Here, analysing endogenous procollagen and a new engineered GFP-tagged form, we show that transport to the Golgi occurs in the absence of large carriers. Large GFP-positive structures are observed occasionally but these are non-dynamic, are not COPII-positive, and label with markers of the ER. We propose a \"short-loop\" model of COPII-dependent ER-to-Golgi traffic that, while consistent with models of ERGIC-dependent expansion of COPII carriers, does not invoke long-range trafficking of large vesicular structures. Our findings provide an important insight into the process of procollagen trafficking and reveal a short-loop pathway from the ER to the Golgi, without the use of large carriers.\n\nSummaryTrafficking of procollagen is essential for normal cell function. Here, imaging of GFP-tagged type I procollagen reveals that it is transported from the endoplasmic reticulum to the Golgi, without the use of large carriers.

cell biology↗

bioRxiv2018

EFFECTORS OF THE SPINDLE ASSEMBLY CHECKPOINT BUT NOT THE MITOTIC EXIT NETWORK ARE CONFINED WITHIN THE NUCLEUS OF SACCHAROMYCES CEREVISIAE

The Spindle Assembly Checkpoint (SAC) prevents erroneous chromosome segregation by delaying mitotic progression when chromosomes are incorrectly attached to the mitotic spindle. This delay is mediated by Mitotic Checkpoint Complexes (MCCs), which assemble at unattached kinetochores and repress the activity of the Anaphase Promoting Complex/Cyclosome (APC/C). The cellular localizations of MCCs are likely critical for proper SAC function, yet remain poorly defined. We recently demonstrated that in mammalian cells, in which the nuclear envelope disassembles during mitosis, MCCs diffuse throughout the spindle region and cytoplasm. Here, we employed binucleate yeast zygotes to examine the localization dynamics of SAC effectors required for MCC assembly and function in budding yeast, in which the nuclear envelope remains intact throughout mitosis. Our findings indicate that in yeast MCCs are confined to the nuclear compartment and excluded from the cytoplasm during mitosis. In contrast, we find that effectors of the Mitotic Exit Network (MEN) - a pathway that initiates disassembly of the anaphase spindle only when it is properly oriented - are in fact freely exchanged between multiple nuclei within a shared cytoplasm. Our study provides insight into how cell cycle checkpoints have evolved to function in diverse cellular contexts.

cell biology↗

bioRxiv2018

Candida albicans hyphal expansion causes phagosomal membrane damage and luminal alkalinization

Macrophages rely on phagosomal acidity to destroy engulfed microorganisms. To survive this hostile response, opportunistic fungi such as Candida albicans developed strategies to evade the acidic environment. C. albicans is polymorphic, able to convert from yeast to hyphae, and this transition is required to subvert the microbicidal activity of the phagosome. However, the phagosomal lumen, which is acidic and nutrient-deprived, inhibits yeast-to-hypha transition. To account for this apparent paradox, it was recently proposed that C. albicans produces ammonia that alkalinizes the phagosome, thus facilitating yeast-to-hypha transformation. We re-examined the mechanism underlying phagosomal alkalinization by applying dual-wavelength ratiometric pH measurements. The phagosomal membrane was found to be highly permeable to ammonia, which is therefore unlikely to account for the pH elevation. Instead, we find that yeast-to-hypha transition begins within acidic phagosomes, and that alkalinization is a consequence of proton leakage induced by excessive membrane distension caused by the expanding hypha.\n\nIMPORTANCEC. albicans is the most common nosocomial fungal infection, and over three million people acquire life-threatening invasive fungal infections every year. Even if antifungal drugs exist, almost half of these patients will die. Despite this, fungi remain underestimated as pathogens. Our study uses quantitative biophysical approaches to demonstrate that the yeast-to-hypha transition occurs within the nutrient deprived, acidic phagosome and that alkalinization is a consequence, as opposed to the cause of hyphal growth.

