Medical Biochemistry Commons^™

Loss Of Peroxiredoxin Iv Protects Mice From Azoxymethane/Dextran Sulfate Sodium-Induced Colorectal Cancer Development, Pratik Thapa, Hong Jiang, Na Ding, Yanning Hao, Aziza Alshahrani, Eun Young Lee, Junichi Fujii, Qiou Wei

Markey Cancer Center Faculty Publications

Peroxiredoxin IV (Prx4), a typical two-cysteine-containing member of the peroxidase family, functions as an antioxidant to maintain cellular redox homeostasis through the reduction of reactive oxygen species (ROS) via cycles of oxidation–reduction reactions. Under oxidative stress, all Prxs including Prx4 are inactivated as their catalytic cysteines undergo hyperoxidation, and hyperoxidized two-cysteine Prxs can be exclusively repaired and revitalized through the reduction cycle catalyzed by sulfiredoxin (Srx). Previously, we showed that Prx4 is a preferred substrate of Srx, and knockout of Srx in mice leads to resistance to azoxymethane/dextran sulfate sodium (AOM/DSS)-induced colon carcinogenesis. To further understand the significance of the …

Harnessing Transcriptionally Driven Chromosomal Instability Adaptation To Target Therapy-Refractory Lethal Prostate Cancer., Brittiny Dhital, Sandra Santasusagna, Perumalraja Kirthika, Michael Xu, Peiyao Li, Marc Carceles-Cordon, Rajesh K. Soni, Zhuoning Li, Ronald C. Hendrickson, Matthew J. Schiewer, William K. Kelly, Cora N. Sternberg, Jun Luo, Amaia Lujambio, Carlos Cordon-Cardo, Monica Alvarez-Fernandez, Marcos Malumbres, Haojie Huang, Adam Ertel, Josep Domingo-Domenech, Veronica Rodriguez-Bravo

Kimmel Cancer Center Papers, Presentations, and Grand Rounds

Metastatic prostate cancer (PCa) inevitably acquires resistance to standard therapy preceding lethality. Here, we unveil a chromosomal instability (CIN) tolerance mechanism as a therapeutic vulnerability of therapy-refractory lethal PCa. Through genomic and transcriptomic analysis of patient datasets, we find that castration and chemotherapy-resistant tumors display the highest CIN and mitotic kinase levels. Functional genomics screening coupled with quantitative phosphoproteomics identify MASTL kinase as a survival vulnerability specific of chemotherapy-resistant PCa cells. Mechanistically, MASTL upregulation is driven by transcriptional rewiring mechanisms involving the non-canonical transcription factors androgen receptor splice variant 7 and E2F7 in a circuitry that restrains deleterious CIN and …

A Complete Methyl-Lysine Binding Aromatic Cage Constructed By Two Domains Of Phf2, John R Horton, Jujun Zhou, Qin Chen, Xing Zhang, Mark T Bedford, Xiaodong Cheng

Faculty, Staff and Student Publications

The N-terminal half of PHF2 harbors both a plant homeodomain (PHD) and a Jumonji domain. The PHD recognizes both histone H3 trimethylated at lysine 4 and methylated nonhistone proteins including vaccinia-related kinase 1 (VRK1). The Jumonji domain erases the repressive dimethylation mark from histone H3 lysine 9 (H3K9me2) at select promoters. The N-terminal amino acid sequences of H3 (AR2TK4) and VRK1 (PR2VK4) bear an arginine at position 2 and lysine at position 4. Here, we show that the PHF2 N-terminal half binds to H3 and VRK1 peptides containing K4me3, with dissociation constants (KD values) of 160 nM and 42 nM, …

Differential Transcriptional Alterations In Detoxification Genes In Parkinson’S Disease In Egypt, Nourhan Shebl

Theses and Dissertations

PD is the most common motor neurodegenerative disease worldwide. The underlying cause of PD is still unknow, owingthis to the complexity of the disease. Often, genetics and environmental factors are collaborating in the initiation of the disease. Despite the diversity of its genetical and environmental profiles, the Egyptian population is one of the mostunderrepresented population in terms of PD research. In this study, we investigated PD through various perspectives tohighlight the complexity of the disease in Egypt, taking into consideration the diversity of the Egyptian population. We recruited PD patients and reference controls from 4 governorates: Cairo, Giza, Alexandria, and …

