Digital Commons Network^™

Superior Antitumor Immune Response Achieved With Proton Over Photon Immunoradiotherapy Is Amplified By The Nanoradioenhancer Nbtxr3, Yun Hu, Sébastien Paris, Narayan Sahoo, Qi Wang, Qianxia Wang, Hampartsoum B Barsoumian, Ailing Huang, Jordan Da Silva, Célia Bienassis, Claudia S Kettlun Leyton, Tiffany A Voss, Fatemeh Masrorpour, Thomas Riad, Carola Leuschner, Nahum Puebla-Osorio, Saumil Gandhi, Quynh-Nhu Nguyen, Jing Wang, Maria Angelica Cortez, James W Welsh

Faculty, Staff and Student Publications

Recent findings suggest that immunoradiotherapy (IRT), combining photon radiotherapy (XRT) or proton radiotherapy (PRT) with immune checkpoint blockade, can enhance systemic tumor control. However, the comparative efficacy of XRT and PRT in IRT remains understudied. To address this, we compared outcomes between XRT + αPD1 and PRT + αPD1 in murine αPD1-resistant lung cancer (344SQR). We also assessed the impact of the nanoparticle radioenhancer NBTXR3 on both XRT + αPD1 and PRT + αPD1 for tumor control and examined the tumor immune microenvironment using single-cell RNA sequencing (scRNAseq). Additionally, mice cured by NBTXR3 + PRT + αPD1 were rechallenged with …

Preventive Treatment With A Cd73 Small Molecule Inhibitor Enhances Immune Surveillance In K-Ras Mutant Pancreatic Intraepithelial Neoplasia, Lincoln N Strickland, Wendao Liu, Usama Hussein, Nicolette Mardik, Xian Chen, Tingting Mills, Lana A Vornik, Michelle I Savage, Shizuko Sei, John Clifford, Holger K Eltzschig, Powel H Brown, Zhongming Zhao, Florencia Mcallister, Jennifer M Bailey-Lundberg

Faculty, Staff and Student Publications

Immunoprevention is an emerging consideration for solid tumors, including pancreatic ductal adenocarcinoma (PDAC). We and others have shown that Kras mutations in genetic models of spontaneous pancreatic intraepithelial neoplasia (PanIN), which is a precursor to PDAC, results in CD73 expression in the neoplastic epithelium and some populations of infiltrating immune cells, including macrophages and CD8 T cells. CD73 is an ecto-enzyme that converts extracellular adenosine monophosphate to adenosine, a critical immune inhibitory molecule in PDAC. We hypothesized inhibition of CD73 would reduce the incidence of PanIN formation and alter the immune microenvironment. To test our hypothesis, we used the KrasG12D; …

Benefits Of Calorie Restriction In Mice Are Mediated Via Energy Imbalance, Not Absolute Energy Or Protein Intake, Daniel L Smith, Sharon E Mitchell, Maria S Johnson, Victoria K Gibbs, Stephanie Dickinson, Beate Henschel, Rui Li, Kathryn A Kaiser, Daniella E Chusyd, Andrew W Brown, David B Allison, John R Speakman, Tim R Nagy

Children’s Nutrition Research Center Staff Publications

Caloric restriction (CR) results in reduced energy and protein intake, raising questions about protein restriction's contribution to CR longevity benefits. We kept ad libitum (AL)-fed male C57BL/6J mice at 27°C (AL27) and pair-fed (PF) mice at 22°C (22(PF27)). The 22(PF27) group was fed to match AL27 while restricted for calories due to cold-induced metabolism. The 22(PF27) mice had significantly lower body weight, lean mass, fat mass, leptin, IGF-1, and TNF-α levels than AL27 mice (p< 0.001 for all). Manipulations over ~11 weeks resulted in significant differences in body temperature, physical activity, and expression of key genes linked to hunger in the hypothalamus. Survival was significantly greater in 22(PF27) compared to AL27 overall (p< 0.001). CR in the context of equivalent energy and protein intake resulted in hormonal, metabolic, and physiological benefits and extended longevity. Hence, energy imbalance, rather than low energy or protein intake per se, mediates the benefits of CR.

