Specific pharmacological dimerization of KDR in lentivirally transduced human hematopoietic cells activates anti-apoptotic and proliferative mechanisms. FASEB J 2005 Oct;19(12):1752-4
Date
08/04/2005Pubmed ID
16076962DOI
10.1096/fj.05-4006fjeScopus ID
2-s2.0-26444514555 (requires institutional sign-in at Scopus site) 10 CitationsAbstract
Selective and regulatable expansion of transduced cells could augment gene therapy for many disorders. The activation of modified growth factor receptors via synthetic chemical inducers of dimerization allows for the coordinated growth of transduced cells. This system can also provide information on specific receptor-mediated signaling without interference from other family members. Although several receptor subunits have been investigated in this context, little is known about the precise molecular events associated with dimerizer-initiated signaling. We have constructed and expressed an AP20187-regulated KDR chimeric receptor in human TF1 cells and analyzed activation of this gene switch using functional, biochemical, and microarray analyses. When deprived of natural ligands, GM-CSF, interleukin-3, or erythropoietin, AP20187 prevented apoptosis of transduced TF1 cells, induced dose-dependent proliferation, and supported long-term growth. In addition, AP20187 stimulation activated the signaling molecules associated with mitogen-activated protein kinase and phosphatidyl-inositol 3-kinase/Akt pathways. Microarray analysis determined that a number of transcripts involved in a variety of cellular processes were differentially expressed. Notably, mRNAs affiliated with heat stress, including Hsp70 and Hsp105, were up-regulated. Functional assays showed that Hsp70 and Hsp105 protected transduced TF1 cells from apoptosis and premature senescence, in part through regulation of Akt. These observations delineate specific roles for kinase insert domain-containing receptor, or KDR, signaling and suggest strategies to endow genetically modified cells with a survival advantage enabling the generation of adequate cell numbers for therapeutic outcomes.
Author List
Siatskas C, Underwood J, Ramezani A, Hawley RG, Medin JAAuthor
Jeffrey A. Medin PhD Professor in the Pediatrics department at Medical College of WisconsinMESH terms used to index this publication - Major topics in bold
ApoptosisCell Line
Cell Line, Tumor
Cell Proliferation
Cell Transformation, Viral
Dimerization
Dose-Response Relationship, Drug
Down-Regulation
Erythropoietin
Flow Cytometry
Gene Transfer Techniques
Genetic Therapy
Genetic Vectors
Granulocyte-Macrophage Colony-Stimulating Factor
Green Fluorescent Proteins
HSP110 Heat-Shock Proteins
HSP70 Heat-Shock Proteins
HeLa Cells
Hematopoietic Stem Cells
Humans
Immunoblotting
Interleukin-3
Lentivirus
Oligonucleotide Array Sequence Analysis
Phosphorylation
Protein Structure, Tertiary
RNA, Messenger
Reverse Transcriptase Polymerase Chain Reaction
Signal Transduction
Tacrolimus
Transfection
Up-Regulation
Vascular Endothelial Growth Factor Receptor-2