Semi-Quantitative Detection Of Pseudouridine Modifications And Type I/Ii I/Ii Hypermodifications In Human Mrnas Using Direct Long-Read Sequencing, Sepideh Tavakoli, Mohammad Nabizadeh, Amr Makhamreh, Howard Gamper, Caroline A Mccormick, Neda K Rezapour, Ya-Ming Hou, Meni Wanunu, Sara H Rouhanifard

Department of Biochemistry and Molecular Biology Faculty Papers

Here, we develop and apply a semi-quantitative method for the high-confidence identification of pseudouridylated sites on mammalian mRNAs via direct long-read nanopore sequencing. A comparative analysis of a modification-free transcriptome reveals that the depth of coverage and specific k-mer sequences are critical parameters for accurate basecalling. By adjusting these parameters for high-confidence U-to-C basecalling errors, we identify many known sites of pseudouridylation and uncover previously unreported uridine-modified sites, many of which fall in k-mers that are known targets of pseudouridine synthases. Identified sites are validated using 1000-mer synthetic RNA controls bearing a single pseudouridine in the center position, demonstrating systematic …

Sgc-Camkk2-1: A Chemical Probe For Camkk2, Carrow Wells, Yi Liang, Thomas L Pulliam, Chenchu Lin, Dominik Awad, Benjamin Eduful, Sean O'Byrne, Mohammad Anwar Hossain, Carolina Moura Costa Catta-Preta, Priscila Zonzini Ramos, Opher Gileadi, Carina Gileadi, Rafael M Couñago, Brittany Stork, Christopher G Langendorf, Kevin Nay, Jonathan S Oakhill, Debarati Mukherjee, Luigi Racioppi, Anthony R Means, Brian York, Donald P Mcdonnell, John W Scott, Daniel E Frigo, David H Drewry

Faculty, Staff and Student Publications

The serine/threonine protein kinase calcium/calmodulin-dependent protein kinase kinase 2 (CAMKK2) plays critical roles in a range of biological processes. Despite its importance, only a handful of inhibitors of CAMKK2 have been disclosed. Having a selective small molecule tool to interrogate this kinase will help demonstrate that CAMKK2 inhibition can be therapeutically beneficial. Herein, we disclose SGC-CAMKK2-1, a selective chemical probe that targets CAMKK2.

Disruption Of The Interaction Between Mutationally Activated Gαq And Gβγ Attenuates Aberrant Signaling, Jenna L Aumiller, Philip B Wedegaertner

Department of Biochemistry and Molecular Biology Faculty Papers

Heterotrimeric G protein stimulation via G protein-coupled receptors promotes downstream proliferative signaling. Mutations can occur in Gα proteins which prevent GTP hydrolysis; this allows the G proteins to signal independently of G protein-coupled receptors and can result in various cancers, such as uveal melanoma (UM). Most UM cases harbor Q209L, Q209P, or R183C mutations in Gαq/11 proteins, rendering the proteins constitutively active (CA). Although it is generally thought that active, GTP-bound Gα subunits are dissociated from and signal independently of Gβγ, accumulating evidence indicates that some CA Gα mutants, such as Gαq/11, retain binding to Gβγ, and this interaction is …

Differentiating Pc12 Cells To Evaluate Neurite Densities Through Live-Cell Imaging, Jordyn Karliner, Diane E Merry

Department of Biochemistry and Molecular Biology Faculty Papers

Although PC12 cells are a valuable tool in neuroscience research, previously published PC12 cell differentiation techniques fail to consider the variability in differentiation rates between different PC12 cell strains and clonal variants. Here, we present a comprehensive protocol to differentiate PC12 cells into equivalent neurite densities through live-cell imaging for morphological, immunocytochemical, and biochemical analyses. We detail steps on optimized substrate coating, plating techniques, culture media, validation steps, and quantification techniques.