Ficture: Scalable Segmentation-Free Analysis Of Submicron-Resolution Spatial Transcriptomics, Yichen Si, Changhee Lee, Yongha Hwang, Jeong H Yun, Weiqiu Cheng, Chun-Seok Cho, Miguel Quiros, Asma Nusrat, Weizhou Zhang, Goo Jun, Sebastian Zöllner, Jun Hee Lee, Hyun Min Kang

Faculty, Staff and Student Publications

Spatial transcriptomics (ST) technologies have advanced to enable transcriptome-wide gene expression analysis at submicron resolution over large areas. However, analysis of high-resolution ST is often challenged by complex tissue structure, where existing cell segmentation methods struggle due to the irregular cell sizes and shapes, and by the absence of segmentation-free methods scalable to whole-transcriptome analysis. Here we present FICTURE (Factor Inference of Cartographic Transcriptome at Ultra-high REsolution), a segmentation-free spatial factorization method that can handle transcriptome-wide data labeled with billions of submicron-resolution spatial coordinates and is compatible with both sequencing-based and imaging-based ST data. FICTURE uses the multilayered Dirichlet model …

The Five Homologous Ciar-Controlled Ccn Srnas Of Streptococcus Pneumoniae Modulate Zn-Resistance, Nicholas R De Lay, Nidhi Verma, Dhriti Sinha, Abigail Garrett, Maximillian K Osterberg, Daisy Porter, Spencer Reiling, David P Giedroc, Malcolm E Winkler

Faculty, Staff and Student Publications

Zinc is a vital transition metal for all bacteria; however, elevated intracellular free Zn levels can result in mis-metalation of Mn-dependent enzymes. For Mn-centric bacteria such as Streptococcus pneumoniae that primarily use Mn instead of Fe as an enzyme cofactor, Zn is particularly toxic at high concentrations. Here, we report our identification and characterization of the function of the five homologous, CiaRH-regulated Ccn sRNAs in controlling S. pneumoniae virulence and metal homeostasis. We show that deletion of all five ccn genes (ccnA, ccnB, ccnC, ccnD, and ccnE) from S. pneumoniae strains D39 (serotype 2) and TIGR4 (serotype 4) causes Zn …

Dual Targeting Macrophages And Microglia Is A Therapeutic Vulnerability In Models Of Pten-Deficient Glioblastoma, Yang Liu, Junyan Wu, Hinda Najem, Yiyun Lin, Lizhi Pang, Fatima Khan, Fei Zhou, Heba Ali, Amy B Heimberger, Peiwen Chen

Faculty, Staff and Student Publications

Tumor-associated macrophages and microglia (TAMs) are critical for tumor progression and therapy resistance in glioblastoma (GBM), a type of incurable brain cancer. We previously identified lysyl oxidase (LOX) and olfactomedin like-3 (OLFML3) as essential macrophage and microglia chemokines, respectively, in GBM. Here, single-cell transcriptomics and multiplex sequential immunofluorescence followed by functional studies demonstrate that macrophages negatively correlate with microglia in the GBM tumor microenvironment. LOX inhibition in PTEN-deficient GBM cells upregulates OLFML3 expression via the NF-κB-PATZ1 signaling pathway, inducing a compensatory increase of microglia infiltration. Dual targeting macrophages and microglia via inhibition of LOX and the CLOCK-OLFML3 axis generates potent …

Inulin-Gel-Based Oral Immunotherapy Remodels The Small Intestinal Microbiome And Suppresses Food Allergy, Kai Han, Fang Xie, Olamide Animasahun, Minal Nenwani, Sho Kitamoto, Yeji Kim, May Thazin Phoo, Jin Xu, Fulei Wuchu, Kehinde Omoloja, Abhinav Achreja, Srinadh Choppara, Zhaoheng Li, Wang Gong, Young Seok Cho, Hannah Dobson, Jinsung Ahn, Xingwu Zhou, Xuehui Huang, Xinran An, Alexander Kim, Yao Xu, Qi Wu, Soo-Hong Lee, Jessica J O'Konek, Yuying Xie, Yu Leo Lei, Nobuhiko Kamada, Deepak Nagrath, James J Moon

Faculty, Staff and Student Publications

Despite the potential of oral immunotherapy against food allergy, adverse reactions and loss of desensitization hinder its clinical uptake. Dysbiosis of the gut microbiota is implicated in the increasing prevalence of food allergy, which will need to be regulated to enable for an effective oral immunotherapy against food allergy. Here we report an inulin gel formulated with an allergen that normalizes the dysregulated ileal microbiota and metabolites in allergic mice, establishes allergen-specific oral tolerance and achieves robust oral immunotherapy efficacy with sustained unresponsiveness in food allergy models. These positive outcomes are associated with enhanced allergen uptake by antigen-sampling dendritic cells …

