Medical Cell Biology Commons^™

Measuring Mitochondrial Respiration In Vivo: From Mouse To Human, Arthur Orchanian, Brennan Schilling, Bruce Berkowitz Phd

Medical Student Research Symposium

Introduction: The mitochondrial energy ecosystem can be non-invasively interrogated in photoreceptors by combing a clinical tool, optical coherence tomography (OCT), with a mitochondrial protonophore (2,4 dinitrophenol, DNP). It remains unclear if only supra-clinical doses of DNP will be useful for mouse studies or if lower but clinically relevant doses of DNP would facilitate translation from mice to humans.

Methods: The experiment was a paired longitudinal design that took place over 2 days. On day 1, C57BL/6J mice were overnight dark adapted, then light-adapted for 5 h before OCT examination before regaining consciousness; a similar procedure was followed on day 2 …

Role Of Ataxia Telangiectasia Mutated In The Repair Of Magnetite Nanoparticles Induced Double Strand Breaks Associated With Heterochromatin, Mie Mohamed Mounir

Theses and Dissertations

Magnetite nanoparticles particulate matter pollution is escalating in large cities like New Mexico, Manchester city, Cairo, and New Delhi. Combustion derived Magnetite nanoparticles was found deposited in internal vital organs of deceased residents of urban areas. A great deal of research focused on the genotoxic and cytotoxic effects of Magnetite Nanoparticles (MNPs). In our work we focus on the mechanisms employed in DNA repair of higher chromatin structures. We employed the sensitive analysis of anti γ-H2AX to detect and monitor the repair of DSBs following exposure to MNPs. A 2- fold delay in the repair of heterochromatin associated DSBs in …

Characterization Of Abin-1 In The Traumatically Injured Brain, Aaron Orellana, Alana C. Conti

Medical Student Research Symposium

Traumatic brain injury (TBI) is associated with chronic pain and persistent neuroinflammation. Opioids are often prescribed in order to relieve pain symptoms, but recent evidence suggests that their use negatively impacts the neuropathology of TBI leading to behavioral impairments and exacerbated neuroinflammation. The de-ubiquinating enzyme tumor necrosis factor alpha-induced protein (TNFAIP)3 or A20, inhibits the inflammatory signaling transcription factor nuclear factor (NF)-kB leading to attenuation of the inflammatory response. A20-binding inhibitor of nuclear factor kB (ABIN-1), which physically interacts with A20, also plays a role in the inhibition of NF-kB and is poorly researched in the central nervous system (CNS). …

Clinical Performance Of The Point-Of-Care Cobas Liat For Detection Of Sars-Cov-2 In 20 Minutes: A Multicenter Study., Glen Hansen, Jamie Marino, Zi-Xuan Wang, Kathleen G Beavis, John Rodrigo, Kylie Labog, Lars F Westblade, Run Jin, Nedra Love, Karen Ding, Sachin Garg, Alan Huang, Joanna Sickler, Nam K Tran

Department of Pathology, Anatomy, and Cell Biology Faculty Papers

Highly accurate testing for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) at the point of care (POC) is an unmet diagnostic need in emergency care and time-sensitive outpatient care settings. Reverse transcription-PCR (RT-PCR) technology is the gold standard for SARS-CoV-2 diagnostics. We performed a multisite U.S. study comparing the clinical performance of the first U.S. Food and Drug Administration (FDA)-authorized POC RT-PCR for detection of SARS-CoV-2 in 20 min, the cobas Liat SARS-CoV-2 and influenza A/B nucleic acid test, to the most widely used RT-PCR laboratory test, the cobas 68/8800 SARS-CoV-2 test. Clinical nasopharyngeal swab specimens from 444 patients with …

'Educated' Osteoblasts Reduce Osteoclastogenesis In A Bone-Tumor Mimetic Microenvironment., Alexus D. Kolb, Jinlu Dai, Evan T. Keller, Karen M. Bussard

Department of Cancer Biology Faculty Papers

Breast cancer (BC) metastases to bone disrupt the balance between osteoblasts and osteoclasts, leading to excessive bone resorption. We identified a novel subpopulation of osteoblasts with tumor-inhibitory properties, called educated osteoblasts (EOs). Here we sought to examine the effect of EOs on osteoclastogenesis during tumor progression. We hypothesized that EOs affect osteoclast development in the bone-tumor niche, leading to suppressed pre-osteoclast fusion and bone resorption. Conditioned media (CM) was analyzed for protein expression of osteoclast factors receptor activator of nuclear factor kappa-β ligand (RANKL), osteoprotegerin (OPG), and tumor necrosis factor alpha (TNFα) via ELISA. EOs were co-cultured with pre-osteoclasts on …