Quantifying Metal Interactions With The Antimicrobial Peptide Calcitermin, Maria G. Aviles-Sandoval, Michael Stevenson

Featured Student Work

Antimicrobial Peptides (AMPs) are novel therapeutic agents that play important roles in the innate immune system with its ability to kill gram-positive and gram-negative bacteria. Antibiotics are losing their efficacy, thus requiring a larger dose of medicine for treatment resulting in the increase of antibiotic resistance. AMPs are an attractive approach for exploration due to their broad-spectrum activity and ease of synthesis. Furthermore, fewer bacteria have developed resistance to AMPs. Several AMPs have demonstrated increased antimicrobial activity with metal ions, like Zn(II) and Cu(II) binding to the peptide. In this study, we performed the purification of Calcitermin through high-performance liquid …

Observing Ceramide Pathway With Ferroptosis Via Mia Paca-2 Cell Treatment With Rsl3, Tazrin Rahman

AUCTUS: The Journal of Undergraduate Research and Creative Scholarship

Composed of sphingosine and a fatty acid, ceramides are lipid molecules that serve as key metabolic signaling molecules of a sphingolipid pathway. While it acts as a precursor of complex sphingolipids, inducing ceramide generation can cause cell stress leading to subsequent cell death via apoptosis, necrosis, and even mitophagy. With regards to cell death specifically, a novel form of regulated cell death, ferroptosis, has recently been recognized of necrotic nature. Its unique morphological features and distinct properties have been observed over the last several decades; however, the molecular features were not identifiable as pure evidence of cell death, until recently …

Role Of The Ormdls In Keratinocytes, Souad Hatmi

Theses and Dissertations

The epidermis, the outermost layer of the skin, serves as a vital barrier protecting the body from environmental stressors, pathogens, and dehydration. Keratinocytes, the predominant cell type in the epidermis, play a central role in maintaining skin integrity and barrier function. In normal human skin, keratinocytes move from the basal layer to the stratum corneum, the top layer of the epidermis. In their travel, keratinocytes undergo a process of terminal differentiation, loss their ability to proliferate, secrete sphingolipids into the extracellular space, and gradually loss their nucleus and organelles. Sphingolipids, particularly ceramides, are key components of the intercellular lipid lamellae …

การมีส่วนร่วมของปัจจัยทางพันธุกรรมต่อการสร้างซีซีซีดีเอ็นเอของไวรัสตับอักเสบบี, กัลยกร สิงห์สังข์

Chulalongkorn University Theses and Dissertations (Chula ETD)

การติดเชื้อไวรัสตับอักเสบบี (hepatitis B virus infection; HBV) เป็นปัญหาทางด้านสาธารณสุขที่สำคัญทั่วโลก เนื่องจากผู้ติดเชื้อไวรัสตับอักเสบบีสามารถพัฒนาความรุนแรงไปเป็นโรคตับแข็ง (cirrhosis) และมะเร็งตับ (hepatocellular carcinoma: HCC) ซึ่งนำไปสู่การเสียชีวิตได้ในที่สุด ถึงแม้ว่าในปัจจุบันโรคติดเชื้อไวรัสตับอักเสบบีสามารถรักษาได้ด้วยยาต้านไวรัส อย่างไรก็ตามยังไม่สามารถรักษาให้หายขาดได้ เนื่องจากการคงอยู่ของ covalently closed circular DNA (cccDNA) ซึ่งเป็น DNA สายคู่แบบวงปิดคล้ายโครโมโซมขนาดเล็กภายในนิวเคลียสของเซลล์ตับ ขบวนการสร้าง cccDNA จำเป็นต้องอาศัยการทำงานของปัจจัยทางพันธุกรรมของเซลล์เจ้าบ้าน (host factors) หลายชนิด โดยเฉพาะอย่างยิ่ง DNA damage binding protein 2 (DDB2) และ DNA polymerase δ (DNA pol δ) อย่างไรก็ตามการศึกษายังไม่พบการศึกษาการแสดงออกของ host factors ดังกล่าวต่อการสร้าง cccDNA ในเซลล์ตับของผู้ป่วยโรคไวรัสตับอักเสบบี โดยเฉพาะไวรัสตับอักเสบบีแบบแอบแฝง (occult HBV infection หรือ OBI) รวมทั้งยังไม่มีวิธีมาตรฐานในการตรวจวัดระดับของ cccDNA ในเซลล์ตับ ดังนั้นการศึกษานี้จึงมีวัตถุประสงค์ในการศึกษาความสัมพันธ์ของระดับของ cccDNA และ host factors ในเซลล์ตับของผู้ป่วยมะเร็งตับที่เกิดจากจากติดเชื้อไวรัสตับอักเสบบี (HBV-related HCC) และไวรัสตับอักเสบบีแบบแอบแฝง (occult HBV (OBI)-related HCC) ด้วยการทำdroplet digital PCR (ddPCR) และ Western blot ตามลำดับ จากเนื้อเยื่อบริเวณ non-tumor หรือ adjacent tissues ผลการศึกษาระดับการแสดงออกของ intrahepatic HBV DNA และ cccDNA ในเซลล์ตับ ของผู้ป่วย พบว่าผู้ป่วยกลุ่ม HBV-related HCC มีการแสดงออกของ intrahepatic HBV …

