Cell & Developmental Biology

Permanent URI for this collection

https://ir.vanderbilt.edu/handle/1803/9926

Browse

Now showing 1 - 17 of 17

Retrograde trafficking of Argonaute 2 acts as a rate-limiting step for de novo miRNP formation on endoplasmic reticulum-attached polysomes in mammalian cells
(Life Science Alliance, 2020-02) Bose, Mainak; Chatterjee, Susanta; Chakrabarty, Yogaditya; Barman, Bahnisikha; Bhattacharyya, Suvendra N.
microRNAs are short regulatory RNAs in metazoan cells. Regulation of miRNA activity and abundance is evident in human cells where availability of target messages can influence miRNA biogenesis by augmenting the Dicer1-dependent processing of precursors to mature microRNAs. Requirement of subcellular compartmentalization of Ago2, the key component of miRNA repression machineries, for the controlled biogenesis of miRNPs is reported here. The process predominantly happens on the polysomes attached with the endoplasmic reticulum for which the subcellular Ago2 trafficking is found to be essential. Mitochondrial tethering of endoplasmic reticulum and its interaction with endosomes controls Ago2 availability. In cells with depolarized mitochondria, miRNA biogenesis gets impaired, which results in lowering of de novo-formed mature miRNA levels and accumulation of miRNA-free Ago2 on endosomes that fails to interact with Dicer1 and to traffic back to endoplasmic reticulum for de novo miRNA loading. Thus, mitochondria by sensing the cellular context regulates Ago2 trafficking at the subcellular level, which acts as a rate-limiting step in miRNA biogenesis process in mammalian cells.
PPARγ-Independent Side Effects of Thiazolidinediones on Mitochondrial Redox State in Rat Isolated Hearts
(Cells, 2020-01) Riess, Matthias L.; Elorbany, Reem; Weihrauch, Dorothee; Stowe, David F.; Camara, Amadou K. S.
The effect of anti-diabetic thiazolidinediones (TZDs) on contributing to heart failure and cardiac ischemia/reperfusion (IR) injury is controversial. In this study we investigated the effect of select TZDs on myocardial and mitochondrial function in Brown Norway rat isolated hearts. In a first set of experiments, the TZD rosiglitazone was given acutely before global myocardial IR, and pre- and post-IR function and infarct size were assessed. In a second set of experiments, different concentrations of rosiglitazone and pioglitazone were administered in the presence or absence of the specific PPAR gamma antagonist GW9662, and their effects on the mitochondrial redox state were measured by online NADH and FAD autofluorescence. The administration of rosiglitazone did not significantly affect myocardial function except for transiently increasing coronary flow, but it increased IR injury compared to the control hearts. Both TZDs resulted in dose-dependent, reversible increases in mitochondrial oxidation which was not attenuated by GW9662. Taken together, these data suggest that TZDs cause excessive mitochondrial uncoupling by a PPAR gamma-independent mechanism. Acute rosiglitazone administration before IR was associated with enhanced cardiac injury. If translated clinically, susceptible patients on PPAR gamma agonists may experience enhanced myocardial IR injury by mitochondrial dysfunction.
Delineating the Molecular Basis of the Calmodulin–bMunc13-2 Interaction by Cross-Linking/Mass Spectrometry—Evidence for a Novel CaM Binding Motif in bMunc13-2
(Cells, 2020-01) Piotrowski, Christine; Moretti, Rocco; Ihling, Christian H.; Haedicke, Andre; Liepold, Thomas; Lipstein, Noa; Meiler, Jens; Jahn, Olaf; Sinz, Andrea
Exploring the interactions between the Ca2+ binding protein calmodulin (CaM) and its target proteins remains a challenging task. Members of the Munc13 protein family play an essential role in short-term synaptic plasticity, modulated via the interaction with CaM at the presynaptic compartment. In this study, we focus on the bMunc13-2 isoform expressed in the brain, as strong changes in synaptic transmission were observed upon its mutagenesis or deletion. The CaM-bMunc13-2 interaction was previously characterized at the molecular level using short bMunc13-2-derived peptides only, revealing a classical 1-5-10 CaM binding motif. Using larger protein constructs, we have now identified for the first time a novel and unique CaM binding site in bMunc13-2 that contains an N-terminal extension of a classical 1-5-10 CaM binding motif. We characterize this motif using a range of biochemical and biophysical methods and highlight its importance for the CaM-bMunc13-2 interaction.