Astrocyte-Induced Cdk5 Expedites Breast Cancer Brain Metastasis By Suppressing Mhc-I Expression To Evade Immune Recognition, Arseniy E Yuzhalin, Frank J Lowery, Yohei Saito, Xiangliang Yuan, Jun Yao, Yimin Duan, Jingzhen Ding, Sunil Acharya, Chenyu Zhang, Abigail Fajardo, Hao-Nien Chen, Yongkun Wei, Yutong Sun, Lin Zhang, Yi Xiao, Ping Li, Philip L Lorenzi, Jason T Huse, Huihui Fan, Zhongming Zhao, Mien-Chie Hung, Dihua Yu

Faculty, Staff and Student Publications

Brain metastases (BrMs) evade the immune response to develop in the brain, yet the mechanisms of BrM immune evasion remains unclear. This study shows that brain astrocytes induce the overexpression of neuronal-specific cyclin-dependent kinase 5 (Cdk5) in breast cancer-derived BrMs, which facilitates BrM outgrowth in mice. Cdk5-overexpressing BrMs exhibit reduced expression and function of the class I major histocompatibility complex (MHC-I) and antigen-presentation pathway, which are restored by inhibiting Cdk5 genetically or pharmacologically, as evidenced by single-cell RNA sequencing and functional studies. Mechanistically, Cdk5 suppresses MHC-I expression on the cancer cell membrane through the Irf2bp1-Stat1-importin α-Nlrc5 pathway, enabling BrMs to …

Antiangiogenic Tyrosine Kinase Inhibitors Have Differential Efficacy In Clear Cell Renal Cell Carcinoma In Bone, Stefan Maksimovic, Nina C Boscolo, Ludovica La Posta, Sergio Barrios, Mohammad Jad Moussa, Emanuela Gentile, Pedro I Pesquera, Wenjiao Li, Jianfeng Chen, Javier A Gomez, Akshay Basi, Jared K Burks, Christopher Alvarez-Breckenridge, Jianjun Gao, Matthew T Campbell, Eleonora Dondossola

Faculty, Staff and Student Publications

Clear cell renal cell carcinoma (ccRCC) is the most prevalent kidney neoplasm; bone metastasis (BM) develops in 35% to 40% of metastatic patients and results in substantial morbidity and mortality, as well as medical costs. A key feature of ccRCC is the loss of function of the von Hippel-Lindau protein, which enhances angiogenesis via vascular endothelial growth factor release. Consequently, antiangiogenic tyrosine kinase inhibitors (TKI) emerged as a treatment for ccRCC. However, limited data about their efficacy in BM is available, and no systematic comparisons have been performed. We developed mouse models of bone and lung ccRCC tumors and compared …

Inhibition Of Hepatic Oxalate Overproduction Ameliorates Metabolic Dysfunction-Associated Steatohepatitis, Sandeep Das, Alexandra C Finney, Sumit Kumar Anand, Sumati Rohilla, Yuhao Liu, Nilesh Pandey, Alia Ghrayeb, Dhananjay Kumar, Kelley Nunez, Zhipeng Liu, Fabio Arias, Ying Zhao, Brenna H Pearson-Gallion, M Peyton Mckinney, Koral S E Richard, Jose A Gomez-Vidal, Chowdhury S Abdullah, Elizabeth D Cockerham, Joseph Eniafe, Andrew D Yurochko, Tarek Magdy, Christopher B Pattillo, Christopher G Kevil, Babak Razani, Md Shenuarin Bhuiyan, Erin H Seeley, Gretchen E Galliano, Bo Wei, Lin Tan, Iqbal Mahmud, Ida Surakka, Minerva T Garcia-Barrio, Philip L Lorenzi, Eyal Gottlieb, Eduardo Salido, Jifeng Zhang, A Wayne Orr, Wanqing Liu, Monica Diaz-Gavilan, Y Eugene Chen, Nirav Dhanesha, Paul T Thevenot, Ari J Cohen, Arif Yurdagul, Oren Rom

Faculty, Staff and Student Publications

The incidence of metabolic dysfunction-associated steatohepatitis (MASH) is on the rise, and with limited pharmacological therapy available, identification of new metabolic targets is urgently needed. Oxalate is a terminal metabolite produced from glyoxylate by hepatic lactate dehydrogenase (LDHA). The liver-specific alanine-glyoxylate aminotransferase (AGXT) detoxifies glyoxylate, preventing oxalate accumulation. Here we show that AGXT is suppressed and LDHA is activated in livers from patients and mice with MASH, leading to oxalate overproduction. In turn, oxalate promotes steatosis in hepatocytes by inhibiting peroxisome proliferator-activated receptor-α (PPARα) transcription and fatty acid β-oxidation and induces monocyte chemotaxis via C-C motif chemokine ligand 2. In …