Role Of Thymidine Phosphorylase In Type 2 Diabetes-Associated High-Risk Of Thrombosis, Adam Belcher

Theses, Dissertations and Capstones

Cardiovascular diseases (CVDs), such as ischemic heart disease and stroke, are the leading causes of death globally that disproportionally affects patients with type 2 diabetes mellitus (T2DM) at a 2-4-fold rate compared to non-diabetic patients. These CVDs result from a blood clot, also known as a thrombus, formed in arteries that can block blood flow resulting in tissue damage and death. Antiplatelet drugs, such as aspirin and clopidogrel, used to prevent thrombosis have been found to be less effective in diabetics and increase the risk of hemorrhagic events. The T2DM epidemic has made it imperative to find therapies that decrease …

Na/K-Atpase Alphα1 Regulates Adipogenesis Via Its Conserved Caveolin Binding Motif, Minqi Huang

Theses, Dissertations and Capstones

The Na/K-ATPase (NKA) was identified in 1957 by Dr. Jens C. Skou. It belongs to the P-type ATPase family, which can actively transport ions across cell membranes by using the energy from adenosine triphosphate (ATP) hydrolysis. During the second half of the 20th century, the molecular mechanism of the NKA catalytic cycle was clarified, and the isoform diversity of NKA in different species and organs was identified. The active ion transport through NKA generates cell membrane ion gradients and the electric potential. Hence, the enzymatic function of NKA is critical for cell viability as well as multiple physiological processes including …

The Epigenetic Effects Of Omega-3 Fatty Acids In Diffuse Large B-Cell Lymphoma, Tanner Jeffrey Bakhshi

Theses, Dissertations and Capstones

Diffuse large B-cell lymphoma (DLBCL) is the most common type of lymphoma. It is an aggressive cancer, with 50-70% of patients diagnosed at an advanced stage and 30-50% of patients not cured by chemoimmunotherapy. DLBCL is almost always caused by genetic damage sustained during the germinal center (GC) reaction. The mechanisms that govern the GC reaction bear a striking resemblance to those that drive DLBCL. Genomic studies have shown that some of its most common mutations occur in genes that encode epigenetic modifiers, including the lysine (histone) acetyltransferases CREBBP and p300. These mutations prevent the acetylation of multiple histone residues, …

Fgdb: Database Of Follicle Stimulating Hormone Glycans, Sushil Kumar Shakyawar, Sanjit Pandey, David J. Harvey, George Bousfield, Chittibabu Guda

Journal Articles: Genetics, Cell Biology & Anatomy

Glycomics, the study of the entire complement of sugars of an organism has received significant attention in the recent past due to the advances made in high throughput mass spectrometry technologies. These analytical advancements have facilitated the characterization of glycans associated with the follicle-stimulating hormones (FSH), which play a central role in the human reproductive system both in males and females utilizing regulating gonadal (testicular and ovarian) functions. The irregularities in FSH activity are also directly linked with osteoporosis. The glycoanalytical studies have been tremendously helpful in understanding the biological roles of FSH. Subsequently, the increasing number of characterized FSH …

Targeting Epigenetic Mechanisms In Endometriosis, Sarah Elizabeth Brunty

Theses, Dissertations and Capstones

Endometriosis is a complex and elusive gynecological disease in which the inner lining of the uterus grows in locations outside of the uterus and forms lesions. It is known to affect 1 in 9 women of reproductive age worldwide. Symptoms of endometriosis include severe pain, heavy periods, and infertility. While multiple theories of origin exist, none fully encompass all aspects of the disease, although all theories agree that this is an inflammation-driven disease. Due to this, many researchers are turning towards epigenetics to explain the initiation and progression of endometriosis. However, what is causing these epigenetic changes is still a …

Crippling Rapid Evolution Of Metastasis And Drug Resistance In A549 Non-Small Cell Lung Cancer Cells With The Clinically Relevant Hsp90 Inhibitor Auy922, Nickolas Anthony Bacon