PP2A(C) Phospho-Tyr(307) Antibodies Are Not Specific for this Modification but Are Sensitive to Other PP2A(C) Modifications Including Leu(309) Methylation
(Cell Reports, 2020-03-03) Frohner, Ingrid E.; Mudrak, Ingrid; Schuechner, Stefan; Anrather, Dorothea; Hartl, Markus; Sontag, Jean-Marie; Sontag, Estelle; Wadzinski, Brian E.; Preglej, Teresa; Ellmeier, Wilfried; Ogris, Egon
Protein phosphatase 2A (PP2A) is an important regulator of signal transduction pathways and a tumor suppressor. Phosphorylation of the PP2A catalytic subunit (PP2A(C)) at tyrosine 307 has been claimed to inactivate PP2A and was examined in more than 180 studies using commercial antibodies, but this modification was never identified using mass spectrometry. Here we show that the most cited pTyr(307) monoclonal antibodies, E155 and F-8, are not specific for phosphorylated Tyr(307) but instead are hampered by PP2A(C) methylation at leucine 309 or phosphorylation at threonine 304. Other pTyr(307)( )antibodies are sensitive to PP2A(C) methylation as well, and some cross-react with pTyr residues in general, including phosphorylated hemagglutinin tags. We identify pTyr(307) using targeted mass spectrometry after transient overexpression of PP2A(C) and Src kinase. Yet under such conditions, none of the tested antibodies show exclusive pTyr(307) specificity. Thus, data generated using these antibodies need to be revisited, and the mechanism of PP2A inactivation needs to be redefined.
A Review of Precision Oncology Knowledgebases for Determining the Clinical Actionability of Genetic Variants
(Frontiers in Cell and Developmental Biology, 2020-02-11) Li, Xuanyi; Warner, Jeremy L.
The increased availability of tumor genetic testing and targeted cancer therapies contributes to the advancement of precision medicine in the field of oncology. Precision oncology knowledgebases provide a way of organizing clinically relevant genetic information in a way that is easily accessible for both oncologists and patients, facilitating the genetic-based clinical decision making. Many organizations and companies have built precision oncology knowledgebases, intended for multiple users. In general, these knowledgebases offer information on cancer-related genetic variants as well as their associated diagnostic, prognostic, and therapeutic implications, but they often differ in their information curations, designs, and user experiences. It is advisable that oncologists use multiple knowledgebases during their practice to have them complement each other. In the future, convergence toward common standards and formats is needed to ensure that the comprehensive knowledge across all sources can be unified to bring the oncology community closer to the achievement of the goal of precision oncology.
Microtubule minus-end stability is dictated by the tubulin off-rate
(Journal of Cell Biology, 2019-09) Strothman, Claire; Farmer, Veronica; Arpag, Goker; Rodgers, Nicole; Podolski, Marija; Norris, Stephen; Ohi, Ryoma; Zanic, Marija
Dynamic organization of microtubule minus ends is vital for the formation and maintenance of acentrosomal microtubule arrays. In vitro, both microtubule ends switch between phases of assembly and disassembly, a behavior called dynamic instability. Although minus ends grow slower, their lifetimes are similar to those of plus ends. The mechanisms underlying these distinct dynamics remain unknown. Here, we use an in vitro reconstitution approach to investigate minus-end dynamics. We find that minus-end lifetimes are not defined by the mean size of the protective GTP-tubulin cap. Rather, we conclude that the distinct tubulin off-rate is the primary determinant of the difference between plus- and minus-end dynamics. Further, our results show that the minus-end-directed kinesin-14 HSET/KIFC1 suppresses tubulin off-rate to specifically suppress minus-end catastrophe. HSET maintains its protective minus-end activity even when challenged by a known microtubule depolymerase, kinesin-13 MCAK. Our results provide novel insight into the mechanisms of minus-end dynamics, essential for our understanding of microtubule minus-end regulation in cells.
