Elizabeth Anne Allen

All Publications

• Therapeutic depletion of CCR8(+) tumor-infiltrating regulatory T cells elicits antitumor immunity and synergizes with anti-PD-1 therapy JOURNAL FOR IMMUNOTHERAPY OF CANCER Van Damme, H., Dombrecht, B., Kiss, M., Roose, H., Allen, E., Van Overmeire, E., Kancheva, D., Martens, L., Murgaski, A., Bardet, P., Blancke, G., Jans, M., Bolli, E., Martins, M., Elkrim, Y., Dooley, J., Boon, L., Schwarze, J., Tacke, F., Movahedi, K., Vandamme, N., Neyns, B., Ocak, S., Scheyltjens, I., Vereecke, L., Nana, F., Merchiers, P., Laoui, D., Van Ginderachter, J. 2021; 9 (2)

  Abstract

  Modulation and depletion strategies of regulatory T cells (Tregs) constitute valid approaches in antitumor immunotherapy but suffer from severe adverse effects due to their lack of selectivity for the tumor-infiltrating (ti-)Treg population, indicating the need for a ti-Treg specific biomarker.We employed single-cell RNA-sequencing in a mouse model of non-small cell lung carcinoma (NSCLC) to obtain a comprehensive overview of the tumor-infiltrating T-cell compartment, with a focus on ti-Treg subpopulations. These findings were validated by flow cytometric analysis of both mouse (LLC-OVA, MC38 and B16-OVA) and human (NSCLC and melanoma) tumor samples. We generated two CCR8-specific nanobodies (Nbs) that recognize distinct epitopes on the CCR8 extracellular domain. These Nbs were formulated as tetravalent Nb-Fc fusion proteins for optimal CCR8 binding and blocking, containing either an antibody-dependent cell-mediated cytotoxicity (ADCC)-deficient or an ADCC-prone Fc region. The therapeutic use of these Nb-Fc fusion proteins was evaluated, either as monotherapy or as combination therapy with anti-programmed cell death protein-1 (anti-PD-1), in both the LLC-OVA and MC38 mouse models.We were able to discern two ti-Treg populations, one of which is characterized by the unique expression of Ccr8 in conjunction with Treg activation markers. Ccr8 is also expressed by dysfunctional CD4+ and CD8+ T cells, but the CCR8 protein was only prominent on the highly activated and strongly T-cell suppressive ti-Treg subpopulation of mouse and human tumors, with no major CCR8-positivity found on peripheral Tregs. CCR8 expression resulted from TCR-mediated Treg triggering in an NF-κB-dependent fashion, but was not essential for the recruitment, activation nor suppressive capacity of these cells. While treatment of tumor-bearing mice with a blocking ADCC-deficient Nb-Fc did not influence tumor growth, ADCC-prone Nb-Fc elicited antitumor immunity and reduced tumor growth in synergy with anti-PD-1 therapy. Importantly, ADCC-prone Nb-Fc specifically depleted ti-Tregs in a natural killer (NK) cell-dependent fashion without affecting peripheral Tregs.Collectively, our findings highlight the efficacy and safety of targeting CCR8 for the depletion of tumor-promoting ti-Tregs in combination with anti-PD-1 therapy.

