Molecular Biology Commons^™

Correlation Between Salivary Levels And Tissue Expression Status Of Her2 In Breast Cancer Patients - A Cross-Sectional Study, Arthi Sri A.S, Massillamani F, Vinothkumar Thilla Sekar

Annual Research Symposium

The study aims to investigate if soluble salivary human epidermal growth factor receptor-2 (HER2) would play a role in the diagnosis of breast cancer patients and also to find out the influence of demographic and hormonal factors on the salivary HER2 levels in breast cancer.

Cold-Inducible Rna Binding Protein Impedes Breast Tumor Growth In The Pymt Murine Model For Breast Cancer, Daniel A. Lujan, Joey L. Ochoa, Ellen J. Beswick, Tamara A. Howard, Helen J. Hathaway, Nora I. Perrone-Bizzozero, Rebecca S. Hartley

Markey Cancer Center Faculty Publications

RNA binding proteins (RBPs) post-transcriptionally regulate gene expression by associating with regulatory sequences in the untranslated regions of mRNAs. Cold-inducible RBP (CIRP) is a stress-induced RBP that was recently shown to modulate inflammation in response to cellular stress, where it increases or decreases pro-tumorigenic (proinflammatory) cytokines in different contexts. CIRP expression is altered in several cancers, including breast cancer, but the effects of CIRP on inflammation in breast cancer is not known. Here, we investigate if CIRP alters growth and the inflammatory profile of breast tumors. Transgenic mice overexpressing CIRP in the mammary epithelium were crossed with the PyMT mouse …

Inflammation And Tumor Progression: The Differential Impact Of Saa In Breast Cancer Models, Daniel Wilhelm Olivier, Carla Eksteen, Manisha Du Plessis, Louis De Jager, Lize Engelbrecht, Nathaniel Wade Mcgregor, Preetha Shridas, Frederick C. De Beer, Willem J. S. De Villiers, Etheresia Pretorius, Anna-Mart Engelbrecht

Saha Cardiovascular Research Center Faculty Publications

Background: Previous research has shown that the Serum Amyloid A (SAA) protein family is intricately involved in inflammatory signaling and various disease pathologies. We have previously demonstrated that SAA is associated with increased colitis disease severity and the promotion of tumorigenesis. However, the specific role of SAA proteins in breast cancer pathology remains unclear. Therefore, we investigated the role of systemic SAA1 and SAA2 (SAA1/2) in a triple-negative breast cancer mouse model.

Methods: Syngeneic breast tumors were established in wild-type mice, and mice lacking the SAA1/2 (SAADKO). Subsequently, tumor volume was monitored, species survival determined, the inflammatory profiles of mice …

The Carboxy Terminus Causes Interfacial Assembly Of Oleate Hydratase On A Membrane Bilayer, Christopher D. Radka, Christy R. Grace, Hale S. Hasdemir, Yupeng Li, Carlos C. Rodriguez, Patrick Rodrigues, Michael L. Oldham, M. Zuhaib Qayyum, Aaron Pitre, William J. Maccain, Ravi C. Kalathur, Emad Tajkhorshid, Charles O. Rock

Markey Cancer Center Faculty Publications

The soluble flavoprotein oleate hydratase (OhyA) hydrates the 9-cis double bond of unsaturated fatty acids. OhyA sub- strates are embedded in membrane bilayers; OhyA must remove the fatty acid from the bilayer and enclose it in the active site. Here, we show that the positively charged helix- turn-helix motif in the carboxy terminus (CTD) is responsible for interacting with the negatively charged phosphatidylgly- cerol (PG) bilayer. Super-resolution microscopy of Staphylo- coccus aureus cells expressing green fluorescent protein fused to OhyA or the CTD sequence shows subcellular localization along the cellular boundary, indicating OhyA is membrane- associated and the CTD sequence …

Strategies For Improving The Performance Of Prediction Models For Response To Immune Checkpoint Blockade Therapy In Cancer, Tiantian Zeng, Jason Z. Zhang, Arnold Stromberg, Jin Chen, Chi Wang

Markey Cancer Center Faculty Publications

Immune checkpoint blockade (ICB) therapy holds promise for bringing long-lasting clinical gains for the treatment of cancer. However, studies show that only a fraction of patients respond to the treatment. In this regard, it is valu- able to develop gene expression signatures based on RNA sequencing (RNAseq) data and machine learning meth- ods to predict a patient’s response to the ICB therapy, which contributes to more personalized treatment strategy and better management of cancer patients. However, due to the limited sample size of ICB trials with RNAseq data available and the vast number of candidate gene expression features, it is …