Temporal Recording Of Mammalian Development And Precancer, Mirazul Islam, Yilin Yang, Alan J Simmons, Vishal M Shah, Krushna Pavan Musale, Yanwen Xu, Naila Tasneem, Zhengyi Chen, Linh T Trinh, Paola Molina, Marisol A Ramirez-Solano, Iannish D Sadien, Jinzhuang Dou, Andrea Rolong, Ken Chen, Mark A Magnuson, Jeffrey C Rathmell, Ian G Macara, Douglas J Winton, Qi Liu, Hamim Zafar, Reza Kalhor, George M Church, Martha J Shrubsole, Robert J Coffey, Ken S Lau

Faculty, Staff and Student Publications

Temporal ordering of cellular events offers fundamental insights into biological phenomena. Although this is traditionally achieved through continuous direct observations1,2, an alternative solution leverages irreversible genetic changes, such as naturally occurring mutations, to create indelible marks that enables retrospective temporal ordering3–5. Using a multipurpose, single-cell CRISPR platform, we developed a molecular clock approach to record the timing of cellular events and clonality in vivo, with incorporation of cell state and lineage information. Using this approach, we uncovered precise timing of tissue-specific cell expansion during mouse embryonic development, unconventional developmental relationships between cell …

Development Of An Engineered Extracellular Vesicles-Based Vaccine Platform For Combined Delivery Of Mrna And Protein To Induce Functional Immunity, Xin Luo, Kathleen M Mcandrews, Kent A Arian, Sami J Morse, Viktoria Boeker, Shreyasee V Kumbhar, Yingying Hu, Krishnan K Mahadevan, Kaira A Church, Sriram Chitta, Nicolas T Ryujin, Janine Hensel, Jianli Dai, Dara P Dowlatshahi, Hikaru Sugimoto, Michelle L Kirtley, Valerie S Lebleu, Shabnam Shalapour, Joe H Simmons, Raghu Kalluri

Faculty, Staff and Student Publications

mRNA incorporated in lipid nanoparticles (LNPs) became a new class of vaccine modality for induction of immunity against COVID-19 and ushered in a new era in vaccine development. Here, we report a novel, easy-to-execute, and cost effective engineered extracellular vesicles (EVs)-based combined mRNA and protein vaccine platform (EVX-M+P vaccine) and explore its utility in proof-of-concept immunity studies in the settings of cancer and infectious disease. As a first example, we engineered EVs, natural nanoparticle carriers shed by all cells, to contain ovalbumin mRNA and protein (EVOvaM+P vaccine) to serve as cancer vaccine against ovalbumin-expressing melanoma tumors. EVOvaM+P administration to mice …

Novel Genes Of The Male Reproductive System: Potential Roles In Male Reproduction And As Non-Hormonal Male Contraceptive Targets, Thomas X Garcia, Martin M Matzuk

Faculty, Staff and Students Publications

The development of novel non-hormonal male contraceptives represents a pivotal frontier in reproductive health, driven by the need for safe, effective, and reversible contraceptive methods. This comprehensive review explores the genetic underpinnings of male fertility, emphasizing the crucial roles of specific genes and structural variants (SVs) identified through advanced sequencing technologies such as long-read sequencing (LRS). LRS has revolutionized the detection of structural variants and complex genomic regions, offering unprecedented precision and resolution over traditional next-generation sequencing (NGS). Key genetic targets, including those implicated in spermatogenesis and sperm motility, are highlighted, showcasing their potential as non-hormonal contraceptive targets. The review …

A Mouse Model For Conditional Expression Of Activated Β-Catenin In Epidermal Keratinocytes, Vineet K Maurya, Yan Ying, John P Lydon

Faculty, Staff and Students Publications

We report the generation and characterization of the K5: CAT bigenic mouse in which the constitutively activated form of β-catenin (ΔN89 β-catenin) is conditionally expressed in cytokeratin-5 (K5) positive epidermal keratinocytes. Following short-term doxycycline intake during the telogen resting phase, the adult K5: CAT bigenic develops enlarged pilosebaceous units that expand deep into the dermis, an expansion usually observed during the anagen growth phase. Prolonged doxycycline treatment results in significant thickening and folding of the K5: CAT epidermis. During this persistent induction period, there is clear evidence of increased keratinocyte proliferation, particularly in the epidermal basal cell layer and the …