Correlated evolution between repertoire size and song plasticity predicts that sexual selection on song promotes open-ended learning
(eLife, 2019-09-03) Robinson, Cristina M.; Snyder, Kate T.; Creanza, Nicole
Some oscine songbird species modify their songs throughout their lives ('adult song plasticity' or 'open-ended learning'), while others crystallize their songs around sexual maturity. It remains unknown whether the strength of sexual selection on song characteristics, such as repertoire size, affects adult song plasticity, or whether adult song plasticity affects song evolution. Here, we compiled data about song plasticity, song characteristics, and mating system and then examined evolutionary interactions between these traits. Across 67 species, we found that lineages with adult song plasticity show directional evolution toward increased syllable and song repertoires, while several other song characteristics evolved faster, but in a non-directional manner. Song plasticity appears to drive bi-directional transitions between monogamous and polygynous social mating systems. Notably, our analysis of correlated evolution suggests that extreme syllable and song repertoire sizes drive the evolution of adult song plasticity or stability, providing novel evidence that sexual selection may indirectly influence open-versus closed-ended learning.
Myeloid Cell-Derived HB-EGF Drives Tissue Recovery After Pancreatitis
(Cellular and Molecular Gastroenterology and Hepatology, 2019) Wen, Hui-Ju; Gao, Shan; Wang, Yin; Ray, Michael; Magnuson, Mark A.; Wright, Christopher V. E.; Di Magliano, Marina Pasca; Frankel, Timothy L.; Crawford, Howard C.
BACKGROUND & AIMS: Pancreatitis is a major cause of morbidity and mortality and is a risk factor for pancreatic tumorigenesis. Upon tissue damage, an inflammatory response, made up largely of macrophages, provides multiple growth factors that promote repair. Here, we examine the molecular pathways initiated by macrophages to promote pancreas recovery from pancreatitis. METHODS: To induce organ damage, mice were subjected to cerulein-induced experimental pancreatitis and analyzed at various times of recovery. CD11b-DTR mice were used to deplete myeloid cells. Hbeg(f/f);LysM-Cre mice were used to ablate myeloid cell-derived heparin-binding epidermal growth factor (EGF)-like growth factor (HB-EGF). To ablate EGFR specifically during recovery, pancreatitis was induced in Egfr(f/f);Ptf1a(Flp0/+);FSF-Rosa26(CAG-CreERT2 )mice followed by tamoxifen treatment. RESULTS: Macrophages infiltrating the pancreas in experimental pancreatitis make high levels of HB-EGF. Both depletion of myeloid cells and ablation of myeloid cell HB-EGF delayed recovery from experimental pancreatitis, resulting from a decrease in cell proliferation and an increase in apoptosis. Mechanistically, ablation of myeloid cell HB-EGF impaired epithelial cell DNA repair, ultimately leading to cell death. Soluble HB-EGF induced EGFR nuclear translocation and methylation of histone H4, facilitating resolution of DNA damage in pancreatic acinar cells in vitro. Consistent with its role as the primary receptor of HB-EGF, in vivo ablation of EGFR from pancreatic epithelium during recovery from pancreatitis resulted in accumulation of DNA damage. CONCLUSIONS: By using novel conditional knockout mouse models, we determined that HB-EGF derived exclusively from myeloid cells induces epithelial cell proliferation and EGFR-dependent DNA repair, facilitating pancreas healing after injury.