Vaginal Lactobacillus Fatty Acid Response Mechanisms Reveal A Metabolite-Targeted Strategy For Bacterial Vaginosis Treatment, Paul C. Blainey, Seth M. Bloom, Douglas S. Kwon

Markey Cancer Center Faculty Publications

Bacterial vaginosis (BV), a common syndrome characterized by Lactobacillus-deficient vaginal microbiota, is associated with adverse health outcomes. BV often recurs after standard antibiotic therapy in part because antibiotics promote microbiota dominance by Lactobacillus iners instead of Lactobacillus crispatus, which has more beneficial health associations. Strategies to promote L. crispatus and inhibit L. iners are thus needed. We show that oleic acid (OA) and similar long-chain fatty acids simultaneously inhibit L. iners and enhance L. crispatus growth. These phenotypes require OA-inducible genes conserved in L. crispatus and related lactobacilli, including an oleate hydratase (ohyA) and putative fatty acid efflux pump (farE). …

Elevated Free Cholesterol Levels Due To Impaired Reverse Cholesterol Transport Are A Risk Factor For Polymicrobial Sepsis In Mice, Qian Wang, Ling Guo, Dan Hao, Misa Ito, Chieko Mineo, Philip W. Shaul, Xiang-An Li

Markey Cancer Center Faculty Publications

Dysregulated lipid metabolism is commonly observed in septic patients, but how it contributes to sepsis remains largely unknown. Reverse cholesterol transport (RCT) is crucial for regulating cholesterol metabolism in circulation. During RCT, high-density lipoprotein collects cholesterol from peripheral tissues and transports it to the liver’s scavenger receptor BI (SR- BI), where SR-BI mediates the uptake of cholesteryl esters (CEs) from high-density lipoprotein for excretion via bile. In this study, we utilized AlbCreSR-BIfl/fl mice, a model with impaired RCT, to investigate the impact of RCT on sepsis. We found that AlbCreSR-BIfl/fl mice were significantly more susceptible to cecal ligation and puncture …

Therapeutic Potential Of Berberine In Attenuating Cholestatic Liver Injury: Insights From A Psc Mouse Model, Yanyan Wang, Derrick Zhao, Lianyong Su, Yun-Ling Tai, Grayson W. Way, Jing Zeng, Qianhua Yan, Ying Xu, Xuan Wang, Emily C. Gurley, Xi-Qiao Zhou, Jinze Liu, Jinpeng Liu, Weidong Chen, Philip B. Hylemon, Huiping Zhou

Markey Cancer Center Faculty Publications

Background and aims

Primary sclerosing cholangitis (PSC) is a chronic liver disease characterized by progressive biliary inflammation and bile duct injury. Berberine (BBR) is a bioactive isoquinoline alkaloid found in various herbs and has multiple beneficial effects on metabolic and inflammatory diseases, including liver diseases. This study aimed to examine the therapeutic effect of BBR on cholestatic liver injury in a PSC mouse model (Mdr2 −/− mice) and eluci‑ date the underlying mechanisms.

Methods

Mdr2−/− mice (12–14 weeks old, both sexes) received either BBR (50 mg/kg) or control solution daily for eight weeks via oral gavage. Histological and serum biochemical …

Mechanisms Of Γδ T Cell Accumulation In Visceral Adipose Tissue With Aging, Sujata Mukherjee, Maria E. C. Bruno, Jason Oakes, Gregory S. Hawk, Arnold Stromberg, Donald A. Cohen, Marlene E. Starr

Markey Cancer Center Faculty Publications

γδ T cells are resident in visceral adipose tissue (VAT) where they show an age- associated increase in numbers and contribute to local and systemic chronic inflammation. However, regulation of this population and mechanisms for the age-dependent accumulation are not known. In this study, we identified a progressive trend of γδ T cell accumulation in VAT over the lifespan in mice and explored physiological mechanisms contributing to accumulation. Using isochronic parabiotic pairs of wild-type (WT) and T cell receptor delta knockout (TCRδ KO) mice at young and old age, we confirmed that VAT γδ T cells are predominately a tissue-resident …