Pkmyt1 Is A Marker Of Treatment Response And A Therapeutic Target For Cdk4/6 Inhibitor-Resistance In Er+ Breast Cancer, Anran Chen, Beom-Jun Kim, Aparna Mitra, Craig T Vollert, Jonathan T Lei, Diana Fandino, Meenakshi Anurag, Matthew V Holt, Xuxu Gou, Jacob B Pilcher, Matthew P Goetz, Donald W Northfelt, Susan G Hilsenbeck, C Gary Marshall, Marc L Hyer, Robert Papp, Shou-Yun Yin, Carmine De Angelis, Rachel Schiff, Suzanne A W Fuqua, Cynthia X Ma, Charles E Foulds, Matthew J Ellis

Faculty, Staff and Students Publications

Endocrine therapies (ET) with cyclin-dependent kinase 4/6 (CDK4/6) inhibition are the standard treatment for estrogen receptor-α-positive (ER+) breast cancer, however drug resistance is common. In this study, proteogenomic analyses of patient-derived xenografts (PDXs) from patients with 22 ER+ breast cancer demonstrated that protein kinase, membrane-associated tyrosine/threonine one (PKMYT1), a WEE1 homolog, is estradiol (E2) regulated in E2-dependent PDXs and constitutively expressed when growth is E2-independent. In clinical samples, high PKMYT1 mRNA levels associated with resistance to both ET and CDK4/6 inhibition. The PKMYT1 inhibitor lunresertib (RP-6306) with gemcitabine selectively and synergistically reduced the viability of ET and palbociclib-resistant ER+ breast …

Linkage Between Fuz And Gpr161 Genes Regulates Sonic Hedgehog Signaling During Mouse Neural Tube Development, Sung-Eun Kim, Hyun-Yi Kim, Bogdan J Wlodarczyk, Richard H Finnell

Faculty, Staff and Students Publications

Sonic hedgehog (Shh) signaling regulates embryonic morphogenesis utilizing the primary cilium, the cell's antenna, which acts as a signaling hub. Fuz, an effector of planar cell polarity signaling, regulates Shh signaling by facilitating cilia formation, and the G protein-coupled receptor 161 (Gpr161) is a negative regulator of Shh signaling. The range of phenotypic malformations observed in mice bearing mutations in either of the genes encoding these proteins is similar; however, their functional relationship has not been previously explored. This study identified the genetic and biochemical linkage between Fuz and Gpr161 in mouse neural tube development. Fuz was found to be …

Galectin 3-Binding Protein (Lgals3bp) Depletion Attenuates Hepatic Fibrosis By Reducing Transforming Growth Factor-Β1 (Tgf-Β1) Availability And Inhibits Hepatocarcinogenesis, Dae-Hwan Kim, Minjeong Sung, Myong-Suk Park, Eun-Gene Sun, Sumin Yoon, Kyung Hyun Yoo, Kamalakannan Radhakrishnan, Sung Yun Jung, Woo-Kyun Bae, Sang-Hee Cho, Ik-Joo Chung

Faculty, Staff and Students Publications

Background: Increased Galectin 3-binding protein (LGALS3BP) serum levels have been used to assess hepatic fibrosis stages and the severity of hepatocellular carcinoma (HCC). Considering the crucial role of transforming growth factor-β1 (TGF-β1) in the emergence of these diseases, the present study tested the hypothesis that LGALS3BP regulates the TGF-β1 signaling pathway.

Methods: The expression levels of LGALS3BP and TGFB1 were analyzed in patients with metabolic dysfunction-associated steatohepatitis (MASH) and HCC. Multiple omics techniques, such as RNA-sequencing, transposase-accessible chromatin-sequencing assay, and liquid chromatography-tandem mass spectrometry proteomics, were used to identify the regulatory mechanisms for the LGALS3BP-TGF-β1 axis. The effects of altered …

Mouse Corneal Immune Cell Heterogeneity Revealed By Single-Cell Rna Sequencing, Ebru Yaman, Nicole Heyer, Cintia S De Paiva, Mary Ann Stepp, Stephen C Pflugfelder, Jehan Alam

Faculty, Staff and Students Publications

PURPOSE: This study aimed to define the heterogeneity, spatial localization, and functional roles of immune cells in the mouse cornea using single-cell RNA sequencing (scRNA-seq) and immunofluorescent staining.

METHODS: Enriched mouse corneal immune cells (C57BL/6 strain, age 16-20 weeks) underwent single-cell RNA sequencing library preparation, sequencing, and analysis with Seurat, Monocle 3, and CellChat packages in R. Pathway analysis used Qiagen Ingenuity Pathway Analysis software. Immunostaining confirmed cell distribution.