  View details for DOI 10.1136/jitc-2020-001749

  View details for Web of Science ID 000620639400001

  View details for PubMedID 33589525

  View details for PubMedCentralID PMC7887378

• Consensus guidelines for the use and interpretation of angiogenesis assays ANGIOGENESIS Nowak-Sliwinska, P., Alitalo, K., Allen, E., Anisimov, A., Aplin, A. C., Auerbach, R., Augustin, H. G., Bates, D. O., van Beijnum, J. R., Bender, R. F., Bergers, G., Bikfalvi, A., Bischoff, J., Boeck, B. C., Brooks, P. C., Bussolino, F., Cakir, B., Carmeliet, P., Castranova, D., Cimpean, A. M., Cleaver, O., Coukos, G., Davis, G. E., De Palma, M., Dimberg, A., Dings, R. M., Djonov, V., Dudley, A. C., Dufton, N. P., Fendt, S., Ferrara, N., Fruttiger, M., Fukumura, D., Ghesquiere, B., Gong, Y., Griffin, R. J., Harris, A. L., Hughes, C. W., Hultgren, N. W., Iruela-Arispe, M., Irving, M., Jain, R. K., Kalluri, R., Kalucka, J., Kerbel, R. S., Kitajewski, J., Klaassen, I., Kleinmann, H. K., Koolwijk, P., Kuczynski, E., Kwak, B. R., Marien, K., Melero-Martin, J. M., Munn, L. L., Nicosia, R. F., Noel, A., Nurro, J., Olsson, A., Petrova, T. V., Pietras, K., Pili, R., Pollard, J. W., Post, M. J., Quax, P. A., Rabinovich, G. A., Raica, M., Randi, A. M., Ribatti, D., Ruegg, C., Schlingemann, R. O., Schulte-Merker, S., Smith, L. H., Song, J. W., Stacker, S. A., Stalin, J., Stratman, A. N., Van de Velde, M., van Hinsbergh, V. M., Vermeulen, P. B., Waltenberger, J., Weinstein, B. M., Xin, H., Yetkin-Arik, B., Yla-Herttuala, S., Yoder, M. C., Griffioen, A. W. 2018; 21 (3): 425-532

  Abstract

  The formation of new blood vessels, or angiogenesis, is a complex process that plays important roles in growth and development, tissue and organ regeneration, as well as numerous pathological conditions. Angiogenesis undergoes multiple discrete steps that can be individually evaluated and quantified by a large number of bioassays. These independent assessments hold advantages but also have limitations. This article describes in vivo, ex vivo, and in vitro bioassays that are available for the evaluation of angiogenesis and highlights critical aspects that are relevant for their execution and proper interpretation. As such, this collaborative work is the first edition of consensus guidelines on angiogenesis bioassays to serve for current and future reference.

  View details for DOI 10.1007/s10456-018-9613-x

  View details for Web of Science ID 000438644400001

  View details for PubMedID 29766399

  View details for PubMedCentralID PMC6237663

• Therapeutic induction of high endothelial venules (HEVs) to enhance T-cell infiltration in tumors ONCOTARGET Allen, E., Missiaen, R., Bergers, G. 2017; 8 (59): 99207-99208

  View details for DOI 10.18632/oncotarget.22276

  View details for Web of Science ID 000419561600003

  View details for PubMedID 29245888

  View details for PubMedCentralID PMC5725079

• Combined antiangiogenic and anti-PD-L1 therapy stimulates tumor immunity through HEV formation SCIENCE TRANSLATIONAL MEDICINE Allen, E., Jabouille, A., Rivera, L. B., Lodewijckx, I., Missiaen, R., Steri, V., Feyen, K., Tawney, J., Hanahan, D., Michael, I. P., Bergers, G. 2017; 9 (385)

  Abstract

  Inhibitors of VEGF (vascular endothelial growth factor)/VEGFR2 (vascular endothelial growth factor receptor 2) are commonly used in the clinic, but their beneficial effects are only observed in a subset of patients and limited by induction of diverse relapse mechanisms. We describe the up-regulation of an adaptive immunosuppressive pathway during antiangiogenic therapy, by which PD-L1 (programmed cell death ligand 1), the ligand of the negative immune checkpoint regulator PD-1 (programmed cell death protein 1), is enhanced by interferon-γ-expressing T cells in distinct intratumoral cell types in refractory pancreatic, breast, and brain tumor mouse models. Successful treatment with a combination of anti-VEGFR2 and anti-PD-L1 antibodies induced high endothelial venules (HEVs) in PyMT (polyoma middle T oncoprotein) breast cancer and RT2-PNET (Rip1-Tag2 pancreatic neuroendocrine tumors), but not in glioblastoma (GBM). These HEVs promoted lymphocyte infiltration and activity through activation of lymphotoxin β receptor (LTβR) signaling. Further activation of LTβR signaling in tumor vessels using an agonistic antibody enhanced HEV formation, immunity, and subsequent apoptosis and necrosis in pancreatic and mammary tumors. Finally, LTβR agonists induced HEVs in recalcitrant GBM, enhanced cytotoxic T cell (CTL) activity, and thereby sensitized tumors to antiangiogenic/anti-PD-L1 therapy. Together, our preclinical studies provide evidence that anti-PD-L1 therapy can sensitize tumors to antiangiogenic therapy and prolong its efficacy, and conversely, antiangiogenic therapy can improve anti-PD-L1 treatment specifically when it generates intratumoral HEVs that facilitate enhanced CTL infiltration, activity, and tumor cell destruction.