A Novel Protozoa Parasite-Derived Protein Adjuvant Is Effective In Immunization With Cancer Cells To Activate The Cancer-Specific Protective Immunity And Inhibit The Cancer Growth In A Murine Model Of Colorectal Cancer, Rajesh Mani, Chloe G. Martin, Kanal E. Balu, Qingding Wang, Piotr G. Rychahou, Tadahide Izumi, B. Mark Evers, Yasuhiro Suzuki

Markey Cancer Center Faculty Publications

Cancer-specific CD8+ cytotoxic T cells play important roles in preventing cancer growth, and IFN-γ, in addition to IL-12 and type I interferon, is critical for activating CD8+ cytotoxic T cells. We recently identified the capability of the amino-terminus region of dense granule protein 6 (GRA6Nt) of Toxoplasma gondii, an intracellular protozoan parasite, to activate IFN-γ production of microglia, a tissue-resident macrophage population. Therefore, in the present study, we examined whether recombinant GRA6Nt protein (rGRA6Nt) functions as an effective adjuvant to potently activate cancer-specific protective immunity using a murine model of MC38 colorectal cancer (CRC). When mice were immunized with non-replicable …

Structure, Dynamics, And Redox Reactivity Of An All-Purpose Flavodoxin, Sharique Khan, Ahmadullah Ansari, Monica Brachi, Debarati Das, Wassim El Housseini, Shelley Minteer, Anne-Francis Miller

Markey Cancer Center Faculty Publications

The flavodoxin of Rhodopseudomonas palustris CGA009 (Rp9Fld) supplies highly reducing equivalents to crucial en- zymes such as hydrogenase, especially when the organism is iron-restricted. By acquiring those electrons from photodriven electron flow via the bifurcating electron transfer flavoprotein, Rp9Fld provides solar power to vital metabolic processes. To understand Rp9Fld’s ability to work with diverse partners, we solved its crystal structure. We observed the canonical fla- vodoxin (Fld) fold and features common to other long-chain Flds but not all the surface loops thought to recognize part- ner proteins. Moreover, some of the loops display alternative structures and dynamics. To advance studies …

Non-Transcriptional Irf7 Interacts With Nf-Κb To Inhibit Viral Inflammation, Shumin Fan, Sonam Popli, Sukanya Chakravarty, Ritu Chakravarty, Saurabh Chattopadhyay

Markey Cancer Center Faculty Publications

Interferon (IFN) regulatory factors (IRF) are key transcrip- tion factors in cellular antiviral responses. IRF7, a virus- inducible IRF, expressed primarily in myeloid cells, is required for transcriptional induction of interferon α and antiviral genes. IRF7 is activated by virus-induced phosphory- lation in the cytoplasm, leading to its translocation to the nu- cleus for transcriptional activity. Here, we revealed a nontranscriptional activity of IRF7 contributing to its antiviral functions. IRF7 interacted with the pro-inflammatory tran- scription factor NF-κB-p65 and inhibited the induction of in- flammatory target genes. Using knockdown, knockout, and overexpression strategies, we demonstrated that IRF7 inhibited NF-κB–dependent inflammatory …

Foxs1 Is Increased In Liver Fibrosis And Regulates Tgfβ Responsiveness And Proliferation Pathways In Human Hepatic Stellate Cells, Evelyn A. Bates, Zachary A. Kipp, Wang-Hsin Lee, Genesee J. Martinez, Lauren Weaver, Kathryn N. Becker, Sally Pauss, Justin F. Creeden, Garrett B. Anspach, Robert N. Helsley, Mei Xu, Maria E. C. Bruno, Marlene E. Starr, Terry D. Hinds Jr.

Markey Cancer Center Faculty Publications

Liver fibrosis commences with liver injury stimulating transforming growth factor beta (TGFβ) activation of hepatic stellate cells (HSCs), causing scarring and irreversible damage. TGFβ induces expression of the transcription factor Forkhead box S1 (FOXS1) in hepatocytes and may have a role in the pathogenesis of hepatocellular carcinoma (HCC). To date, no studies have determined how it affects HSCs. We analyzed human livers with cirrhosis, HCC, and a murine fibrosis model and found that FOXS1 expression is significantly higher in fibrotic livers but not in HCC. Next, we treated human LX2 HSC cells with TGFβ to activate fibrotic pathways, and FOXS1 …