ความหลากหลายทางพันธุกรรมของโรคไขมันพอกตับที่สัมพันธ์กับการติดเชื้อเอชไอวีและโรคมะเร็งตับ, กมลชนก มูลลิสาร

Chulalongkorn University Theses and Dissertations (Chula ETD)

โรคไขมันพอกตับเป็นหนึ่งในสาเหตุหลักของโรคตับอักเสบเรื้อรังที่สามารถดำเนินโรคไปสู่โรคมะเร็งตับได้ ความชุกของโรคไขมันพอกตับในประเทศไทยพบประมาณร้อยละ 20-30 สาเหตุของการเกิดไขมันพอกตับที่พบบ่อยได้แก่โรคอ้วน เบาหวาน กลุ่มโรคเมตาบอลิกและปัจจัยทางพันธุกรรม การศึกษาที่ผ่านมาพบว่ามียีนหลายตำแหน่งที่มีบทบาทเกี่ยวข้องกับกลไกการเกิดโรค ที่สำคัญได้แก่ PNPLA3, TM6SF2 และ HSD17B13 งานวิจัยนี้มีจุดประสงค์เพื่อหาความสัมพันธ์ของความหลากหลายทางพันธุกรรมของยีนดังกล่าวกับการเกิดโรคไขมันพอกตับในคนไทย โดยศึกษาจีโนไทป์ของทั้งสามยีนด้วยเทคนิค TaqMan Real-time PCR จากตัวอย่างเลือด 900 ตัวอย่างของ 4 กลุ่มได้แก่ กลุ่มควบคุมที่มีสุขภาพดี ผู้ป่วยโรคไขมันพอกตับที่มีและไม่มีการติดเชื้อเอชไอวี และกลุ่มผู้ป่วยมะเร็งตับ ผลการศึกษาพบว่าจีโนไทป์ GG ของยีน PNPLA3 rs738409 เป็นปัจจัยเสี่ยงของการเกิดโรคไขมันพอกตับในทุกกลุ่มที่ศึกษาเมื่อเปรียบเทียบกับกลุ่มควบคุม ดังนี้ กลุ่มผู้ป่วยที่ไม่มีการติดเชื้อเอชไอวี [odds ratio (OR)=6.52, 95% confidence interval (CI) 2.64-16.08, P<0.001] กลุ่มผู้ป่วยที่มีการติดเชื้อเอชไอวี(OR=2.76, 95% CI 1.09-7.00, P=0.03) และกลุ่มผู้ป่วยมะเร็งตับ (OR=9.96, 95% CI 4.06-24.44, P<0.001) อย่างไรก็ตามไม่ พบความสัมพันธ์ของยีน TM6SF2 rs58542926 และ HSD17B13 rs6834314 กับการเกิดโรคของ กลุ่มตัวอย่างต่างๆ จากการวิเคราะห์หาปัจจัยเสี่ยงอิสระด้วยสมการถดถอยพบว่า ค่าดัชนีมวลกาย ที่สูง โรคเบาหวาน ระดับเอนไซม์ALT ที่สูงและการไม่ติดเชื้อเอชไอวีเป็นปัจจัยเสี่ยงในการทำให้ เกิดความรุนแรงในการสะสมไขมันในตับ โดยสรุปผลการวิจัยนี้พบว่าความหลากหลายทาง พันธุกรรมของ PNPLA3 มีความสัมพันธ์กับการเกิดโรคไขมันพอกตับและเพิ่มความเสี่ยงต่อการเกิด โรคมะเร็งตับในประชากรไทย แต่ไม่พบความสัมพันธ์ดังกล่าวสำหรับยีน TM6SF2 และ HSD17B13