  View details for DOI 10.1126/scitranslmed.aak9679

  View details for Web of Science ID 000399009200006

  View details for PubMedID 28404866

  View details for PubMedCentralID PMC5554432

• Metabolic Symbiosis Enables Adaptive Resistance to Anti-angiogenic Therapy that Is Dependent on mTOR Signaling CELL REPORTS Allen, E., Mieville, P., Warren, C. M., Saghafinia, S., Li, L., Peng, M., Hanahan, D. 2016; 15 (6): 1144-1160

  Abstract

  Therapeutic targeting of tumor angiogenesis with VEGF inhibitors results in demonstrable, but transitory efficacy in certain human tumors and mouse models of cancer, limited by unconventional forms of adaptive/evasive resistance. In one such mouse model, potent angiogenesis inhibitors elicit compartmental reorganization of cancer cells around remaining blood vessels. The glucose and lactate transporters GLUT1 and MCT4 are induced in distal hypoxic cells in a HIF1α-dependent fashion, indicative of glycolysis. Tumor cells proximal to blood vessels instead express the lactate transporter MCT1, and p-S6, the latter reflecting mTOR signaling. Normoxic cancer cells import and metabolize lactate, resulting in upregulation of mTOR signaling via glutamine metabolism enhanced by lactate catabolism. Thus, metabolic symbiosis is established in the face of angiogenesis inhibition, whereby hypoxic cancer cells import glucose and export lactate, while normoxic cells import and catabolize lactate. mTOR signaling inhibition disrupts this metabolic symbiosis, associated with upregulation of the glucose transporter GLUT2.

  View details for DOI 10.1016/j.celrep.2016.04.029

  View details for Web of Science ID 000376164800003

  View details for PubMedID 27134166

  View details for PubMedCentralID PMC4872464

• Trimming the Vascular Tree in Tumors: Metabolic and Immune Adaptations Allen, E., Missiaen, R., Bergers, G., Stewart, D., Stillman, B. COLD SPRING HARBOR LABORATORY PRESS. 2016: 21-29

  Abstract

  Angiogenesis, the formation of new blood vessels, has become a well-established hallmark of cancer. Its functional importance for the manifestation and progression of tumors has been further validated by the beneficial therapeutic effects of angiogenesis inhibitors, most notably ones targeting the vascular endothelial growth factor (VEGF) signaling pathways. However, with the transient and short-lived nature of the patient response, it has become evident that tumors have the ability to adapt to the pressures of vascular growth restriction. Several escape mechanisms have been described that adapt tumors to therapy-induced low-oxygen tension by either reinstating tumor growth by vascular rebound or by altering tumor behavior without the necessity to reinitiate revascularization. We review here two bypass mechanisms that either instigate angiogenic and immune-suppressive polarization of intratumoral innate immune cells to facilitate VEGF-independent angiogenesis or enable metabolic adaptation and reprogramming of endothelial cells and tumor cells to adapt to low-oxygen tension.

  View details for DOI 10.1101/sqb.2016.81.030940

  View details for Web of Science ID 000483100500003

  View details for PubMedID 28396525

  View details for PubMedCentralID PMC8335596

• Development of a Bioluminescent Nitroreductase Probe for Preclinical Imaging PLOS ONE Vorobyeva, A. G., Stanton, M., Godinat, A., Lund, K. B., Karateev, G. G., Francis, K. P., Allen, E., Gelovani, J. G., McCormack, E., Tangney, M., Dubikovskaya, E. A. 2015; 10 (6): e0131037

  Abstract

  Bacterial nitroreductases (NTRs) have been widely utilized in the development of novel antibiotics, degradation of pollutants, and gene-directed enzyme prodrug therapy (GDEPT) of cancer that reached clinical trials. In case of GDEPT, since NTR is not naturally present in mammalian cells, the prodrug is activated selectively in NTR-transformed cancer cells, allowing high efficiency treatment of tumors. Currently, no bioluminescent probes exist for sensitive, non-invasive imaging of NTR expression. We therefore developed a "NTR caged luciferin" (NCL) probe that is selectively reduced by NTR, producing light proportional to the NTR activity. Here we report successful application of this probe for imaging of NTR in vitro, in bacteria and cancer cells, as well as in vivo in mouse models of bacterial infection and NTR-expressing tumor xenografts. This novel tool should significantly accelerate the development of cancer therapy approaches based on GDEPT and other fields where NTR expression is important.