Evaluating The Efficacy Of The Archaeal Proteasome To Alleviate Protein-Misfolding Retinopathy, Celine Brooks

Graduate Theses, Dissertations, and Problem Reports (ETD)

Nearly all neurodegenerative diseases (NDs) and some eye diseases are characterized by the presence of misfolded and aggregated proteins. The ubiquitin proteasome system (UPS) is responsible for degrading the majority of proteins including misfolded and aggregated proteins. However, in NDs the performance of the UPS is impaired, and this impairment partially comes from these protein aggregates inhibiting the proteasome. This inhibition leads to an accumulation of protein aggregates, more proteasomal inhibition, and fewer chaperones for newly synthesized proteins, triggering a collapse in proteostasis, which is thought to contribute to the pathogenesis of NDs. Consequently, stimulating proteasome function is being investigated …

A Novel Role Of Tgfbi In Macrophage Polarization And Macrophage-Induced Pancreatic Cancer Growth And Therapeutic Resistance, Jing Zhou, Nan Lyu, Qiongling Wanga, Ming Yang, Eric T. Kimchi, Kun Chenge, Trupti Joshi, Adama R. Tukulif, Kevin F. Staveley-O’Carroll, Guangfu Li

Biomedical Sciences

Tumor-associated macrophages (TAMs), as a major and essential component of tumor microenvironment (TME), play a critical role in orchestrating pancreatic cancer (PaC) tumorigenesis from initiation to angiogenesis, growth, and systemic dissemination, as well as immunosuppression and resistance to chemotherapy and immunotherapy; however, the critical intrinsic factors responsible for TAMs reprograming and function remain to be identified. By performing single-cell RNA sequencing, transforming growth factor-beta-induced protein (TGFBI) was identified as TAM-producing factor in murine PaC tumors. TAMs express TGFBI in human PaC and TGFBI expression is positively related with human PaC growth. By inducing TGFBI loss-of-function in macrophage (MΦs) in vitro …

Cell Aggregation Activates Small Gtpase Rac1 And Induces Cd44 Cleavage By Maintaining Lipid Raft Integrity, Dong Li, Younhee Park, Hami Hemati, Xia Liu

Markey Cancer Center Faculty Publications

Lipid rafts are highly ordered membrane domains that are enriched in cholesterol and glycosphingolipids and serve as major platforms for signal transduction. Cell detachment from the extracellular matrix (ECM) triggers lipid raft disruption and anoikis, which is a barrier for cancer cells to metastasize. Compared to single circulating tumor cells (CTCs), our recent studies have demonstrated that CD44-mediatd cell aggregation enhances the stemness, survival and metastatic ability of aggregated cells. Here, we investigated whether and how lipid rafts are involved in CD44-mediated cell aggregation. We found that cell detachment, which mimics the condition when tumor cells detach from the ECM …

Deficiency Of Acute-Phase Serum Amyloid A Exacerbates Sepsis-Induced Mortality And Lung Injury In Mice, Ailing Ji, Andrea C. Trumbauer, Victoria P. Noffsinger, Luke W. Meredith, Brittany Dong, Qian Wang, Ling Guo, Xiangan Li, Frederick C. De Beer, Nancy R. Webb, Lisa R. Tannock, Marlene E. Starr, Christopher M. Waters, Preetha Shridas

Saha Cardiovascular Research Center Faculty Publications

Serum amyloid A (SAA) is a family of proteins, the plasma levels of which may increase >1000-fold in acute inflammatory states. We investigated the role of SAA in sepsis using mice deficient in all three acute-phase SAA isoforms (SAA-TKO). SAA deficiency significantly increased mortality rates in the three experimental sepsis mouse models: cecal ligation and puncture (CLP), cecal slurry (CS) injection, and lipopolysaccharide (LPS) treatments. SAA-TKO mice had exacerbated lung pathology compared to wild-type (WT) mice after CLP. A bulk RNA sequencing performed on lung tissues excised 24 h after CLP indicated significant enrichment in the expression of genes associated …

Loss Of Carnitine Palmitoyltransferase 1a Reduces Docosahexaenoic Acid-Containing Phospholipids And Drives Sexually Dimorphic Liver Disease In Mice, Mikala M. Zelows, Corissa Cady, Nikitha Dharanipragada, Anna E. Mead, Zachary A. Kipp, Evelyn A. Bates, Venkateshwari Varadharajan, Rakhee Banerjee, Se-Hyung Park, Nathan R. Shelman, Harrison A. Clarke, Tara R. Hawkinson, Terrymar Medina, Ramon C. Sun, Todd A. Lydic, Terry D. Hinds, Jr., J. Mark Brown, Samir Softic, Gregory A. Graf, Robert N. Helsley

Saha Cardiovascular Research Center Faculty Publications

Background and aims: Genome and epigenome wide association studies identified variants in carnitine palmitoyltransferase 1a (CPT1a) that associate with lipid traits. The goal of this study was to determine the role of liver-specific CPT1a on hepatic lipid metabolism.

