scholarly journals Protein Transduction Domains Fused to Virus Receptors Improve Cellular Virus Uptake and Enhance Oncolysis by Tumor-Specific Replicating Vectors

2004 ◽  
Vol 78 (24) ◽  
pp. 13743-13754 ◽  
Author(s):  
Florian Kühnel ◽  
Bernd Schulte ◽  
Thomas Wirth ◽  
Norman Woller ◽  
Sonja Schäfers ◽  
...  
Keyword(s):  
Viral Vectors ◽  
Protein Transduction ◽  
Virus Binding ◽  
Adenoviral Infection ◽  
Protein Transduction Domains ◽  
Oncolytic Therapy ◽  
Uptake Rates ◽  
Immunodeficiency Virus ◽  
Transactivator Of Transcription ◽  
Adenovirus Receptor

ABSTRACT Expression of cellular receptors determines viral tropism and limits gene delivery by viral vectors. Protein transduction domains (PTDs) have been shown to deliver proteins, antisense oligonucleotides, liposomes, or plasmid DNA into cells. In our study, we investigated the role of several PTD motifs in adenoviral infection. When physiologically expressed, a PTD from human immunodeficiency virus transactivator of transcription (Tat) did not improve adenoviral infection. We therefore fused PTDs to the ectodomain of the coxsackievirus-adenovirus receptor (CARex) to attach PTDs to adenoviral fiber knobs. CARex-Tat and CARex-VP22 allowed efficient adenoviral infection in nonpermissive cells and significantly improved viral uptake rates in permissive cells. Dose-dependent competition of CARex-PTD-mediated infection using CARex and inhibition experiments with heparin showed that binding of CARex-PTD to both adenoviral fiber and cellular glycosaminoglycans is essential for the improvement of infection. CARex-PTD-treated adenoviruses retained their properties after density gradient ultracentrifugation, indicating stable binding of CARex-PTD to adenoviral particles. Consequently, the mechanism of CARex-PTD-mediated infection involves coating of the viral fiber knobs by CARex-PTD, rather than placement of CARex domains on cell surfaces. Expression of CARex-PTDs led to enhanced lysis of permissive and nonpermissive tumor cells by replicating adenoviruses, indicating that CARex-PTDs are valuable tools to improve the efficacy of oncolytic therapy. Together, our study shows that CARex-PTDs facilitate gene transfer in nonpermissive cells and improve viral uptake at reduced titers and infection times. The data suggest that PTDs fused to virus binding receptors may be a valuable tool to overcome natural tropism of vectors and could be of great interest for gene therapeutic approaches.

scholarly journals Tumor Antigen LRRC15 Impedes Adenoviral Infection: Implications for Virus-Based Cancer Therapy

2008 ◽  
Vol 82 (12) ◽  
pp. 5933-5939 ◽  
Author(s):  
Jim O'Prey ◽  
Simon Wilkinson ◽  
Kevin M. Ryan
Keyword(s):  
Cell Death ◽  
Tumor Antigen ◽  
Tumor Suppressor P53 ◽  
Adenoviral Infection ◽  
Multiple Tumor ◽  
Oncolytic Therapy ◽  
Adenoviral Delivery ◽  
Per Se ◽  
Tumor Types ◽  
Adenovirus Receptor

ABSTRACT Adenoviruses for gene or oncolytic therapy are under development. Notable among these strategies is adenoviral delivery of the tumor suppressor p53. Since all therapeutics have limitations in certain settings, we have undertaken retroviral suppressor screens to identify genes conferring resistance to adenovirus-delivered p53. These studies identified the tumor antigen LRRC15, which is frequently overexpressed in multiple tumor types, as a repressor of cell death due to adenoviral p53. LRRC15, however, does not impede p53 function per se but impedes adenoviral infection. Specifically, LRRC15 causes redistribution of the coxsackievirus-adenovirus receptor away from the cell surface. This effect is manifested in less adenoviral binding to the surfaces of LRRC15-expressing cells. This discovery, therefore, not only is important for understanding adenoviral biology but also has potentially important implications for adenovirus-based anticancer therapeutics.

scholarly journals TAT-Conjugated NDUFS8 Can Be Transduced into Mitochondria in a Membrane-Potential-Independent Manner and Rescue Complex I Deficiency

2021 ◽  
Vol 22 (12) ◽  
pp. 6524
Author(s):  
Bo-Yu Lin ◽  
Gui-Teng Zheng ◽  
Kai-Wen Teng ◽  
Juan-Yu Chang ◽  
Chao-Chang Lee ◽  
...  
Keyword(s):  
Membrane Potential ◽  
Fusion Proteins ◽  
Complex I ◽  
Nadh Dehydrogenase ◽  
Leigh Syndrome ◽  
Protein Transduction ◽  
Independent Manner ◽  
Complex I Deficiency ◽  
Transactivator Of Transcription ◽  
Hiv 1