Theses, Dissertations and Capstones

The ability for species to evolve new features in response to changing circumstances in order to survive and propagate is a ubiquitous observation on both the macroscopic and microscopic levels of living systems. It should be no surprise, then, that diseases such as cancer utilize their own forms of adaptation to perpetuate themselves when exposed to external threats. Indeed, concepts drawn from Darwinian evolution are now widely accepted to help explain certain aspects of carcinogenesis and malignant progression, the sum of which have come to be known as the theory of tumor evolution. Since metastasis and drug resistance are features …

Inhibiting Fatty Acid Binding Protein Family Members Decreases Multiple Myeloma Cell Proliferation Through Effecting The Myc Pathway, Mariah Farrell, Heather Fairfield, Anastasia D'Amico, Carolyne Falank, Connor Murphy, Michaela Reagan

Costas T. Lambrew Research Retreat 2021

FABP inhibition leads to increased survival of myeloma bearing mice and decreased cell growth by inhibiting MYC signaling.

The Effect Of High Fat Diet On Marrow Adipocytes From C57bl/6j (B6) Mice, Gisela Pachón-­Peña, Clifford J. Rosen, Samantha Costa

Costas T. Lambrew Research Retreat 2021

In mice models, the administration of a high fat diet (HFD) is an accelerating factor for metabolic syndrome, impaired glucose tolerance, and early type 2 diabetes mellitus (T2DM) (1)

Il-15/Il-15rα Heterodimeric Complex As Cancer Immunotherapy In Murine Breast Cancer Models, Siqi Guo, Ronald B. Smeltz, Anthony Nanajian, Richard Heller

Bioelectrics Publications

Interleukin 15 (IL-15) has been evaluated as a potential treatment for solid tumors in clinical trials, but the effectiveness of systemic IL-15 administration as a monotherapy has not been realized. IL-15 receptor alpha (IL-15Rα) can stabilize IL-15 and enhance its bioactivity. The goal of this study was to examine the activity of IL-15/IL-15Rα complex (IL-15cx) to CD8(+) T cells and evaluate its potential efficacy in murine breast cancer models. The antitumor efficacy was studied in mouse mammary carcinoma models (Her2/neu transgenic and 4T1-luc mammary cancers) treated with systemic recombinant protein with/without the depletion of myeloid-derived suppressor cells or intra-tumoral gene …

Cold Atmospheric-Pressure Plasma Caused Protein Damage In Methicillin-Resistant Staphylococcus Aureus Cells In Biofilms, Li Guo, Lu Yang, Yu Qi, Gulimire Niyazi, Lingling Huang, Lu Gou, Zifeng Wang, Lei Zhang, Dingxin Liu, Xiaohua Wang, Hailan Chen, Michael G. Kong

Bioelectrics Publications

Biofilms formed by multidrug-resistant bacteria are a major cause of hospital-acquired infections. Cold atmospheric-pressure plasma (CAP) is attractive for sterilization, especially to disrupt biofilms formed by multidrug-resistant bacteria. However, the underlying molecular mechanism is not clear. In this study, CAP effectively reduced the living cells in the biofilms formed by methicillin-resistant Staphylococcus aureus, and 6 min treatment with CAP reduced the S. aureus cells in biofilms by 3.5 log₁₀. The treatment with CAP caused the polymerization of SaFtsZ and SaClpP proteins in the S. aureus cells of the biofilms. In vitro analysis demonstrated that recombinant SaFtsZ lost …

Effect Of He Plasma Jet Versus Surface Plasma On The Metabolites Of Acute Myeloid Leukemia Cells, Dehui Xu, Ning Ning, Yujing Xu, Wenjie Xia, Hai-Lan Chen, Michael G. Kong

Bioelectrics Publications

Cold atmospheric plasma, including plasma jet and surface plasma, can promote the apoptosis of cancer cells without causing significant damage to surrounding normal cells, which was hopeful to be applied to the clinical cancer therapy. However, experimental plasma devices used directly to clinical experiments has challenges in technology and methods, especially the difference in killing tumor cells efficiency of these two common plasma sources. Therefore, it is great necessity to explore the differences in treating tumors between different plasma sources. This paper achieved good killing efficiency by using two kinds of cold atmospheric plasma generating devices, namely plasma jet and …

The Role Of Vascular Endothelial Growth Factor In Leukemia Trafficking, Shaw Powell