RESULTS: We identified 14 distinct immune cell clusters (56% myeloid and 44% lymphoid). Myeloid populations included resident macrophages, conventional dendritic cells (cDC2s), Langerhans cells, neutrophils, monocytes, and mast cells. Additionally, lymphocyte …

Htlv-1 Infected T Cells Cause Bone Loss Via Small Extracellular Vesicles, Nitin Kumar Pokhrel, Haniya Habib, Shamreethaa Seeniraj, Ancy Joseph, Daniel Rauch, Linda Cox, Robert Sprung, Petra Erdmann Gilmore, Qiang Zhang, Robert Reid Townsend, Lee Ratner, Katherine N Weilbaecher, Deborah J Veis, Et Al.

2020-Current year OA Pubs

Adult T cell leukaemia (ATL), caused by infection with human T- lymphotropic virus type 1 (HTLV-1), is often complicated by hypercalcemia and osteolytic lesions. Therefore, we studied the communication between patient-derived ATL cells (ATL-PDX) and HTLV-1 immortalized CD4+ T cell lines (HTLV/T) with osteoclasts and their effects on bone mass in mice. Intratibial inoculation of some HTLV/T leads to a profound local decrease in bone mass similar to marrow-replacing ATL-PDX, despite the fact that few HTLV/T cells persisted in the bone. To study the direct effect of HTLV/T and ATL-PDX on osteoclasts, supernatants were added to murine and human osteoclast …

Pkmyt1 Is A Marker Of Treatment Response And A Therapeutic Target For Cdk4/6 Inhibitor-Resistance In Er+ Breast Cancer, Anran Chen, Cynthia X Ma, Et Al.

2020-Current year OA Pubs

Endocrine therapies (ET) with cyclin-dependent kinase 4/6 (CDK4/6) inhibition are the standard treatment for estrogen receptor-α-positive (ER+) breast cancer, however drug resistance is common. In this study, proteogenomic analyses of patient-derived xenografts (PDXs) from patients with 22 ER+ breast cancer demonstrated that protein kinase, membrane-associated tyrosine/threonine one (PKMYT1), a WEE1 homolog, is estradiol (E2) regulated in E2-dependent PDXs and constitutively expressed when growth is E2-independent. In clinical samples, high PKMYT1 mRNA levels associated with resistance to both ET and CDK4/6 inhibition. The PKMYT1 inhibitor lunresertib (RP-6306) with gemcitabine selectively and synergistically reduced the viability of ET and palbociclib-resistant ER+ breast …

Sequential Early-Life Viral Infections Modulate The Microbiota And Adaptive Immune Responses To Systemic And Mucosal Vaccination, Yuhao Li, Jerome M. Molleston, Crystal Lovato, Jasmine Wright, Isabel Erickson, Duyen Bui, Andrew H. Kim, Harshad Ingle, Somya Aggarwal, Lila S Nolan, Ahmed O. Hassan, Lynne Foster, Michael S. Diamond, Megan T. Baldridge

2020-Current year OA Pubs

Increasing evidence points to the microbial exposome as a critical factor in maturing and shaping the host immune system, thereby influencing responses to immune challenges such as infections or vaccines. To investigate the effect of early-life viral exposures on immune development and vaccine responses, we inoculated mice with six distinct viral pathogens in sequence beginning in the neonatal period, and then evaluated their immune signatures before and after intramuscular or intranasal vaccination against SARS-CoV-2. Sequential viral infection drove profound changes in all aspects of the immune system, including increasing circulating leukocytes, altering innate and adaptive immune cell lineages in tissues, …

Impact Of Essential Genes On The Success Of Genome Editing Experiments Generating 3313 New Genetically Engineered Mouse Lines., Hillary Elrick, Kevin A Peterson, Brandon J Willis, Denise G Lanza, Elif F Acar, Edward J Ryder, Lydia Teboul, Petr Kasparek, Marie-Christine Birling, David J Adams, Allan Bradley, Robert E Braun, Steve D Brown, Adam Caulder, Gemma F Codner, Francesco J Demayo, Mary E Dickinson, Brendan Doe, Graham Duddy, Marina Gertsenstein, Leslie O Goodwin, Yann Hérault, Lauri G Lintott, K C Kent Lloyd, Isabel Lorenzo, Matthew Mackenzie, Ann-Marie Mallon, Colin Mckerlie, Helen Parkinson, Ramiro Ramirez-Solis, John R Seavitt, Radislav Sedlacek, William C Skarnes, Damien Smedley, Sara Wells, Jacqueline K White, Joshua A Wood, International Mouse Phenotyping Consortium, Stephen A Murray, Jason D Heaney, Lauryl M J Nutter