Hif-1 Activation And Inflammatory Responses To Hypoxia, Shazreen Shaharuddin, Nik Mohd Afizan Nik Abd Rahman, Mas Jaffri Masarudin, Mohammedarfat N Alamassi

International Journal of Aviation, Aeronautics, and Aerospace

Acute hypoxia is a significant physiological danger during high-altitude flying and military aircraft missions. The human brain requires a constant supply of oxygen to function properly, and is susceptible to settings with low availability of air oxygen. Hypoxia can influence inflammatory signalling, and both central and systemic responses can activate HIF pathway genes. HIFs are critical molecules that regulate inflammation andhypoxia, ensuring appropriate cell function and survival. Hypoxia is the condition in which insufficient oxygen reaches the body's tissues. It can be caused by a decrease in partial oxygen pressure (PO2) in the environment, problems with breathing and/or oxygen transport, …

Microrna-1 Attenuates The Growth And Metastasis Of Small Cell Lung Cancer Through Cxcr4/Foxm1/Rrm2 Axis, Parvez Khan, Jawed A. Siddiqui, Prakash Kshirsagar Dr., Ramakanth Chirravuri Venkata, Shailendra K. Maurya, Tamara Mirzapoiazova, Naveenkumar Perumal, Sanjib Chaudhary, Ranjana K. Kanchan, Mahek Fatima, Md Arafat Khan, Asad Ur Rehman, Imayavaramban Lakshmanan, Sidharth Mahapatra, Geoffrey A. Talmon, Prakash Kulkarni, Apar Kishor Ganti, Maneesh Jain, Ravi Salgia, Surinder K. Batra, Mohd W. Nasser

Journal Articles: Biochemistry & Molecular Biology

BACKGROUND: Small cell lung cancer (SCLC) is an aggressive lung cancer subtype that is associated with high recurrence and poor prognosis. Due to lack of potential drug targets, SCLC patients have few therapeutic options. MicroRNAs (miRNAs) provide an interesting repertoire of therapeutic molecules; however, the identification of miRNAs regulating SCLC growth and metastasis and their precise regulatory mechanisms are not well understood.

METHODS: To identify novel miRNAs regulating SCLC, we performed miRNA-sequencing from donor/patient serum samples and analyzed the bulk RNA-sequencing data from the tumors of SCLC patients. Further, we developed a nanotechnology-based, highly sensitive method to detect microRNA-1 (miR-1, …

The Mucin Family Of Proteins: Candidates As Potential Biomarkers For Colon Cancer, Kristin E. Cox, Shanglei Liu, Thinzar M. Lwin, Robert M. Hoffman, Surinder K. Batra, Michael Bouvet

Journal Articles: Biochemistry & Molecular Biology

Mucins (MUC1-MUC24) are a family of glycoproteins involved in cell signaling and barrier protection. They have been implicated in the progression of numerous malignancies including gastric, pancreatic, ovarian, breast, and lung cancer. Mucins have also been extensively studied with respect to colorectal cancer. They have been found to have diverse expression profiles amongst the normal colon, benign hyperplastic polyps, pre-malignant polyps, and colon cancers. Those expressed in the normal colon include MUC2, MUC3, MUC4, MUC11, MUC12, MUC13, MUC15 (at low levels), and MUC21. Whereas MUC5, MUC6, MUC16, and MUC20 are absent from the normal colon and are expressed in colorectal …