Theses and Dissertations--Medical Sciences

Vascular endothelial growth factor (VEGF) is a signaling protein involved in inducing and regulating endothelial cell proliferation and function (Duffy et al 2000). VEGF is also involved in cancer progression, as it induces vascular permeability and promotes angiogenesis to tumor laden areas, giving cancer cells critical oxygen and nutrients (Hoeppner et al.,2012. Studies indicate VEGF prevents lymphoblast apoptosis, which may contribute to leukemia formation and enable the proliferation of leukemic cells (Duffy et al 2000). Ongoing research seeks to further examine VEGF in leukemia, using a rag2:GFP-Myc expressing transgenic zebrafish as the animal model of T-cell Acute Lymphoblastic Leukemia (T-ALL). …

Novel Mammalian Models For Understanding And Treating Spinal Cord Injury, Michael B. Orr

Theses and Dissertations--Physiology

Spinal cord injury (SCI) is devastating and often leaves the injured individual with persistent dysfunction. The injury persists because humans have poor wound repair and there are no pharmacologic treatments to induce wound repair after SCI. The continued efforts to discover therapeutic targets and develop treatments heavily relies on animal models. The purpose of this project is to develop and study novel mammalian models of SCI to provide insights for the development and effective implementation of SCI therapies.

Lab mice (Mus musculus) are a powerful tool for recapitulating the progression and persistent damage evident in human SCI, but …

Systemic Versus Local Injection Of Bone Marrow Mesenchymal Stem Cells On 5-Fluorouracil Treated Parotid Glands Of Albino Rats, Caroline Maged Massieh, Medhat El Zainy, Reham Magdy Amin, Iman Fathy

Dentistry

ABSTRACT Objective: Head and neck cancer is the sixth leading cancer by incidence worldwide and eighth by death. Recent reports revealed that, not only radiotherapy but also chemotherapy may induce xerostomia. The aim of this study was to compare the possible regenerative effect of BMSCs through systemic and local injections. Material and Methods: 52 male Albino rats were randomly divided into 4 groups: Group 1: 10 rats received 0.5 ml of PBS by injection. Group 2: 14 rats received an intraperitoneal injection of 5-FU drug. Group 3: 14 rats were injected the same dose of 5-FU then received an intraglandular …

Protein-Functionalized Poly(Ethylene Glycol) Hydrogels As Scaffolds For Monolayer Organoid Culture, Reid L Wilson, Ganesh Swaminathan, Khalil Ettayebi, Carolyn Bomidi, Xi-Lei Zeng, Sarah E Blutt, Mary K Estes, K Jane Grande-Allen

Faculty, Staff and Students Publications

Stem cell-derived, organotypic in vitro models, known as organoids, have emerged as superior alternatives to traditional cell culture models due to their unparalleled ability to recreate complex physiological and pathophysiological processes. For this reason, they are attractive targets of tissue-engineering efforts, as constructs that include organoid technology would be expected to better simulate the many functions of the desired tissue or organ. While the 3D spheroidal architecture that is the default architecture of most organoid models may be preferred for some applications, 2D monolayer arrangements remain the preferred organization for many applications in tissue engineering. Therefore, in this work, we …

Epigenetic States Regulate Tumor Aggressiveness And Response To Targeted Therapies In Lung Adenocarcinoma, Fan Chen

Theses and Dissertations--Toxicology and Cancer Biology

Non-small cell lung cancer (NSCLC) is the leading cause of cancer-related death worldwide, 85% of which are lung adenocarcinomas (LUAD). Although molecular studies of NSCLC identified targetable mutations of some oncogenes including BRAF, EGFR and ALK, no targeted therapies exist for most cases. Cancer epigenetics is the study of epigenetic modifications, including histone modifications, that control gene expression in cancer. Recent advances have revealed numerous epigenetic abnormalities in human cancers, and targeting epigenetic regulators could be effective at reversing dysregulated epigenetic programs or driving sensitivity to other targeted therapies.