  View details for DOI 10.1371/journal.pone.0131037

  View details for Web of Science ID 000356933800111

  View details for PubMedID 26110789

  View details for PubMedCentralID PMC4482324

• Nemitin, a Novel Map8/Map1s Interacting Protein with Wd40 Repeats PLOS ONE Wang, W., Lundin, V. F., Millan, I., Zeng, A., Chen, X., Yang, J., Allen, E., Chen, N., Bach, G., Hsu, A., Maloney, M. T., Kapur, M., Yang, Y. 2012; 7 (4)

  Abstract

  In neurons, a highly regulated microtubule cytoskeleton is essential for many cellular functions. These include axonal transport, regional specialization and synaptic function. Given the critical roles of microtubule-associated proteins (MAPs) in maintaining and regulating microtubule stability and dynamics, we sought to understand how this regulation is achieved. Here, we identify a novel LisH/WD40 repeat protein, tentatively named nemitin (neuronal enriched MAP interacting protein), as a potential regulator of MAP8-associated microtubule function. Based on expression at both the mRNA and protein levels, nemitin is enriched in the nervous system. Its protein expression is detected as early as embryonic day 11 and continues through adulthood. Interestingly, when expressed in non-neuronal cells, nemitin displays a diffuse pattern with puncta, although at the ultrastructural level it localizes along the microtubule network in vivo in sciatic nerves. These results suggest that the association of nemitin to microtubules may require an intermediary protein. Indeed, co-expression of nemitin with microtubule-associated protein 8 (MAP8) results in nemitin losing its diffuse pattern, instead decorating microtubules uniformly along with MAP8. Together, these results imply that nemitin may play an important role in regulating the neuronal cytoskeleton through an interaction with MAP8.

  View details for DOI 10.1371/journal.pone.0033094

  View details for Web of Science ID 000305345000007

  View details for PubMedID 22523538

  View details for PubMedCentralID PMC3327699

• Brivanib, a Dual FGF/VEGF Inhibitor, Is Active Both First and Second Line against Mouse Pancreatic Neuroendocrine Tumors Developing Adaptive/Evasive Resistance to VEGF Inhibition CLINICAL CANCER RESEARCH Allen, E., Walters, I. B., Hanahan, D. 2011; 17 (16): 5299-5310

  Abstract

  Preclinical trials of a mouse model of pancreatic neuroendocrine tumors (PNET) were conducted to determine whether dual FGF/VEGF pathway inhibition with brivanib can improve first-line efficacy in comparison with VEGF inhibitors lacking fibroblast growth factor (FGF)-inhibitory activity and to characterize second-line brivanib activity before and after the onset of evasive resistance to VEGF-selective therapy.An anti-VEGFR2 monoclonal antibody (DC101), an inhibitor of FGF signaling (FGF ligand trap), sorafenib, and brivanib were comparatively evaluated in first-line monotherapy in short and longer term fixed endpoint intervention trials in the RIP-Tag2 mouse model of PNET. Brivanib was also tested second line aiming to block adaptive resistance to selective VEGF therapies, assessing tumor growth, vascularity, hypoxia, invasion, and metastasis. The effects of initiating second-line brivanib therapy prior to or following overt relapse on sorafenib therapy were compared in overall survival trials to first-line therapies.Brivanib produced enduring tumor stasis and angiogenic blockade, both first and second line following the failure of DC101 or sorafenib. Overall survival was significantly extended by brivanib versus sorafenib, both first-line and when second-line therapy was initiated prior to sorafenib failure; second-line brivanib was less beneficial when initiated later, after the initiation of revascularization and incipient tumor progression.Brivanib holds promise and deserves consideration for clinical evaluation as an antiangiogenic therapy, both in the context of impending failures of VEGF-selective therapy and in a first-line setting aiming to limit the adaptive response to VEGF inhibitors that results in evasive resistance.