Enteropathogenic Escherichia coli remodels host endosomes to promote endocytic turnover and breakdown of surface polarity
(PLOS Pathogens, 2019-06) Kassa, Ephrem G.; Zlotkin-Rivkin, Efrat; Friedman, Gil; Ramachandran, Rachana P.; Melamed-Book, Naomi; Weiss, Aryeh M.; Belenky, Michael; Reichmann, Dana; Breuer, William; Pal, Ritesh Ranjan; Rosenshine, Ilan; Lapierre, Lynne A.; Goldenring, James R.; Aroeti, Benjamin
Enteropathogenic E. coli (EPEC) is an extracellular diarrheagenic human pathogen which infects the apical plasma membrane of the small intestinal enterocytes. EPEC utilizes a type III secretion system to translocate bacterial effector proteins into its epithelial hosts. This activity, which subverts numerous signaling and membrane trafficking pathways in the infected cells, is thought to contribute to pathogen virulence. The molecular and cellular mechanisms underlying these events are not well understood. We investigated the mode by which EPEC effectors hijack endosomes to modulate endocytosis, recycling and transcytosis in epithelial host cells. To this end, we developed a flow cytometry-based assay and imaging techniques to track endosomal dynamics and membrane cargo trafficking in the infected cells. We show that type-III secreted components prompt the recruitment of clathrin (clathrin and AP2), early (Rab5a and EEA1) and recycling (Rab4a, Rab11a, Rab11b, FIP2, Myo5b) endocytic machineries to peripheral plasma membrane infection sites. Protein cargoes, e.g. transferrin receptors, beta 1 integrins and aquaporins, which exploit the endocytic pathways mediated by these machineries, were also found to be recruited to these sites. Moreover, the endosomes and cargo recruitment to infection sites correlated with an increase in cargo endocytic turnover (i.e. endocytosis and recycling) and transcytosis to the infected plasma membrane. The hijacking of endosomes and associated endocytic activities depended on the translocated EspF and Map effectors in non-polarized epithelial cells, and mostly on EspF in polarized epithelial cells. These data suggest a model whereby EPEC effectors hijack endosomal recycling machineries to mislocalize and concentrate host plasma membrane proteins in endosomes and in the apically infected plasma membrane. We hypothesize that these activities contribute to bacterial colonization and virulence. Author summary Enteropathogenic Escherichia coli (EPEC) are pathogenic bacteria that cause infantile diarrhea. Upon ingestion, EPEC reaches the small intestine, where an injection device termed the type III secretion system is utilized to inject a set of effector proteins from the bacteria into the host cell. These proteins manipulate the localization and functions of host proteins, lipids and organelles and contribute to the emergence of the EPEC disease. The molecular mechanisms underlying the functions of the EPEC effector proteins are not completely understood. Here we show that early upon infection, two such effector proteins, EspF and Map, hijack host endosomes at bacterial adherence sites to facilitate endocytosis and recycling of plasma membrane proteins at these sites. The consequence of this event is the enrichment and mislocalization of host plasma membrane proteins at infection sites. One such protein is the transferrin receptor, which is a carrier for transferrin, whose function is to mediate cellular uptake of iron. Iron is a critical nutrient for bacterial growth and survival. We postulate that the unique manipulation of transferrin receptor endocytic membrane trafficking by EPEC plays an important role in its survival on the luminal surface of the intestinal epithelium.
USP9X Deubiquitylates DVL2 to Regulate WNT Pathway Specification
(Cell Reports, 2019-07-23) Nielsen, Casey P.; Jemigan, Kristin K.; Diggins, Nicole L.; Webb, Donna J.; MacGurn, Jason A.
The WNT signaling network is comprised of multiple receptors that relay various input signals via distinct transduction pathways to execute multiple complex and context-specific output processes. Integrity of the WNT signaling network relies on proper specification between canonical and noncanonical pathways, which presents a regulatory challenge given that several signal transducing elements are shared between pathways. Here, we report that USP9X, a deubiquitylase, and WWP1, an E3 ubiquitin ligase, regulate a ubiquitin rheostat on DVL2, a WNT signaling protein. Our findings indicate that USP9X-mediated deubiquitylation of DVL2 is required for canonical WNT activation, while increased DVL2 ubiquitylation is associated with localization to actin-rich projections and activation of the planar cell polarity (PCP) pathway. We propose that a WWP1-USP9X axis regulates a ubiquitin rheostat on DVL2 that specifies its participation in either canonical WNT or WNT-PCP pathways. These findings have important implications for therapeutic targeting of USP9X in human cancer.