Inhibitors of the histone methyltransferase EZH2 have recently been approved as …

Janus Kinase 1 Drives Endoplasmic Reticulum Stress-Induced Transcriptional Reprogramming In Astrocytes, Savannah Graham Sims

Graduate Theses, Dissertations, and Problem Reports (ETD)

Neurological and neurodegenerative diseases are heterogenous and devastating diseases with limited therapeutic options and no cures. The broad, long-term goal of this project was to elucidate therapeutic targets for neurodegenerative conditions that attenuate damaging inflammation while leaving the beneficial immune response intact and avoiding broad immunosuppression. Inflammation and the accumulation of misfolded proteins are associated with a wide variety of neurological diseases. Here, we have examined how the accumulation of misfolded proteins shapes inflammatory signaling in the glial cell population astrocytes. Astrocytes are the most populous cell in the central nervous system (CNS) and provide physical and trophic support to …

Extracellular Vesicles Released By Human Retinal Pigment Epithelium Mediate Increased Polarised Secretion Of Drusen Proteins In Response To Amd Stressors, Miguel Flores-Bellver, Jason Mighty, Silvia Aparicio-Domingo, Kang V. Li, Cui Shi, Jing Zhou, Hannah Cobb, Patrick Mcgrath, German Michelis, Patricia Lenhart, Ganna Bilousova, Søren Heissel, Michael J. Rudy, Christina Coughlan, Andrew E. Goodspeed, S. Patricia Becerra, Stephen Redenti, M. Valeria Canto-Soler

Publications and Research

Age-related macular degeneration (AMD) is a leading cause of blindness worldwide. Drusen are key contributors to the etiology of AMD and the ability to modulate drusen biogenesis could lead to therapeutic strategies to slow or halt AMD progression. The mechanisms underlying drusen biogenesis, however, remain mostly unknown. Here we demonstrate that under homeostatic conditions extracellular vesicles (EVs) secreted by retinal pigment epithelium (RPE) cells are enriched in proteins associated with mechanisms involved in AMD pathophysiology, including oxidative stress, immune response, inflammation, complement system and drusen composition. Furthermore, we provide first evidence that drusen-associated proteins are released as cargo of extracellular …

Human Retinal Organoids Release Extracellular Vesicles That Regulate Gene Expression In Target Human Retinal Progenitor Cells, Jing Zhou, Miguel Flores‑Bellver, Jianbo Pan, Alberto Benito‑Martin, Cui Shi, Onyekwere Onwumere, Jason Mighty, Jiang Qian, Xiufeng Zhong, Tasmim Hogue, Baffour Amponsah‑Antwi, Linda Einbond, Rajendra Gharbaran, Hao Wu, Bo‑Juen Chen, Zhiliang Zheng, Tatyana Tchaikovskaya, Xusheng Zhang, Hector Peinado, Maria Valeria Canto‑Soler, Stephen Redenti

Publications and Research

The mechanisms underlying retinal development have not been completely elucidated. Extracellular vesicles (EVs) are novel essential mediators of cell‑to‑cell communication with emerging roles in developmental processes. Nevertheless, the identification of EVs in human retinal tissue, characterization of their cargo, and analysis of their potential role in retina development has not been accomplished. Three‑dimensional retinal tissue derived from human induced pluripotent stem cells (hiPSC) provide an ideal developmental system to achieve this goal. Here we report that hiPSC‑derived retinal organoids release exosomes and microvesicles with small noncoding RNA cargo. EV miRNA cargo‑predicted targetome correlates with Gene Ontology (GO) pathways involved in …

Plasma-Treated Solutions (Pts) In Cancer Therapy, Hiromasa Tanaka, Sander Bekeschus, Dayun Yan, Masaru Hori, Michael Keidar, Mounir Laroussi

Electrical & Computer Engineering Faculty Publications

Cold physical plasma is a partially ionized gas generating various reactive oxygen and nitrogen species (ROS/RNS) simultaneously. ROS/RNS have therapeutic effects when applied to cells and tissues either directly from the plasma or via exposure to solutions that have been treated beforehand using plasma processes. This review addresses the challenges and opportunities of plasma-treated solutions (PTSs) for cancer treatment. These PTSs include plasma-treated cell culture media in experimental research as well as clinically approved solutions such as saline and Ringer’s lactate, which, in principle, already qualify for testing in therapeutic settings. Several types of cancers were found to succumb to …

Conservation Of Epithelial-To-Mesenchymal Transition Process In Neural Crest Cells And Metastatic Cancer, April Zhang, Hira Aslam, Neha Sharma, Aryeh Warmflash, Walid D Fakhouri