Specific Targeting And Labeling Of Colonic Polyps In Cpc-Apc Mice With Mucin 5ac Fluorescent Antibodies: A Model For Detection Of Early Colon Cancer, Michael A. Turner, Kristin E. Cox, Shanglei Liu, Nicholas Neel, Siamak Amirfakhri, Hiroto Nishino, Mojgan Hosseini, Joshua A. Alcantara, Amer Ali Abd El-Hafeez, Thinzar M. Lwin, Kavita Mallya, Joseph R. Pisegna, Satish K. Singh, Pradipta Ghosh, Robert M. Hoffman, Surinder K. Batra, Michael Bouvet

Journal Articles: Biochemistry & Molecular Biology

Poor visualization of polyps can limit colorectal cancer screening. Fluorescent antibodies to mucin5AC (MUC5AC), a glycoprotein upregulated in adenomas and colorectal cancer, could improve screening colonoscopy polyp detection rate. Adenomatous polyposis coli flox mice with a Cdx2-Cre transgene (CPC-APC) develop colonic polyps that contain both dysplastic and malignant tissue. Mice received MUC5AC-IR800 or IRdye800 as a control IV and were sacrificed after 48 h for near-infrared imaging of their colons. A polyp-to-background ratio (PBR) was calculated for each polyp by dividing the mean fluorescence intensity of the polyp by the mean fluorescence intensity of the background tissue. The mean 25 …

Gpcrs And Fibroblast Heterogeneity In Fibroblast-Associated Diseases, Nidhi V. Dwivedi, Souvik Datta, Karim El-Kersh, Ruxana Sadikot Md, Mrcp, Apar Kishor Ganti, Surinder K. Batra, Maneesh Jain

Journal Articles: Biochemistry & Molecular Biology

G protein-coupled receptors (GPCRs) are the largest and most diverse class of signaling receptors. GPCRs regulate many functions in the human body and have earned the title of "most targeted receptors". About one-third of the commercially available drugs for various diseases target the GPCRs. Fibroblasts lay the architectural skeleton of the body, and play a key role in supporting the growth, maintenance, and repair of almost all tissues by responding to the cellular cues via diverse and intricate GPCR signaling pathways. This review discusses the dynamic architecture of the GPCRs and their intertwined signaling in pathological conditions such as idiopathic …

Immunotherapy: An Emerging Modality To Checkmate Brain Metastasis, Aatiya Ahmad, Parvez Khan, Asad Ur Rehman, Surinder K. Batra, Mohd W. Nasser

Journal Articles: Biochemistry & Molecular Biology

The diagnosis of brain metastasis (BrM) has historically been a dooming diagnosis that is nothing less than a death sentence, with few treatment options for palliation or prolonging life. Among the few treatment options available, brain radiotherapy (RT) and surgical resection have been the backbone of therapy. Within the past couple of years, immunotherapy (IT), alone and in combination with traditional treatments, has emerged as a reckoning force to combat the spread of BrM and shrink tumor burden. This review compiles recent reports describing the potential role of IT in the treatment of BrM in various cancers. It also examines …

Roles Unveiled For Membrane-Associated Mucins At The Ocular Surface Using A Muc4 Knockout Mouse Model, Rafael Martinez-Carrasco, Satyanarayan Rachagani, Surinder K. Batra, Pablo Argüeso, M Elizabeth Fini

Journal Articles: Biochemistry & Molecular Biology

Membrane-associated mucins (MAMs) are proposed to play critical roles at the ocular surface; however, in vivo evidence has been lacking. Here we investigate these roles by phenotyping of a Muc4 KO mouse. Histochemical analysis for expression of the beta-galactosidase transgene replacing Muc4 revealed a spiraling ribbon pattern across the corneal epithelium, consistent with centripetal cell migration from the limbus. Depletion of Muc4 compromised transcellular barrier function, as evidenced by an increase in rose bengal staining. In addition, the corneal surface was less smooth, consistent with disruption of tear film stability. While surface cells presented with well-developed microprojections, an increase in …