CpG-related SNPs in the MS4A region have a dose-dependent effect on risk of late-onset Alzheimer disease
(Aging Cell, 2019-08) Thornton-Wells, Tricia A.
CpG-related single nucleotide polymorphisms (CGS) have the potential to perturb DNA methylation; however, their effects on Alzheimer disease (AD) risk have not been evaluated systematically. We conducted a genome-wide association study using a sliding-window approach to measure the combined effects of CGSes on AD risk in a discovery sample of 24 European ancestry cohorts (12,181 cases, 12,601 controls) from the Alzheimer's Disease Genetics Consortium (ADGC) and replication sample of seven European ancestry cohorts (7,554 cases, 27,382 controls) from the International Genomics of Alzheimer's Project (IGAP). The potential functional relevance of significant associations was evaluated by analysis of methylation and expression levels in brain tissue of the Religious Orders Study and the Rush Memory and Aging Project (ROSMAP), and in whole blood of Framingham Heart Study participants (FHS). Genome-wide significant (p < 5 x 10(-8)) associations were identified with 171 1.0 kb-length windows spanning 932 kb in the APOE region (top p < 2.2 x 10(-308)), five windows at BIN1 (top p = 1.3 x 10(-13)), two windows at MS4A6A (top p = 2.7 x 10(-10)), two windows near MS4A4A (top p = 6.4 x 10(-10)), and one window at PICALM (p = 6.3 x 10(-9)). The total number of CGS-derived CpG dinucleotides in the window near MS4A4A was associated with AD risk (p = 2.67 x 10(-10)), brain DNA methylation (p = 2.15 x 10(-10)), and gene expression in brain (p = 0.03) and blood (p = 2.53 x 10(-4)). Pathway analysis of the genes responsive to changes in the methylation quantitative trait locus signal at MS4A4A (cg14750746) showed an enrichment of methyltransferase functions. We confirm the importance of CGS in AD and the potential for creating a functional CpG dosage-derived genetic score to predict AD risk.
Point mutations in the PDX1 transactivation domain impair human beta-cell development and function
(Molecular Metabolism, 2019-06) Wright, Christopher V. E.
Objective: Hundreds of missense mutations in the coding region of PDX1 exist; however, if these mutations predispose to diabetes mellitus is unknown. Methods: In this study, we screened a large cohort of subjects with increased risk for diabetes and identified two subjects with impaired glucose tolerance carrying common, heterozygous, missense mutations in the PDX1 coding region leading to single amino acid exchanges (P33T, C18R) in its transactivation domain. We generated iPSCs from patients with heterozygous PDX1(P33T/)(+), PDX1(C18R/+) mutations and engineered isogenic cell lines carrying homozygous PDX1(P33T/P33T), PDX1(C18R/C18R) mutations and a heterozygous PDX1 loss-of-function mutation (PDX1(+/-)). Results: Using an in vitro beta-cell differentiation protocol, we demonstrated that both, heterozygous PDXP33T/+, PDX1(C18R/+)and homozygous PDX1(P33T/P33T), PDX1(C18R/C18R) mutations impair beta-cell differentiation and function. Furthermore, PDX1(+/-) and PDX1(P33T/P33T )mutations reduced differentiation efficiency of pancreatic progenitors (PPs), due to downregulation of PDX1 -bound genes, including transcription factors MNX1 and PDX1 as well as insulin resistance gene CES1. Additionally, both PDX1(P33T/+ )and PDX1(P33T/P33T) mutations in PPs reduced the expression of PDX1-bound genes including the long-noncoding RNA, MEG3 and the imprinted gene NNAT, both involved in insulin synthesis and secretion. Conclusions: Our results reveal mechanistic details of how common coding mutations in PDX1 impair human pancreatic endocrine lineage formation and beta-cell function and contribute to the predisposition for diabetes. (C) 2019 The Authors. Published by Elsevier GmbH.