  View details for DOI 10.1158/1078-0432.CCR-10-2847

  View details for Web of Science ID 000293843700011

  View details for PubMedID 21622725

  View details for PubMedCentralID PMC3156934

• Antiangiogenic Therapy Elicits Malignant Progression of Tumors to Increased Local Invasion and Distant Metastasis CANCER CELL Paez-Ribes, M., Allen, E., Hudock, J., Takeda, T., Okuyama, H., Vinals, F., Inoue, M., Bergers, G., Hanahan, D., Casanovas, O. 2009; 15 (3): 220-231

  Abstract

  Multiple angiogenesis inhibitors have been therapeutically validated in preclinical cancer models, and several in clinical trials. Here we report that angiogenesis inhibitors targeting the VEGF pathway demonstrate antitumor effects in mouse models of pancreatic neuroendocrine carcinoma and glioblastoma but concomitantly elicit tumor adaptation and progression to stages of greater malignancy, with heightened invasiveness and in some cases increased lymphatic and distant metastasis. Increased invasiveness is also seen by genetic ablation of the Vegf-A gene in both models, substantiating the results of the pharmacological inhibitors. The realization that potent angiogenesis inhibition can alter the natural history of tumors by increasing invasion and metastasis warrants clinical investigation, as the prospect has important implications for the development of enduring antiangiogenic therapies.

  View details for DOI 10.1016/j.ccr.2009.01.027

  View details for Web of Science ID 000263998200009

  View details for PubMedID 19249680

  View details for PubMedCentralID PMC2874829

• Giant axonal neuropathy CELLULAR AND MOLECULAR LIFE SCIENCES Yang, Y., Allen, E., Ding, J., Wang, W. 2007; 64 (5): 601-609

  Abstract

  Giant axonal neuropathy (GAN) is a rare autosomal recessive disorder affecting both the central and peripheral nervous systems. Cytopathologically, the disorder is characterized by giant axons with derangements of cytoskeletal components. Geneticists refined the chromosomal interval containing the locus, culminating in the cloning of the defective gene, GAN. To date, many distinct mutations scattered throughout the coding region of the locus have been reported by researchers from different groups around the world. GAN encodes the protein, gigaxonin. Recently, a genetic mouse model of the disease was generated by targeted disruption of the locus. Over the years, the molecular mechanisms underlying GAN have attracted much interest. Studies have revealed that gigaxonin appears to play an important role in cytoskeletal functions and dynamics by directing ubiquitin-mediated degradations of cytoskeletal proteins. Aberrant accumulations of cytoskeletal-associated proteins caused by a defect in the ubiquitin-proteasome system (UPS) have been shown to be responsible for neurodegeneration occurring in GAN-null neurons, providing strong support for the notion that UPS plays crucial roles in cytoskeletal functions and dynamics. However, many key questions about the disease remain unanswered.

  View details for DOI 10.1007/s00018-007-6396-4

  View details for Web of Science ID 000244713900009

  View details for PubMedID 17256086

• Gene targeting of GAN in mouse causes a toxic accumulation of microtubule-associated protein 8 and impaired retrograde axonal transport HUMAN MOLECULAR GENETICS Ding, J. Q., Allen, E., Wang, W., Valle, A., Wu, C. B., Nardine, T., Cui, B. X., Yi, J., Taylor, A., Jeon, N. L., Chu, S., So, Y., Vogel, H., Tolwani, R., Mobley, W., Yang, Y. M. 2006; 15 (9): 1451-1463

  Abstract

  Mutations in gigaxonin were identified in giant axonal neuropathy (GAN), an autosomal recessive disorder. To understand how disruption of gigaxonin's function leads to neurodegeneration, we ablated the gene expression in mice using traditional gene targeting approach. Progressive neurological phenotypes and pathological lesions that developed in the GAN null mice recapitulate characteristic human GAN features. The disruption of gigaxonin results in an impaired ubiquitin-proteasome system leading to a substantial accumulation of a novel microtubule-associated protein, MAP8, in the null mutants. Accumulated MAP8 alters the microtubule network, traps dynein motor protein in insoluble structures and leads to neuronal death in cultured wild-type neurons, which replicates the process occurring in GAN null mutants. Defective axonal transport is evidenced by the in vitro assays and is supported by vesicular accumulation in the GAN null neurons. We propose that the axonal transport impairment may be a deleterious consequence of accumulated, toxic MAP8 protein.

