Expression, purification, and evaluation of in vivo anti-fibrotic activity for soluble truncated TGF-β receptor II as a cleavable His-SUMO fusion protein

In animals, oral administration of the cholera toxin B (CTB) subunit conjugated to the autoantigen insulin enhances the specific immune-unresponsive state. This is called oral tolerance and is capable of suppressing autoimmune type 1 diabetes (T1D). However, the process by which the CTB-insulin (CTB-INS) protein works as a therapy for T1Din vivoremains unclear. Here, we successfully expressed a green fluorescent protein- (GFP-) tagged CTB-Ins (CTB-Ins-GFP) fusion protein in silkworms in a pentameric form that retained the native ability to activate the mechanism. Oral administration of the CTB-Ins-GFP protein induced special tolerance, delayed the development of diabetic symptoms, and suppressed T1D onset in nonobese diabetic (NOD) mice. Moreover, it increased the numbers of CD4+CD25+Foxp3+T regulatory (Treg) cells in peripheral lymph tissues and affected the biological activity of spleen cells. This study demonstrated that the CTB-Ins-GFP protein produced in silkworms acted as an oral protein vaccine, inducing immunological tolerance involving CD4+CD25+Foxp3+Treg cells in treating T1D.

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In vivo delivery of a XIAP (BIR3-RING) fusion protein containing the protein transduction domain protects against neuronal death induced by seizures

Experimental Neurology ◽

10.1016/j.expneurol.2005.08.021 ◽

2006 ◽

Vol 197 (2) ◽

pp. 301-308 ◽

Cited By ~ 14

Author(s):

Tianfu Li ◽

Yongfeng Fan ◽

Yumin Luo ◽

Baoguo Xiao ◽

Chuanzhen Lu

Keyword(s):

Fusion Protein ◽

Neuronal Death ◽

Protein Transduction ◽

Protein Transduction Domain ◽

Ring Fusion ◽

In Vivo Delivery

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GAGA Factor Isoforms Have Distinct but Overlapping Functions In Vivo

Molecular and Cellular Biology ◽

10.1128/mcb.21.24.8565-8574.2001 ◽

2001 ◽

Vol 21 (24) ◽

pp. 8565-8574 ◽

Cited By ~ 20

Author(s):

Anthony J. Greenberg ◽

Paul Schedl

Keyword(s):

Fusion Protein ◽

Transcriptional Activation ◽

Functional Difference ◽

Wild Type ◽

Gaga Factor ◽

Phenotypic Analysis ◽

Alternatively Spliced

ABSTRACT The Drosophila melanogaster GAGA factor (encoded by the Trithorax-like [Trl] gene) is required for correct chromatin architecture at diverse chromosomal sites. The Trl gene encodes two alternatively spliced isoforms of the GAGA factor (GAGA-519 and GAGA-581) that are identical except for the length and sequence of the C-terminal glutamine-rich (Q) domain. In vitro and tissue culture experiments failed to find any functional difference between the two isoforms. We made a set of transgenes that constitutively express cDNAs coding for either of the isoforms with the goal of elucidating their roles in vivo. Phenotypic analysis of the transgenes in Trl mutant background led us to the conclusion that GAGA-519 and GAGA-581 perform different, albeit largely overlapping, functions. We also expressed a fusion protein with LacZ disrupting the Q domain of GAGA-519. This LacZ fusion protein compensated for the loss of wild-type GAGA factor to a surprisingly large extent. This suggests that the Q domain either is not required for the essential functions performed by the GAGA protein or is exclusively used for tetramer formation. These results are inconsistent with a major role of the Q domain in chromatin remodeling or transcriptional activation. We also found that GAGA-LacZ was able to associate with sites not normally occupied by the GAGA factor, pointing to a role of the Q domain in binding site choice in vivo.

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A Novel Recombinant Fcγ Receptor-Targeted Survivin Combines with Chemotherapy for Efficient Cancer Treatment

Biomedicines ◽

10.3390/biomedicines9070806 ◽

2021 ◽

Vol 9 (7) ◽

pp. 806

Author(s):

Chiao-Chieh Wu ◽

Chen-Yi Chiang ◽

Shih-Jen Liu ◽

Hsin-Wei Chen

Keyword(s):