NADH dehydrogenase (ubiquinone) Fe-S protein 8 (NDUFS8) is a nuclear-encoded core subunit of human mitochondrial complex I. Defects in NDUFS8 are associated with Leigh syndrome and encephalomyopathy. Cell-penetrating peptide derived from the HIV-1 transactivator of transcription protein (TAT) has been successfully applied as a carrier to bring fusion proteins into cells without compromising the biological function of the cargoes. In this study, we developed a TAT-mediated protein transduction system to rescue complex I deficiency caused by NDUFS8 defects. Two fusion proteins (TAT-NDUFS8 and NDUFS8-TAT) were exogenously expressed and purified from Escherichia coli for transduction of human cells. In addition, similar constructs were generated and used in transfection studies for comparison. The results showed that both exogenous TAT-NDUFS8 and NDUFS8-TAT were delivered into mitochondria and correctly processed. Interestingly, the mitochondrial import of TAT-containing NDUFS8 was independent of mitochondrial membrane potential. Treatment with TAT-NDUFS8 not only significantly improved the assembly of complex I in an NDUFS8-deficient cell line, but also partially rescued complex I functions both in the in-gel activity assay and the oxygen consumption assay. Our current findings suggest the considerable potential of applying the TAT-mediated protein transduction system for treatment of complex I deficiency.

scholarly journals The Coxsackievirus and Adenovirus Receptor: Glycosylation and the Extracellular D2 Domain Are Not Required for Coxsackievirus B3 Infection

2016 ◽  
Vol 90 (12) ◽  
pp. 5601-5610 ◽  
Author(s):  
Sandra Pinkert ◽  
Carsten Röger ◽  
Jens Kurreck ◽  
Jeffrey M. Bergelson ◽  
Henry Fechner
Keyword(s):  
Virus Infection ◽  
Immunoglobulin Superfamily ◽  
Minor Importance ◽  
Receptor Function ◽  
Virus Binding ◽  
D2 Domain ◽  
Cvb3 Infection ◽  
Coxsackie B Viruses ◽  
Coxsackievirus And Adenovirus Receptor ◽  
Adenovirus Receptor

ABSTRACTThe coxsackievirus and adenovirus receptor (CAR) is a member of the immunoglobulin superfamily (IgSF) and functions as a receptor for coxsackie B viruses (CVBs). The extracellular portion of CAR comprises two glycosylated immunoglobulin-like domains, D1 and D2. CAR-D1 binds to the virus and is essential for virus infection; however, it is not known whether D2 is also important for infection, and the role of glycosylation has not been explored. To understand the function of these structural components in CAR-mediated CVB3 infection, we generated a panel of human (h) CAR deletion and substitution mutants and analyzed their functionality as CVB receptors, examining both virus binding and replication. Lack of glycosylation of the CAR-D1 or -D2 domains did not adversely affect CVB3 binding or infection, indicating that the glycosylation of CAR is not required for its receptor functions. Deletion of the D2 domain reduced CVB3 binding, with a proportionate reduction in the efficiency of virus infection. Replacement of D2 with the homologous D2 domain from chicken CAR, or with the heterologous type C2 immunoglobulin-like domain from IgSF11, another IgSF member, fully restored receptor function; however, replacement of CAR-D2 with domains from CD155 or CD80 restored function only in part. These data indicate that glycosylation of the extracellular domain of hCAR plays no role in CVB3 receptor function and that CAR-D2 is not specifically required. The D2 domain may function largely as a spacer permitting virus access to D1; however, the data may also suggest that D2 affects virus binding by influencing the conformation of D1.IMPORTANCEAn important step in virus infection is the initial interaction of the virus with its cellular receptor. Although the role in infection of the extracellular CAR-D1, cytoplasmic, and transmembrane domains have been analyzed extensively, nothing is known about the function of CAR-D2 and the extracellular glycosylation of CAR. Our data indicate that glycosylation of the extracellular CAR domain has only minor importance for the function of CAR as CVB3 receptor and that the D2 domain is not essential per se but contributes to receptor function by promoting the exposure of the D1 domain on the cell surface. These results contribute to our understanding of the coxsackievirus-receptor interactions.

Modulation of human dendritic-cell function following transduction with viral vectors: implications for gene therapy

Blood ◽  
2005 ◽  
Vol 105 (10) ◽  
pp. 3824-3832 ◽  
Author(s):  
Peng H. Tan ◽  
Sven C. Beutelspacher ◽  
Shao-An Xue ◽  
Yao-He Wang ◽  
Peter Mitchell ◽  
...  
Keyword(s):  
Dendritic Cell ◽  
Mhc Class Ii ◽  
Viral Vectors ◽  
Cell Function ◽  
Leukemia Virus ◽  
Moloney Murine Leukemia Virus ◽  
Histocompatibility Complex ◽  
Immunodeficiency Virus ◽  
Anemia Virus ◽  
And Function

AbstractGenetic modification of dendritic-cell (DC) function is an attractive approach to treat disease, either using mature DCs (mDCs) to immunize patients, or immature DCs (iDCs) to induce tolerance. Viral vectors are efficient at transducing DCs, and we have investigated the effect of transduction with a variety of viral vectors on the phenotype and function of DCs. Adenovirus (Ad), human immunodeficiency virus (HIV), equine anemia virus (EIAV), and Moloney murine leukemia virus (MMLV) all up-regulate costimulatory molecules and major histocompatibility complex (MHC) class II expression on DCs, as well as, in the case of Ad and lentiviral vectors, inducing production of Th1 and proinflammatory cytokines. Following transduction there is activation of double-stranded (ds) RNA-triggered pathways resulting in interferon (IFN) α/β production. In addition, the function of virally infected DCs is altered; iDCs have an increased, and mDCs a decreased, ability to stimulate a mixed lymphocyte reaction (MLR). Viral transduction of mDCs results in up-regulation of the indoleamine 2,3-dioxygenase (IDO) enzyme, which down-regulates T-cell responsiveness. Inhibition of IDO restores the ability of mDCs to stimulate an MLR, indicating that IDO is responsible for the modulation of mDC function. These data have important implications for the use of viral vectors in the transduction of DCs.

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