  View details for DOI 10.1093/hmg/ddl069

  View details for PubMedID 16565160

• Microtubule-associated protein 8 contains two microtubule binding sites BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS Ding, J. Q., Valle, A., Allen, E., Wang, W., Nardine, T., Zhang, Y. J., Peng, L. L., Yang, Y. M. 2006; 339 (1): 172-179

  Abstract

  Microtubule-associated proteins (MAPs) are critical regulators of microtubule dynamics and functions, and have long been proposed to be essential for many cellular events including neuronal morphogenesis and functional maintenance. In this study, we report the characterization of a new microtubule-associated protein, we named MAP8. The protein of MAP8 is mainly restricted to the nervous system postnatally in mouse. Its expression could first be detected as early as at embryonic day 10, levels plateau during late embryonic and neonatal periods, and subsequently decrease moderately to remain constant into adulthood. In addition to its carboxyl terminal binding site, the MAP8 polyprotein also contains a functional microtubule-binding domain at its N-terminal segment. The association of the carboxyl terminal of the light chain with actin microfilaments could also be detected. Our findings define MAP8 as a novel microtubule associated protein containing two microtubule binding domains.

  View details for DOI 10.1016/j.bbrc.2005.10.199

  View details for PubMedID 16297881

• Gigaxonin interacts with tubulin folding cofactor B and controls its degradation through the ubiquitin-proteasome pathway CURRENT BIOLOGY Wang, W., Ding, J. Q., Allen, E., Zhu, P., Zhang, L., Vogel, H., Yang, Y. M. 2005; 15 (22): 2050-2055

  Abstract

  Gigaxonin is mutated in human giant axonal neuropathy (GAN), an autosomal recessive neurodegenerative disorder. The presence of generalized cytoskeletal abnormalities , including few microtubules and accumulated intermediate filaments (IFs), in GAN suggests an essential role of gigaxonin in cytoskeletal organization and dynamics. However, the molecular mechanisms underlying the cytoskeletal pathology remain to be elucidated. Over the years, the ubiquitin-proteasome system (UPS) of intracellular protein degradation has been implicated in the control of many fundamental cellular processes. Defects in this system seem to be directly linked to the development of human diseases, including cancers and neurodegenerative diseases . Here, we show that gigaxonin controls protein degradation of tubulin folding cofactor B (TBCB) , a function disrupted by GAN-associated mutations. The substantial TBCB protein accumulation caused by impaired UPS may be a causative factor of cytoskeletal pathology in GAN. Our study provides important insight into pathogenesis of neurodegenerative diseases associated with cytoskeletal abnormalities.

  View details for DOI 10.1016/j.cub.2005.10.052

  View details for PubMedID 16303566

• Gigaxonin-controlled degradation of MAP1B light chain is critical to neuronal survival NATURE Allen, E., Ding, J. Q., Wang, W., Pramanik, S., Chou, J., Yau, V., Yang, Y. M. 2005; 438 (7065): 224-228

  Abstract

  Giant axonal neuropathy (GAN) is a devastating sensory and motor neuropathy caused by mutations in the GAN gene, which encodes the ubiquitously expressed protein gigaxonin. Cytopathological features of GAN include axonal degeneration, with accumulation and aggregation of cytoskeletal components. Little is currently known about the molecular mechanisms underlying this recessive disorder. Here we show that gigaxonin controls protein degradation, and is essential for neuronal function and survival. We present evidence that gigaxonin binds to the ubiquitin-activating enzyme E1 through its amino-terminal BTB domain, while the carboxy-terminal kelch repeat domain interacts directly with the light chain (LC) of microtubule-associated protein 1B (MAP1B). Overexpression of gigaxonin leads to enhanced degradation of MAP1B-LC, which can be antagonized by proteasome inhibitors. Ablation of gigaxonin causes a substantial accumulation of MAP1B-LC in GAN-null neurons. Moreover, we show that overexpression of MAP1B in wild-type cortical neurons leads to cell death characteristic of GAN-null neurons, whereas reducing MAP1B levels significantly improves the survival rate of null neurons. Our results identify gigaxonin as a ubiquitin scaffolding protein that controls MAP1B-LC degradation, and provide insight into the molecular mechanisms underlying human neurodegenerative disorders.