Approach and results: Male and female liver-specific knockout (LKO) and littermate controls were placed on a low-fat or high-fat diet (60% kcal fat) for 15 weeks. Mice were necropsied after a 16 h fast, and tissues were collected for lipidomics, matrix-assisted laser desorption ionization mass spectrometry imaging, kinome analysis, RNA-sequencing, and protein expression by immunoblotting. Female LKO mice had increased serum alanine …

Cyclin D1 Extensively Reprograms Metabolism To Support Biosynthetic Pathways In Hepatocytes, Heng Wu, Betsy T. Kren, Andrew N. Lane, Teresa A. Cassel, Richard M. Higashi, Teresa Whei-Mei Fan, George S. Scaria, Laurie L. Shekels, Mark A. Klein, Jeffrey H. Albrecht

Markey Cancer Center Faculty Publications

Cell proliferation requires metabolic reprogramming to accommodate biosynthesis of new cell components, and similar alterations occur in cancer cells. However, the mechanisms linking the cell cycle machinery to metabolism are not well defined. Cyclin D1, along with its main partner cyclindependent kinase 4 (Cdk4), is a pivotal cell cycle regulator and driver oncogene that is overexpressed in many cancers. Here, we examine hepatocyte proliferation to define novel effects of cyclin D1 on biosynthetic metabolism. Metabolomic studies reveal that cyclin D1 broadly promotes biosynthetic pathways including glycolysis, the pentose phosphate pathway, and the purine and pyrimidine nucleotide synthesis in hepatocytes. Proteomic …

Md_Harmonize: A Python Package For Atom-Level Harmonization Of Public Metabolic Databases, Huan Jin, Hunter N. B. Moseley

Markey Cancer Center Faculty Publications

A major challenge to integrating public metabolic resources is the use of different nomenclatures by individual databases. This paper presents md_harmonize, an open-source Python package for harmonizing compounds and metabolic reactions across various metabolic databases. The md_harmonize package utilizes a neighborhood-specific graph coloring method for generating a unique identifier for each compound via atom identifiers based on a compound’s chemical structure. The resulting harmonized compounds and reactions can be used for various downstream analyses, including the construction of atom-resolved metabolic networks and models for metabolic flux analysis. Parts of the md_harmonize package have been optimized using a variety of computational …

Design And Characterization Of A Novel Eef2k Degrader With Potent Therapeutic Efficacy Against Triple-Negative Breast Cancer, Changxin Zhong, Rongfeng Zhu, Ting Jiang, Sheng Tian, Xiaobao Zhao, Xiaoya Wan, Shilong Jiang, Zonglin Chen, Rong Gong, Linhao He, Jin-Ming Yang, Na Ye, Yan Cheng

Markey Cancer Center Faculty Publications

Dysregulated eEF2K expression is implicated in the pathogenesis of many human cancers, including triple-negative breast cancer (TNBC), making it a plausible therapeutic target. However, specific eEF2K inhibitors with potent anti-cancer activity have not been available so far. Targeted protein degradation has emerged as a new strategy for drug discovery. In this study, a novel small molecule chemical is designed and synthesized, named as compound C1, which shows potent activity in degrading eEF2K. C1 selectively binds to F8, L10, R144, C146, E229, and Y236 of the eEF2K protein and promotes its proteasomal degradation by increasing the interaction between eEF2K and the …

Benchmark Dataset For Training Machine Learning Models To Predict The Pathway Involvement Of Metabolites, Erik D. Huckvale, Christian D. Powell, Huan Jin, Hunter N. B. Moseley

