Efficient gene transfer by hybrid retroviral vectors to murine spermatogenic cells.
Using murine spermatogenic cell lines GC-1 spg and GC-2 spd(ts) as target cells, an attempt was made to design a retroviral vector that would transduce genes efficiently. Promoter activities of various retroviral long terminal repeats (LTRs) were examined by using chloramphenicol acetyltransferase (CAT) as a reporter. The U3 region of spleen focus-forming virus (SFFVp) showed higher enhancer activity than that of Moloney murine leukemia virus (Mo-MuLV) in both cell lines. The U3 region of myeloproliferative sarcoma virus (MPSV) showed higher activity only in GC-1 spg cells. Expression was suppressed by the repressor element of the primer-binding site (PBS) of the Moloney-related virus. The efficiency of transduction of the multidrug-resistance gene (mdr-1) by an Mo-MuLV-based vector was compared with hybrid vectors consisting of the murine embryonic stem cell virus (MESV) PBS and the LTR of either SFFVp or MPSV. Rhodamine efflux assays and colchicine-resistant colony-forming assays demonstrated higher gene expression by the hybrid vectors. Amphotropic and ecotropic receptors were found to be expressed and functional in both cell lines. Thus, these hybrid vectors represent a powerful tool by which to transfer genes into spermatogenic cells.
Danno
S
S
Department of Clinical Molecular Biology, Faculty of Medicine, Kyoto University, Japan.
Itoh
K
K
Baum
C
C
Ostertag
W
W
Ohnishi
N
N
Kido
T
T
Tomiwa
K
K
Matsuda
T
T
Fujita
J
J
D000818:Animals; D002460:Cell Line; D015500:Chloramphenicol O-Acetyltransferase / Q000378:metabolism; D003078:Colchicine / Q000494:pharmacology; D005434:Flow Cytometry; D015870:Gene Expression; D018014:Gene Transfer Techniques; D019450:Genes, MDR; D017930:Genes, Reporter; D005822:Genetic Vectors; D008297:Male; D051379:Mice; D010957:Plasmids; D012190:Retroviridae / Q000235:genetics*; D012235:Rhodamines / Q000378:metabolism; D013091:Spermatogenesis; D013094:Spermatozoa; D014161:Transduction, Genetic
D012235:Rhodamines; D015500:Chloramphenicol O-Acetyltransferase; D003078:Colchicine