Faculty Research 2024

The International Mouse Phenotyping Consortium (IMPC) systematically produces and phenotypes mouse lines with presumptive null mutations to provide insight into gene function. The IMPC now uses the programmable RNA-guided nuclease Cas9 for its increased capacity and flexibility to efficiently generate null alleles in the C57BL/6N strain. In addition to being a valuable novel and accessible research resource, the production of 3313 knockout mouse lines using comparable protocols provides a rich dataset to analyze experimental and biological variables affecting in vivo gene engineering with Cas9. Mouse line production has two critical steps - generation of founders with the desired allele and …

Impact Of Essential Genes On The Success Of Genome Editing Experiments Generating 3313 New Genetically Engineered Mouse Lines, Hillary Elrick, Kevin A Peterson, Brandon J Willis, Denise G Lanza, Elif F Acar, Edward J Ryder, Lydia Teboul, Petr Kasparek, Marie-Christine Birling, David J Adams, Allan Bradley, Robert E Braun, Steve D Brown, Adam Caulder, Gemma F Codner, Francesco J Demayo, Mary E Dickinson, Brendan Doe, Graham Duddy, Marina Gertsenstein, Leslie O Goodwin, Yann Hérault, Lauri G Lintott, K C Kent Lloyd, Isabel Lorenzo, Matthew Mackenzie, Ann-Marie Mallon, Colin Mckerlie, Helen Parkinson, Ramiro Ramirez-Solis, John R Seavitt, Radislav Sedlacek, William C Skarnes, Damien Smedley, Sara Wells, Jacqueline K White, Joshua A Wood, International Mouse Phenotyping Consortium, Stephen A Murray, Jason D Heaney, Lauryl M J Nutter

Faculty, Staff and Students Publications

The International Mouse Phenotyping Consortium (IMPC) systematically produces and phenotypes mouse lines with presumptive null mutations to provide insight into gene function. The IMPC now uses the programmable RNA-guided nuclease Cas9 for its increased capacity and flexibility to efficiently generate null alleles in the C57BL/6N strain. In addition to being a valuable novel and accessible research resource, the production of 3313 knockout mouse lines using comparable protocols provides a rich dataset to analyze experimental and biological variables affecting in vivo gene engineering with Cas9. Mouse line production has two critical steps - generation of founders with the desired allele and …

S6k1 Controls Dna Damage Signaling Modulated By The Mrn Complex To Induce Radioresistance In Lung Cancer, Ali Calderon-Aparicio, Jun He, Nicole L. Simone

Department of Radiation Oncology Faculty Papers

Radiation is a mainstay of lung cancer treatment; however, resistance frequently develops. Identifying novel therapeutic targets to increase radiation sensitivity is crucial. S6K1 is a serine/threonine kinase known to regulate protein translation which is associated with radioresistance, but the mechanisms involved are unknown. We proposed to determine whether S6K1 promotes radioresistance by regulating DNA repair in lung cancer. Colony formation, protein expression and proliferation were assessed. S6K1 was modulated pharmacologically by either PF-4708671 or genetically by Crispr-Cas9. Higher radioresistance levels in lung cancer cells were associated with lower phosphoactivation of MRN complex members, a key activator of radiation-induced DNA repair …

Usf2 And Tfeb Compete In Regulating Lysosomal And Autophagy Genes, Jaebeom Kim, Young Suk Yu, Yehwa Choi, Do Hui Lee, Soobin Han, Junhee Kwon, Taichi Noda, Masahito Ikawa, Dongha Kim, Hyunkyung Kim, Andrea Ballabio, Keun Il Kim, Sung Hee Baek

Duncan NRI Faculty and Staff Publications

Autophagy, a highly conserved self-digestion process crucial for cellular homeostasis, is triggered by various environmental signals, including nutrient scarcity. The regulation of lysosomal and autophagy-related processes is pivotal to maintaining cellular homeostasis and basal metabolism. The consequences of disrupting or diminishing lysosomal and autophagy systems have been investigated; however, information on the implications of hyperactivating lysosomal and autophagy genes on homeostasis is limited. Here, we present a mechanism of transcriptional repression involving upstream stimulatory factor 2 (USF2), which inhibits lysosomal and autophagy genes under nutrient-rich conditions. We find that USF2, together with HDAC1, binds to the CLEAR motif within lysosomal …

Spatial Organization Of Adenylyl Cyclase And Its Impact On Dopamine Signaling In Neurons, Léa Ripoll, Yong Li, Carmen W Dessauer, Mark Von Zastrow