  View details for DOI 10.1038/nature04256

  View details for Web of Science ID 000233133500049

  View details for PubMedID 16227972

• BPAG1 n4 is essential for retrograde axonal transport in sensory neurons JOURNAL OF CELL BIOLOGY Liu, J. J., Ding, J. Q., Kowal, A. S., Nardine, T., Allen, E., Delcroix, J. D., Wu, C. B., Mobley, W., Fuchs, E., Yang, Y. M. 2003; 163 (2): 223-229

  Abstract

  Disruption of the BPAG1 (bullous pemphigoid antigen 1) gene results in progressive deterioration in motor function and devastating sensory neurodegeneration in the null mice. We have previously demonstrated that BPAG1n1 and BPAG1n3 play important roles in organizing cytoskeletal networks in vivo. Here, we characterize functions of a novel BPAG1 neuronal isoform, BPAG1n4. Results obtained from yeast two-hybrid screening, blot overlay binding assays, and coimmunoprecipitations demonstrate that BPAG1n4 interacts directly with dynactin p150Glued through its unique ezrin/radixin/moesin domain. Studies using double immunofluorescent microscopy and ultrastructural analysis reveal physiological colocalization of BPAG1n4 with dynactin/dynein. Disruption of the interaction between BPAG1n4 and dynactin results in severe defects in retrograde axonal transport. We conclude that BPAG1n4 plays an essential role in retrograde axonal transport in sensory neurons. These findings might advance our understanding of pathogenesis of axonal degeneration and neuronal death.

  View details for DOI 10.1083/jcb.200306075

  View details for Web of Science ID 000186331100004

  View details for PubMedID 14581450

  View details for PubMedCentralID PMC2173519

• Discovery of gene families and alternatively spliced variants by RecA-mediated cloning GENOMICS Zeng, H., Allen, E., Lehman, C. W., Sargent, R. G., Pati, S., Zarling, D. A. 2002; 80 (5): 543-551

  Abstract

  Probing the functional complexity of the human genome will require new gene cloning techniques, not only to discover intraspecies gene homologs and interspecies gene orthologs, but also to identify alternatively spliced gene variants. We report homologous cDNA cloning methods that allow cloning of gene family members, genes from different species, and alternatively spliced gene variants. We cloned human 14-3-3 gene family members using DNA probes with as much as 35% sequence divergence, cloned alternatively spliced gene forms of Rad51D, and cloned a novel splice form of the human 14-3-3 theta gene with a unique expression pattern. Interspecies gene cloning was demonstrated for the mouse Rad51C and mouse beta-actin genes using human gene probes. The gene family cloning method is fast, efficient, and free from PCR errors; moreover, it exploits the abilities of RecA protein to pair homologous or partially homologous DNA sequences stably in kinetically trapped, multistranded DNA hybrids that can be used for subsequent gene clone enrichment.

  View details for DOI 10.1006/geno.2002.6855

  View details for Web of Science ID 000178994300013

  View details for PubMedID 12408972

• Genetic disorders of the cytoskeleton Fuchs, E., Yang, Y., Dowling, J., Lo, S., Yu, Q., Allen, E., Degenstein, L., Lee, E. FEDERATION AMER SOC EXP BIOL. 1997: A1006

  View details for Web of Science ID 000073305601356

• Mice expressing a mutant desmosomal cadherin exhibit abnormalities in desmosomes, proliferation, and epidermal differentiation JOURNAL OF CELL BIOLOGY Allen, E., Yu, Q. C., Fuchs, E. 1996; 133 (6): 1367-1382