Markey Cancer Center Faculty Publications

Metabolic pathways are a human-defined grouping of life sustaining biochemical reactions, metabolites being both the reactants and products of these reactions. But many public datasets include identified metabolites whose pathway involvement is unknown, hindering metabolic interpretation. To address these shortcomings, various machine learning models, including those trained on data from the Kyoto Encyclopedia of Genes and Genomes (KEGG), have been developed to predict the pathway involvement of metabolites based on their chemical descriptions; however, these prior models are based on old metabolite KEGG-based datasets, including one benchmark dataset that is invalid due to the presence of over 1500 duplicate entries. …

Human Schwann Cells In Vitro I. Nerve Tissue Processing, Pre-Degeneration, Isolation, And Culturing Of Primary Cells, Gabriela I. Aparicio, Paula V. Monje

Markey Cancer Center Faculty Publications

This paper presents versatile protocols to prepare primary human Schwann cell (hSC) cultures from mature peripheral nervous system tissues, including fascicles from long spinal nerves, nerve roots, and ganglia. This protocol starts with a description of nerve tissue procurement, handling, and dissection to obtain tissue sections suitable for hSC isolation and culturing. A description follows on how to disintegrate the nerve tissue by delayed enzymatic dissociation, plate the initial cell suspensions on a two-dimensional substrate, and culture the primary hSCs. Each section contains detailed procedures, technical notes, and background information to aid investigators in understanding and managing all steps. Some …

Phosphorylation Of Ahr By Plk1 Promotes Metastasis Of Luad Via Dio2-Th Signaling, Chaohao Li, Derek B. Allison, Daheng He, Fengyi Mao, Xinyi Wang, Piotr G. Rychahou, Ibrahim A. Imam, Yifan Kong, Qiongsi Zhang, Yanquan Zhang, Jinghui Liu, Ruixin Wang, Xiongjian Rao, Sai Wu, B. Mark Evers, Qing Shao, Chi Wang, Zhiguo Li, Xiaoqi Liu

Markey Cancer Center Faculty Publications

Metastasis of lung adenocarcinoma (LUAD) is a major cause of death in patients. Aryl hydrocarbon receptor (AHR), an important transcription factor, is involved in the initiation and progression of lung cancer. Polo-like kinase 1 (PLK1), a serine/threonine kinase, acts as an oncogene promoting the malignancy of multiple cancer types. However, the interaction between these two factors and their significance in lung cancer remain to be determined. In this study, we demonstrate that PLK1 phosphorylates AHR at S489 in LUAD, leading to epithelial-mesenchymal transition (EMT) and metastatic events. RNA-seq analyses reveal that type 2 deiodinase (DIO2) is responsible for EMT and …

Copy Number Variation That Influences The Ionizing Radiation Sensitivity Of Oral Squamous Cell Carcinoma, Tadahide Izumi, Piotr G. Rychahou, Li Chen, Molly H. Smith, Joseph Valentino

Markey Cancer Center Faculty Publications

Genome instability in cancer cells causes not only point mutations but also structural variations of the genome, including copy number variations (CNVs). It has recently been proposed that CNVs arise in cancer to adapt to a given microenvironment to survive. However, how CNV influences cellular resistance against ionizing radiation remains unknown. PRMT5 (protein arginine methyltransferase 5) and APE1 (apurinic/apyrimidinic endonuclease 1), which enhance repair of DNA double-strand breaks and oxidative DNA damage, are closely localized in the chromosome 14 of the human genome. In this study, the genomics data for the PRMT5 and APE1 genes, including their expression, CNVs, and …

Distinct Camp Signaling Microdomains Differentially Regulate Melanosomal Ph And Pigmentation, Maftuna Yusupova, Dalee Zhou, Jaewon You, Jeydi Gonzalez-Guzman, Megha B. Ghanta, Hong Pu, Zalfa Abdel-Malek, Qiuying Chen, Steven S. Gross, John August D'Orazio, Shosuke Ito, Kazumasa Wakamatsu, Melissa L. Harris, Jonathan H. Zippin

Markey Cancer Center Faculty Publications

cAMP signaling is a well-established regulator of melanin synthesis. Two distinct cAMP signaling pathways—the transmembrane adenylyl cyclase pathway, activated primarily by the MC1R, and the soluble adenylyl cyclase (sAC) pathway—affect melanin synthesis. The sAC pathway affects melanin synthesis by regulating melanosomal pH, and the MC1R pathway affects melanin synthesis by regulating gene expression and post-translational modifications. However, whether MC1R genotype affects melanosomal pH is poorly understood. We now report that loss of function MC1R does not affect melanosomal pH. Thus, sAC signaling appears to be the only cAMP signaling pathway that regulates melanosomal pH. We also addressed whether MC1R genotype …