Immune Responses ◽

Fusion Protein ◽

Human Cancer ◽

Formyl Peptide Receptor ◽

Antigen Delivery ◽

Potential Candidate ◽

Fcγ Receptor ◽

Formyl Peptide ◽

Efficient Uptake

Formyl peptide receptor-like 1 inhibitor (FLIPr), an Fcγ receptor (FcγR) antagonist, can be used as a carrier to guide antigen-FLIPr fusion protein to FcγR then enhances antigen-specific immune responses. Survivin, a tumor-associated antigen, is over-expressed in various types of human cancer. In this study, we demonstrate that recombinant survivin-FLIPr fusion protein (rSur-FLIPr) binds to FcγRs, and efficient uptake by dendritic cells in vivo. In addition, rSur-FLIPr alone stimulates survivin-specific immune responses, which effectively suppresses the tumor growth. The antitumor immunities are through TAP-mediated and CD8-dependent pathways. Furthermore, preexisting anti-FLIPr antibody does not abolish antitumor responses induced by rSur-FLIPr immunization. These results suggest that FLIPr is an effective antigen delivery vector and can be repeatedly used. Combination of chemotherapy with rSur-FLIPr treatment reveals a great benefit to tumor-bearing mice. Altogether, these findings suggest that rSur-FLIPr is a potential candidate for efficient cancer therapy.

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The N-terminal domain of Sxl protein disrupts Sxl autoregulation in females and promotes female-specific splicing of tra in males

Development ◽

10.1242/dev.126.13.2841 ◽

1999 ◽

Vol 126 (13) ◽

pp. 2841-2853 ◽

Cited By ~ 1

Author(s):

G. Deshpande ◽

G. Calhoun ◽

P.D. Schedl

Keyword(s):

Fusion Protein ◽

Rna Binding ◽

Chimeric Protein ◽

Dominant Negative ◽

Terminal Domain ◽

Binding Domains ◽

Transcriptional Regulatory ◽

Dominant Negative Activity ◽

Female Specific

Sex determination in Drosophila depends upon the post-transcriptional regulatory activities of the Sex-lethal (Sxl) gene. Sxl maintains the female determined state and activates female differentiation pathways by directing the female-specific splicing of Sxl and tra pre-mRNAs. While there is compelling evidence that Sxl proteins regulate splicing by directly binding to target RNAs, previous studies indicate that the two Sxl RNA-binding domains are not in themselves sufficient for biological activity and that an intact N-terminal domain is also critical for splicing function. To further investigate the functions of the Sxl N terminus, we ectopically expressed a chimeric protein consisting of the N-terminal 99 amino acids fused to ss-galactosidase. The Nss-gal fusion protein behaves like a dominant negative, interfering with the Sxl autoregulatory feedback loop and killing females. This dominant negative activity can be attributed to the recruitment of the fusion protein into the large Sxl:Snf splicing complexes that are found in vivo and the consequent disruption of these complexes. In addition to the dominant negative activity, the Nss-gal fusion protein has a novel gain-of-function activity in males: it promotes the female-specific processing of tra pre-mRNAs. This novel activity is discussed in light of the blockage model for the tra splicing regulation.

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Abrogation of mesenchyme-specific TGF-β signaling results in lung malformation with prenatal pulmonary cysts in mice

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00299.2020 ◽

2021 ◽

Author(s):

Qing Miao ◽

Hui Chen ◽

Yongfeng Luo ◽

Joanne Chiu ◽

Ling Chu ◽

...

Keyword(s):

Lung Development ◽

Muscle Growth ◽

Branching Morphogenesis ◽

Mouse Lung ◽

Cystic Lesions ◽

Alveolar Epithelial ◽

Pulmonary Cysts ◽

Β Receptor ◽

Airway Branching

The TGF-β signaling pathway plays a pivotal role in controlling organogenesis during fetal development. Although the role of TGF-β signaling in promoting lung alveolar epithelial growth has been determined, mesenchymal TGF-β signaling in regulating lung development has not been studied in vivo due to a lack of genetic tools for specifically manipulating gene expression in lung mesenchymal cells. Therefore, the integral roles of TGF-β signaling in regulating lung development and congenital lung diseases are not completely understood. Using a Tbx4 lung enhancer-driven Tet-On inducible Cre transgenic mouse system, we have developed a mouse model in which lung mesenchyme-specific deletion of TGF-β receptor 2 gene (Tgfbr2) is achieved. Reduced airway branching accompanied by defective airway smooth muscle growth and later peripheral cystic lesions occurred when lung mesenchymal Tgfbr2 was deleted from embryonic day 13.5 to 15.5, resulting in postnatal death due to respiratory insufficiency. Although cell proliferation in both lung epithelium and mesenchyme was reduced, epithelial differentiation was not significantly affected. Tgfbr2 downstream Smad-independent ERK1/2 may mediate these mesenchymal effects of TGF-β signaling through the GSK3β--β-catenin--Wnt canonical pathway in fetal mouse lung. Our study suggests that Tgfbr2-mediated TGF-β signaling in prenatal lung mesenchyme is essential for lung development and maturation, and defective TGF-β signaling in lung mesenchyme may be related to abnormal airway branching morphogenesis and congenital airway cystic lesions.

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