Faculty, Staff and Student Publications

The cAMP cascade is increasingly recognized to transduce physiological effects locally through spatially limited cAMP gradients. However, little is known about how adenylyl cyclase enzymes that initiate cAMP gradients are localized. Here we address this question in physiologically relevant striatal neurons and investigate how AC localization impacts downstream signaling function. We show that the major striatal AC isoforms are differentially sorted between ciliary and extraciliary domains of the plasma membrane, and that one isoform, AC9, is uniquely concentrated in endosomes. We identify key sorting determinants in the N-terminal cytoplasmic domain responsible for isoform-specific localization. We further show that AC9-containing endosomes …

An Approach To Analyze Spatiotemporal Patterns Of Gene Expression At Single-Cell Resolution In Candida Albicans-Infected Mouse Tongues, Elena Lindemann-Perez, Diana L Rodríguez, J Christian Pérez

Faculty, Staff and Student Publications

Microbial gene expression measurements derived from infected organs are invaluable to understand pathogenesis. However, current methods are limited to "bulk" analyses that neglect microbial cell heterogeneity and the lesion's spatial architecture. Here, we report the use of hybridization chain reaction RNA fluorescence in situ hybridization (HCR RNA-FISH) to visualize and quantify Candida albicans transcripts at single-cell resolution in tongues of infected mice. The method is compatible with fixed-frozen and formalin-fixed paraffin-embedded tissues. We document cell-to-cell variation and intriguing spatiotemporal expression patterns for C. albicans mRNAs that encode products implicated in oral candidiasis. The approach provides a spatial dimension to gene …

B Cells Drive Neuropathic Pain-Related Behaviors In Mice Through Igg-Fc Gamma Receptor Signaling, Michael J Lacagnina, Kendal F Willcox, Nabila Boukelmoune, Alexis Bavencoffe, Ishwarya Sankaranarayanan, Daniel T Barratt, Younus A Zuberi, Dorsa Dayani, Melissa V Chavez, Jonathan T Lu, Alex Bersellini Farinotti, Stephanie Shiers, Allison M Barry, Juliet M Mwirigi, Diana Tavares-Ferreira, Geoffrey A Funk, Anna M Cervantes, Camilla I Svensson, Edgar T Walters, Mark R Hutchinson, Cobi J Heijnen, Theodore J Price, Nathan T Fiore, Peter M Grace

Faculty, Staff and Student Publications

Neuroimmune interactions are essential for the development of neuropathic pain, yet the contributions of distinct immune cell populations have not been fully unraveled. Here, we demonstrate the critical role of B cells in promoting mechanical hypersensitivity (allodynia) after peripheral nerve injury in male and female mice. Depletion of B cells with a single injection of anti-CD20 monoclonal antibody at the time of injury prevented the development of allodynia. B cell-deficient (muMT) mice were similarly spared from allodynia. Nerve injury was associated with increased immunoglobulin G (IgG) accumulation in ipsilateral lumbar dorsal root ganglia (DRGs) and dorsal spinal cords. IgG was …

Dense And Persistent Odor Representations In The Olfactory Bulb Of Awake Mice, Delaram Pirhayati, Cameron L Smith, Ryan Kroeger, Saket Navlakha, Paul Pfaffinger, Jacob Reimer, Benjamin R Arenkiel, Ankit Patel, Elizabeth H Moss

Children’s Nutrition Research Center Staff Publications

Recording and analysis of neural activity are often biased toward detecting sparse subsets of highly active neurons, masking important signals carried in low-magnitude and variable responses. To investigate the contribution of seemingly noisy activity to odor encoding, we used mesoscale calcium imaging from mice of both sexes to record odor responses from the dorsal surface of bilateral olfactory bulbs (OBs). The outer layer of the mouse OB is comprised of dendrites organized into discrete "glomeruli," which are defined by odor receptor-specific sensory neuron input. We extracted activity from a large population of glomeruli and used logistic regression to classify odors …

Cd8Α Structural Domains Enhance Gucy2c Car-T Cell Efficacy, Trevor R. Baybutt, Ariana A. Entezari, Adi Caspi, Ross E. Staudt, Robert D. Carlson, Scott A. Waldman, Adam E. Snook

Department of Pharmacology, Physiology, and Cancer Biology Faculty Papers

Despite success in treating some hematological malignancies, CAR-T cells have not yet produced similar outcomes in solid tumors due, in part, to the tumor microenvironment, poor persistence, and a paucity of suitable target antigens. Importantly, the impact of the CAR components on these challenges remains focused on the intracellular signaling and antigen-binding domains. In contrast, the flexible hinge and transmembrane domains have been commoditized and are the least studied components of the CAR. Here, we compared the hinge and transmembrane domains derived from either the CD8ɑ or CD28 molecule in identical GUCY2C-targeted third-generation designs for colorectal cancer. While these structural …