Characterization of constitutive promoters for piggyBac transposon-mediated stable transgene expression in mesenchymal stem cells (MSCs).
Multipotent mesenchymal stem cells (MSCs) can undergo self-renewal and give rise to multi-lineages under given differentiation cues. It is frequently desirable to achieve a stable and high level of transgene expression in MSCs in order to elucidate possible molecular mechanisms through which MSC self-renewal and lineage commitment are regulated. Retroviral or lentiviral vector-mediated gene expression in MSCs usually decreases over time. Here, we choose to use the piggyBac transposon system and conduct a systematic comparison of six commonly-used constitutive promoters for their abilities to drive RFP or firefly luciferase expression in somatic HEK-293 cells and MSC iMEF cells. The analyzed promoters include three viral promoters (CMV, CMV-IVS, and SV40), one housekeeping gene promoter (UbC), and two composite promoters of viral and housekeeping gene promoters (hEFH and CAG-hEFH). CMV-derived promoters are shown to drive the highest transgene expression in HEK-293 cells, which is however significantly reduced in MSCs. Conversely, the composite promoter hEFH exhibits the highest transgene expression in MSCs whereas its promoter activity is modest in HEK-293 cells. The reduced transgene expression driven by CMV promoters in MSCs may be at least in part caused by DNA methylation, or to a lesser extent histone deacetlyation. However, the hEFH promoter is not significantly affected by these epigenetic modifications. Taken together, our results demonstrate that the hEFH composite promoter may be an ideal promoter to drive long-term and high level transgene expression using the piggyBac transposon vector in progenitor cells such as MSCs.
Stem Cell Biology and Therapy Laboratory of Ministry of Education Key Laboratory for Pediatrics, Chongqing Stem Cell Therapy and Engineering Center, and Department of Urology, The Children's Hospital of Chongqing Medical University, Chongqing, China.
Sheng
S
Wen
Chongqing Key Laboratory for Oral Diseases and Biomedical Sciences, and the Affiliated Hospital of Stomatology, Chongqing Medical University, Chongqing, China; Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America.
Hongmei
H
Zhang
Stem Cell Biology and Therapy Laboratory of Ministry of Education Key Laboratory for Pediatrics, Chongqing Stem Cell Therapy and Engineering Center, and Department of Urology, The Children's Hospital of Chongqing Medical University, Chongqing, China.
Yasha
Y
Li
Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America; Departments of Cell Biology and Oncology of the Affiliated Southwest Hospital, the Third Military Medical University, Chongqing, China.
Ning
N
Wang
Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America; Ministry of Education Key Laboratory of Diagnostic Medicine and School of Clinical Diagnostic Medicine, and the Affiliated Hospitals, Chongqing Medical University, Chongqing, China.
Wenwen
W
Zhang
Stem Cell Biology and Therapy Laboratory of Ministry of Education Key Laboratory for Pediatrics, Chongqing Stem Cell Therapy and Engineering Center, and Department of Urology, The Children's Hospital of Chongqing Medical University, Chongqing, China; Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America.
Ke
K
Yang
Stem Cell Biology and Therapy Laboratory of Ministry of Education Key Laboratory for Pediatrics, Chongqing Stem Cell Therapy and Engineering Center, and Department of Urology, The Children's Hospital of Chongqing Medical University, Chongqing, China; Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America; Ministry of Education Key Laboratory of Diagnostic Medicine and School of Clinical Diagnostic Medicine, and the Affiliated Hospitals, Chongqing Medical University, Chongqing, China.
Ningning
N
Wu
Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America; Ministry of Education Key Laboratory of Diagnostic Medicine and School of Clinical Diagnostic Medicine, and the Affiliated Hospitals, Chongqing Medical University, Chongqing, China.
Xian
X
Chen
Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America; Departments of Cell Biology and Oncology of the Affiliated Southwest Hospital, the Third Military Medical University, Chongqing, China.
Fang
F
Deng
Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America; Department of Orthopaedic Surgery, the Affiliated Xiang-Ya Hospital of Central South University, Changsha, China.
Zhan
Z
Liao
Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America; Ministry of Education Key Laboratory of Diagnostic Medicine and School of Clinical Diagnostic Medicine, and the Affiliated Hospitals, Chongqing Medical University, Chongqing, China.
Junhui
J
Zhang
Stem Cell Biology and Therapy Laboratory of Ministry of Education Key Laboratory for Pediatrics, Chongqing Stem Cell Therapy and Engineering Center, and Department of Urology, The Children's Hospital of Chongqing Medical University, Chongqing, China; Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America.
Qian
Q
Zhang
Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America; Ministry of Education Key Laboratory of Diagnostic Medicine and School of Clinical Diagnostic Medicine, and the Affiliated Hospitals, Chongqing Medical University, Chongqing, China.
Zhengjian
Z
Yan
Stem Cell Biology and Therapy Laboratory of Ministry of Education Key Laboratory for Pediatrics, Chongqing Stem Cell Therapy and Engineering Center, and Department of Urology, The Children's Hospital of Chongqing Medical University, Chongqing, China; Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America.
Wei
W
Liu
Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America; Department of Surgery, the Affiliated Zhongnan Hospital of Wuhan University, Wuhan, China.
Zhonglin
Z
Zhang
Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America; School of Bioengineering, Chongqing University, Chongqing, China.
Jixing
J
Ye
Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America; Ministry of Education Key Laboratory of Diagnostic Medicine and School of Clinical Diagnostic Medicine, and the Affiliated Hospitals, Chongqing Medical University, Chongqing, China.
Youlin
Y
Deng
Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America.
Guolin
G
Zhou
Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America.
Hue H
HH
Luu
Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America.
Rex C
RC
Haydon
Molecular Oncology Laboratory, Department of Orthopaedic Surgery and Rehabilitation Medicine, The University of Chicago Medical Center, Chicago, Illinois, United States of America.
Lewis L
LL
Shi
Stem Cell Biology and Therapy Laboratory of Ministry of Education Key Laboratory for Pediatrics, Chongqing Stem Cell Therapy and Engineering Center, and Department of Urology, The Children's Hospital of Chongqing Medical University, Chongqing, China; Ministry of Education Key Laboratory of Diagnostic Medicine and School of Clinical Diagnostic Medicine, and the Affiliated Hospitals, Chongqing Medical University, Chongqing, China.
Tong-Chuan
TC
He
Stem Cell Biology and Therapy Laboratory of Ministry of Education Key Laboratory for Pediatrics, Chongqing Stem Cell Therapy and Engineering Center, and Department of Urology, The Children's Hospital of Chongqing Medical University, Chongqing, China.
Guanghui
G
Wei
D000107:Acetylation; D002460:Cell Line; D019175:DNA Methylation; D004251:DNA Transposable Elements; D015870:Gene Expression; D020868:Gene Silencing; D018014:Gene Transfer Techniques; D017930:Genes, Reporter; D005822:Genetic Vectors; D057809:HEK293 Cells; D006657:Histones / Q000378:metabolism; D059765:Homologous Recombination; D006801:Humans; D059630:Mesenchymal Stem Cells / Q000378:metabolism*; D011401:Promoter Regions, Genetic; D019076:Transgenes
D004251:DNA Transposable Elements; D006657:Histones