cell biology↗

bioRxiv2018

Regulation of EGFR endocytosis by CBL during mitosis

Endocytosis plays important roles in regulating EGFR signaling. We previously found that EGFR endocytosis during mitosis is mediated differently than during interphase. While the regulation of EGFR endocytosis in interphase is well understood, little is known regarding the regulation of EGFR endocytosis during mitosis. Here, we studied the mechanisms regulating mitotic EGFR endocytosis. We found that contrary to interphase cells, mitotic EGFR endocytosis is more reliant on the activation of the E3 ligase CBL. At high EGF doses, inhibition of inhibited EGFR endocytosis of mitotic cells, but not of interphase cells. Moreover, the endocytosis of mutant EGFR Y1045F-YFP was strongly delayed. The endocytosis of truncated EGFR {Delta}1044-YFP that does not bind to CBL was completely inhibited. EGF induced stronger ubiquitination of mitotic EGFR than interphase EGFR and mitotic EGFR is trafficked to lysosome for degradation. Furthermore, during mitosis low doses of EGF also stimulate EGFR endocytosis by NCE. Contrary to interphase, CBL and the CBL-binding regions of EGFR were required for mitotic EGFR endocytosis at low doses. This may be due to the mitotic ubiquitination of the EGFR even at low EGF doses. In conclusion, mitotic EGFR endocytosis solely proceed through CBL-mediated NCE.

cell biology↗

bioRxiv2018

ER shaping proteins regulate mitochondrial fission, outer membrane permeabilization and apoptosis

The mitochondrial fission machinery, comprising a dynamin-related GTPase, DRP-1, is crucial for the regulation of mitochondrial membrane dynamics. Recent reports suggest that the tubular architecture of the endoplasmic reticulum (ER) marks the constriction sites on the mitochondria to facilitate DRP-1-mediated mitochondrial fission. However, the role of several ER shaping proteins that maintain the elaborate network of tubes and sheets in mitochondrial constriction and fission is not yet known. In this report, we demonstrate that modulation of the expression levels of key ER shaping proteins, namely Reticulon1 (RTN-1), Reticulon 4 (RTN-4), Lunapark-1 (LNP-1) and CLIMP-63, markedly decreased the extent of mitochondrial fission mediated by BH3 mimetics, despite no detectable changes in DRP-1 recruitment to the mitochondria. Furthermore, modulation of ER shaping proteins significantly decreased other hallmarks of apoptosis, such as mitochondrial outer membrane permeabilization, caspase activation and phosphatidylserine externalization, and functioned independently of mitochondrial cristae remodeling, thus demonstrating a requirement of ER shaping proteins and ER structural integrity for the efficient execution of the instrinsic apoptotic pathway.

cell biology↗

bioRxiv2018

Cell type-dependent differential activation of ERK by oncogenic KRAS or BRAF in the mouse intestinal epithelium

Mutations activating the KRAS GTPase or the BRAF kinase are frequent in colorectal cancer and are thought to constitutively activate the terminal mitogen-activated protein kinase, ERK. Using mass cytometry, we found graded phosphorylation of ERK anti-correlated with cell differentiation in patient-derived colorectal cancer organoids, independent of KRAS mutational status. Reporter, single cell transcriptome and mass cytometry analyses showed that transgenic KRASG12V activated ERK in a cell type-specific pattern in mouse intestinal organoids. In contrast, transgenic BRAFV600E triggered high ERK activity and downstream gene expression in all intestinal cell types, followed by epithelial disorganisation. Quantitative network modelling from perturbation data revealed that activation of ERK is shaped by cell type-specific MEK to ERK feed forward and negative feedback signalling. We identified dual-specificity phosphatases as candidate modulators of ERK activity between intestinal cell types. Furthermore, we found that oncogenic KRAS, together with {beta}-Catenin, favoured expansion of crypt cells with high ERK activity. Our experiments highlight key differences between ERK activity elicited by the BRAF or KRAS oncogenes in colorectal cancer and find unexpected heterogeneity in a signalling pathway with fundamental relevance for cancer therapy.

cell biology↗