Faculty, Staff and Student Publications

Epithelial to mesenchymal transition (EMT) is a highly conserved cellular process in several species, from worms to humans. EMT plays a fundamental role in early embryogenesis, wound healing, and cancer metastasis. For neural crest cell (NCC) development, EMT typically results in forming a migratory and potent cell population that generates a wide variety of cell and tissue, including cartilage, bone, connective tissue, endocrine cells, neurons, and glia amongst many others. The degree of conservation between the signaling pathways that regulate EMT during development and metastatic cancer (MC) has not been fully established, despite ample studies. This systematic review and meta-analysis …

Hippo-Yap Pathway Orchestrates Neural Crest Ontogenesis, Xiaolei Zhao, Tram P Le, Shannon Erhardt, Tina O Findley, Jun Wang

Faculty, Staff and Student Publications

Neural crest (NC) cells are a migratory stem cell population in vertebrate embryogenesis that can give rise to multiple cell types, including osteoblasts, chondrocytes, smooth muscle cells, neurons, glia, and melanocytes, greatly contributing to the development of different tissues and organs. Defects in NC development are implicated in many human diseases, such as numerous syndromes, craniofacial aberration and congenital heart defects. Research on NC development has gained intense interest and made significant progress. Recent studies showed that the Hippo-Yap pathway, a conserved fundamental pathway with key roles in regulation of cell proliferation, survival, and differentiation, is indispensable for normal NC …

Endmt Regulation By Small Rnas In Diabetes-Associated Fibrotic Conditions: Potential Link With Oxidative Stress, Roberta Giordo, Yusra M A Ahmed, Hilda Allam, Salah Abusnana, Lucia Pappalardo, Gheyath K Nasrallah, Arduino Aleksander Mangoni, Gianfranco Pintus

Faculty, Staff and Student Publications

Diabetes-associated complications, such as retinopathy, nephropathy, cardiomyopathy, and atherosclerosis, the main consequences of long-term hyperglycemia, often lead to organ dysfunction, disability, and increased mortality. A common denominator of these complications is the myofibroblast-driven excessive deposition of extracellular matrix proteins. Although fibroblast appears to be the primary source of myofibroblasts, other cells, including endothelial cells, can generate myofibroblasts through a process known as endothelial to mesenchymal transition (EndMT). During EndMT, endothelial cells lose their typical phenotype to acquire mesenchymal features, characterized by the development of invasive and migratory abilities as well as the expression of typical mesenchymal products such as α-smooth …

Enhancing Glucose Metabolism Via Gluconeogenesis Is Therapeutic In A Zebrafish Model Of Dravet Syndrome, Rajeswari Banerji, Christopher Huynh, Francisco Figueroa, Matthew T Dinday, Scott C Baraban, Manisha Patel

Faculty, Staff and Students Publications

Energy-producing pathways are novel therapeutic targets for the treatment of neurodevelopmental disorders. Here, we focussed on correcting metabolic defects in a catastrophic paediatric epilepsy, Dravet syndrome which is caused by mutations in sodium channel NaV1.1 gene, SCN1A. We utilized a translatable zebrafish model of Dravet syndrome (scn1lab) which exhibits key characteristics of patients with Dravet syndrome and shows metabolic deficits accompanied by down-regulation of gluconeogenesis genes, pck1 and pck2. Using a metabolism-based small library screen, we identified compounds that increased gluconeogenesis via up-regulation of pck1 gene expression in scn1lab larvae. Treatment with PK11195, a pck1 activator …

The E3 Ligase Traf4 Promotes Igf Signaling By Mediating Atypical Ubiquitination Of Irs-1, Wenjuan Yu, Ramesh Singh, Zhao Wang, Bert W O'Malley, Ping Yi

Faculty, Staff and Students Publications

Insulin-like growth factor (IGF) is a potent mitogen that activates the IGF receptor (IGFR)/insulin receptor substrate (IRS) axis, thus stimulating growth in normal cells and uncontrolled cell proliferation in cancer. Posttranslational modifications of IRS such as ubiquitination tightly control IGF signaling, and we previously identified IRS-1 as a potential substrate for the E3 ubiquitin ligase TRAF4 using an unbiased screen. Here we provide evidence that TRAF4-mediated ubiquitination of IRS-1 is physiologically relevant and crucial for IGF signal transduction. Through site-directed mutagenesis we found that TRAF4 promotes an atypical K29-linked ubiquitination at the C-terminal end of IRS-1. Its depletion abolishes AKT …