  Abstract

  Desmogleins are members of the cadherin superfamily which form the core of desmosomes. In vitro studies indicate that the cytoplasmic domain of desmogleins associates with plakoglobin; however, little is known about the role of this domain in desmosome recognition or assembly in vivo, or about the possible relation of desmoglein mutations to epidermal differentiation and disease. To address these questions we used transgenic mouse technology to produce an NH2-terminally truncated desmoglein (Pemphigus Vulgaris Antigen or Dsg3) in cells known to express its wild-type counterpart. Within 2 d, newborn transgenic animals displayed swelling of their paws, flakiness on their back, and blackening of the tail tip. When analyzed histologically and ultrastructurally, widening of intercellular spaces and disruption of desmosomes were especially striking in the paws and tail. Desmosomes were reduced dramatically in number and were smaller and often peculiar in structure. Immunofluorescence and immunoelectron microscopy revealed no major abnormalities in localization of hemidesmosomal components, but desmosomal components organized aberrantly, resulting in a loss of ultrastructure within the plaque. In regions where desmosome loss was prevalent but where some adhesive structures persisted, the epidermis was thickened, with a marked increase in spinous and stratum corneum layers, variability in granular layer thickness, and parakeratosis in some regions. Intriguingly, a dramatic increase in cell proliferation was also observed concomitant with biochemical changes, including alterations in integrin expression, known to be associated with hyperproliferation. An inflammatory response was also detected in some skin regions. Collectively, these findings demonstrate that a mutation in a desmoglein can perturb epidermal cell-cell adhesion, triggering a cascade of changes in the skin.

  View details for DOI 10.1083/jcb.133.6.1367

  View details for Web of Science ID A1996UT40700018

  View details for PubMedID 8682871

  View details for PubMedCentralID PMC2120897

• THE HUMAN X-LINKED STEROID SULFATASE GENE AND A Y-ENCODED PSEUDOGENE - EVIDENCE FOR AN INVERSION OF THE Y-CHROMOSOME DURING PRIMATE EVOLUTION CELL YEN, P. H., MARSH, B., ALLEN, E., TSAI, S. P., ELLISON, J., CONNOLLY, L., NEISWANGER, K., SHAPIRO, L. J. 1988; 55 (6): 1123-1135

  Abstract

  The mammalian X and Y chromosomes are thought to have evolved from a common, nearly homologous chromosome pair. Although there is little sequence similarity between the mouse or the human X and Y, there are several regions in which moderate to extensive sequence homologies have been found, including, but not limited to, the so-called pseudoautosomal segment, in which X-Y pairing and recombination take place. The steroid sulfatase gene is in the pseudoautosomal region of the mouse, but not in man. We have cloned and characterized the human STS X-encoded locus and a pseudogene that is present on the long arm of the Y chromosome. Our data in humans and other primates suggest that there has been a pericentric inversion of the Y chromosome during primate evolution that has disrupted the former pseudoautosomal arrangement of these genes. These results provide additional insight into the evolution of the sex chromosomes and into the nature of this interesting portion of the human genome.

  View details for DOI 10.1016/0092-8674(88)90257-7

  View details for Web of Science ID A1988R562400021

  View details for PubMedID 3203382

• CLONING AND EXPRESSION OF STEROID SULFATASE CDNA AND THE FREQUENT OCCURRENCE OF DELETIONS IN STS DEFICIENCY - IMPLICATIONS FOR X-Y INTERCHANGE CELL YEN, P. H., ALLEN, E., MARSH, B., MOHANDAS, T., WANG, N., TAGGART, R. T., SHAPIRO, L. J. 1987; 49 (4): 443-454

  Abstract

  Human STS is a microsomal enzyme important in steroid metabolism. The gene encoding STS is pseudoautosomal in the mouse but not in humans, and escapes X inactivation in both species. We have prepared monoclonal and polyclonal antibodies to the protein which has been purified and from which partial amino acid sequence data have been obtained. cDNA clones containing the entire coding sequence were isolated, sequenced, and expressed in heterologous cells. Variable length transcripts have been shown to be present and due to usage of alternative poly(A) addition sites. The functional gene maps to Xp22.3-Xpter and there is a pseudogene on Yq suggesting a recent pericentric inversion. Absence of STS enzymatic activity occurs frequently in human populations and produces a visible phenotype of scaly skin or ichthyosis. Ten patients with inherited STS deficiency were studied and eight had complete gene deletions. The possibility that STS deficiency results from aberrant X-Y interchange is discussed.

  View details for DOI 10.1016/0092-8674(87)90447-8

  View details for Web of Science ID A1987H430500003

  View details for PubMedID 3032454