Diaspora linkages benefit both sides: a single partnership experience
(Global Health Action, 2019-01) Kramer, Beverley; Zent, Roy
The emigration of physicians and scientists from resource-constrained countries decreases the country's ability to undertake research. Re-establishing research environments and increasing capacity reduced by these losses are important, particularly in the health sciences. One mechanism for re-establishing strong health sciences research is the introduction of an Alumni Diaspora Fellowship Programme. We define the beneficial effects of a successful single partnership in an Alumni Diaspora Programme. This Host/Alumnus collaboration demonstrates that bi-directional advantages have accrued for both the Host Institution situated in a resource-constrained country and the Alumni's Institution, located in a high-income country. In addition to expanding research in the resource-constrained country, collaborations expanded to other faculty beyond the Alumnus in the sending Institution, in multiple fields including those not readily available in the high-income country (HIV, TB, malaria). The environment at the host Institution in the resource-constrained country has been enriched by increased research publications, training of young scholars (over 200 trained in manuscript and grant application writing), and substantial advances in biomedical informatics. There has been considerable knowledge exchange and development between both Institutions, showing that 'brain circulation' and Diaspora Programmes are valuable strategies for expanding research.
Deletion of Tet proteins results in quantitative disparities during ESC differentiation partially attributable to alterations in gene expression
(BMC Developmental Biology, 2019-07-08) Reimer, Michael, Jr.; Pulakanti, Kirthi; Shi, Linzheng; Abel, Alex; Liang, Mingyu; Malarkannan, Subramaniam; Rao, Sridhar
BackgroundThe Tet protein family (Tet1, Tet2, and Tet3) regulate DNA methylation through conversion of 5-methylcytosine to 5-hydroxymethylcytosine which can ultimately result in DNA demethylation and play a critical role during early mammalian development and pluripotency. While multiple groups have generated knockouts combining loss of different Tet proteins in murine embryonic stem cells (ESCs), differences in genetic background and approaches has made it difficult to directly compare results and discern the direct mechanism by which Tet proteins regulate the transcriptome. To address this concern, we utilized genomic editing in an isogenic pluripotent background which permitted a quantitative, flow-cytometry based measurement of pluripotency in combination with genome-wide assessment of gene expression and DNA methylation changes. Our ultimate goal was to generate a resource of large-scale datasets to permit hypothesis-generating experiments.ResultsWe demonstrate a quantitative disparity in the differentiation ability among Tet protein deletions, with Tet2 single knockout exhibiting the most severe defect, while loss of Tet1 alone or combinations of Tet genes showed a quantitatively intermediate phenotype. Using a combination of transcriptomic and epigenomic approaches we demonstrate an increase in DNA hypermethylation and a divergence of transcriptional profiles in pluripotency among Tet deletions, with loss of Tet2 having the most profound effect in undifferentiated ESCs.ConclusionsWe conclude that loss of Tet2 has the most dramatic effect both on the phenotype of ESCs and the transcriptome compared to other genotypes. While loss of Tet proteins increased DNA hypermethylation, especially in gene promoters, these changes in DNA methylation did not correlate with gene expression changes. Thus, while loss of different Tet proteins alters DNA methylation, this change does not appear to be directly responsible for transcriptome changes. Thus, loss of Tet proteins likely regulates the transcriptome epigenetically both through altering 5mC but also through additional mechanisms. Nonetheless, the transcriptome changes in pluripotent Tet2(-/-) ESCs compared to wild-type implies that the disparities in differentiation can be partially attributed to baseline alterations in gene expression.