Phosphatidic Acid Binding To Patched Contributes To The Inhibition Of Smoothened And Hedgehog Signaling In Drosophila Wing Development, Jie Zhang, Yajuan Liu, Chi Wang, Craig W. Vander Kooi, Jianhang Jia

Markey Cancer Center Faculty Publications

Hedgehog (Hh) signaling controls growth and patterning during embryonic development and homeostasis in adult tissues. Hh binding to the receptor Patched (Ptc) elicits intracellular signaling by relieving Ptc-mediated inhibition of the transmembrane protein Smoothened (Smo). We uncovered a role for the lipid phosphatidic acid (PA) in the regulation of the Hh pathway in Drosophila melanogaster. Deleting the Ptc C-terminal tail or mutating the predicted PA-binding sites within it prevented Ptc from inhibiting Smo in wing discs and in cultured cells. The C-terminal tail of Ptc directly interacted with PA in vitro, an association that was reduced by Hh, and increased …

Hiv-1 Vpu Protein Forms Stable Oligomers In Aqueous Solution Via Its Transmembrane Domain Self-Association, Saman Majeed, Lan Dang, Md Majharul Islam, Olamide Ishola, Peter P Borbat, Steven J Ludtke, Elka R Georgieva

Faculty, Staff and Students Publications

We report our findings on the assembly of the HIV-1 protein Vpu into soluble oligomers. Vpu is a key HIV-1 protein. It has been considered exclusively a single-pass membrane protein. Previous observations show that this protein forms stable oligomers in aqueous solution, but details about these oligomers still remain obscure. This is an interesting and rather unique observation, as the number of proteins transitioning between soluble and membrane embedded states is limited. In this study we made use of protein engineering, size exclusion chromatography, cryoEM and electron paramagnetic resonance (EPR) spectroscopy to better elucidate the nature of the soluble oligomers. …

Protocol For Optimizing Production And Quality Control Of Infective Ecohiv Virions, Hammodah Rawhi Hammodah Alfar, Daphne N. Pariser, Harry Chanzu, Smita Joshi, Daniёlle M. Coenen, Joshua Lykins, Kanakanagavalli Shravani Prakhya, Mary Jane Potash, Wei Chao, Jennifer Kelschenbach, David J. Volsky, Kelly Metcalf-Pate

Saha Cardiovascular Research Center Faculty Publications

EcoHIV is a model of HIV infection that recapitulates aspects of HIV-1 pathology in mice. However, there are limited published protocols to guide EcoHIV virion production. Here, we present a protocol for producing infective EcoHIV virions and essential quality controls. We describe steps for viral purification, titering, and multiple techniques to analyze infection efficacy. This protocol produces high infectivity in C57BL/6 mice which will aid investigators in generating preclinical data.

Bacteroides Fragilis In The Gut Microbiomes Of Alzheimer’S Disease Activates Microglia And Triggers Pathogenesis In Neuronal C/Ebpβ Transgenic Mice, Yiyuan Xia, Yifan Xiao, Zi-Hao Wang, Ashfaqul M. Alam, John P. Haran, Beth A. Mccormick, Xiji Shu, Xiaochuan Wang, Keqiang Ye

Markey Cancer Center Faculty Publications

Gut dysbiosis contributes to Alzheimer’s disease (AD) pathogenesis, and Bacteroides strains are selectively elevated in AD gut microbiota. However, it remains unknown which Bacteroides species and how their metabolites trigger AD pathologies. Here we show that Bacteroides fragilis and their metabolites 12-hydroxy-heptadecatrienoic acid (12-HHTrE) and Prostaglandin E2 (PGE2) activate microglia and induce AD pathogenesis in neuronal C/EBPβ transgenic mice. Recolonization of antibiotics cocktail-pretreated Thy1-C/EBPβ transgenic mice with AD patient fecal samples elicits AD pathologies, associated with C/EBPβ/Asparaginyl endopeptidase (AEP) pathway upregulation, microglia activation, and cognitive disorders compared to mice receiving healthy donors’ fecal microbiota transplantation (FMT). Microbial 16S rRNA sequencing …