Regulation of the Pancreatic Exocrine Differentiation Program and Morphogenesis by Onecut 1/Hnf6
(Cellular and Molecular Gastroenterology and Hepatology, 2019) Kropp, Peter A.; Zhu, Xiaodong; Gannon, Maureen
BACKGROUND & AIMS: The Onecut 1 transcription factor (Oc1, a.k.a. HNF6) promotes differentiation of endocrine and duct cells of the pancreas; however, it has no known role in acinar cell differentiation. We sought to better understand the role of Oc1 in exocrine pancreas development and to identify its direct transcriptional targets. METHODS: Pancreata from Oc1(Delta panc) (Oc1(fl/fl); Pdx1-Cre) mouse embryos and neonates were analyzed morphologically. High-throughput RNA-sequencing was performed on control and Oc1-deficient pancreas; chromatin immunoprecipitation sequencing was performed on wild-type embryonic mouse pancreata to identify direct Oc1 transcriptional targets. Immunofluorescence labeling was used to confirm the RNA-sequencing/chromatin immuno precipitation sequencing results and to further investigate the effects of Oc1 loss on acinar cells. RESULTS: Loss of Oc1 from the developing pancreatic epithelium resulted in disrupted duct and acinar cell development. RNA-sequencing revealed decreased expression of acinar cell regulatory factors (Nr5a2, Ptf1a, Gata4, Mist1) and functional genes (Amylase, Cpa1, Prss1, Spink1) at embryonic day (e) 18.5 in Oc1(Delta panc) samples. Approximately 1000 of the altered genes were also identified as direct Oc1 targets by chromatin immunoprecipitation sequencing, including most of the previously noted genes. By immunolabeling, we confirmed that Amylase, Mist1, and GATA4 protein levels are significantly decreased by P2, and Spink1 protein levels were significantly reduced and mislocalized. The pancreatic duct regulatory factors Hnf1 beta and FoxA2 were also identified as direct Oc1 targets. CONCLUSIONS: These findings confirm that Oc1 is an important regulator of both duct and acinar cell development in the embryonic pancreas. Novel transcriptional targets of Oc1 have now been identified and provide clarity into the mechanisms of Oc1 transcriptional regulation in the developing exocrine pancreas. Oc1 can now be included in the gene-regulatory network of acinar cell regulatory genes. Oc1 regulates other acinar cell regulatory factors and acinar cell functional genes directly, and it can also regulate some acinar cell regulatory factors (eg, Mist1) indirectly. Oc1 therefore plays an important role in acinar cell development.
Structural state recognition facilitates tip tracking of EB1 at growing microtubule ends
(eLIFE, 2019-09-03) Reid, Taylor A.; Coombes, Courtney; Mukherjee, Soumya; Goldblum, Rebecca R.; White, Kyle; Parmar, Sneha; McClellan, Mark; Zanic, Marija; Courtemanche, Naomi; Gardner, Melissa K.
The microtubule binding protein EB1 specifically targets the growing ends of microtubules in cells, where EB1 facilitates the interactions of cellular proteins with microtubule plus-ends. Microtubule end targeting of EB1 has been attributed to high-affinity binding of EB1 to GTP-tubulin that is present at growing microtubule ends. However, our 3D single-molecule diffusion simulations predicted a similar to 6000% increase in EB1 arrivals to open, tapered microtubule tip structures relative to closed lattice conformations. Using quantitative fluorescence, single-molecule, and electron microscopy experiments, we found that the binding of EB1 onto opened, structurally disrupted microtubules was dramatically increased relative to closed, intact microtubules, regardless of hydrolysis state. Correspondingly, in cells, the blunting of growing microtubule plus-ends by Vinblastine was correlated with reduced EB1 targeting. Together, our results suggest that microtubule structural recognition, based on a fundamental diffusion-limited binding model, facilitates the tip tracking of EB1 at growing microtubule ends.
Targeting WDR5: A WINning Anti-Cancer Strategy?
(EPIGENETICS INSIGHTS, 2019-07-18) Aho, Erin R.; Weissmiller, April M.; Fesik, Stephen W.; Tansey, William P.
WDR5 is a component of multiple epigenetic regulatory complexes, including the mixed lineage leukemia (MLL)/SET complexes that deposit histone H3 lysine 4 methylation. Inhibitors of an arginine-binding cavity in WDR5, known as the WDR5-interaction (WIN) site, have been proposed to selectively kill MLL-rearranged malignancies via an epigenetic mechanism. We discovered potent WIN site inhibitors and found that they kill MLL cancer cells not through changes in histone methylation. but by displacing WDR5 from chromatin at protein synthesis genes, choking the translational capacity of these cells, and inducing death via a nucleolar stress response. The mechanism of action of WIN site inhibitors reveals new aspects of WDR5 function and forecasts broad therapeutic utility as anti-cancer agents.

Browse

Recent Submissions