In order to evaluate the efficiency of the tetracycline-regulated gene expression system in Drosophila, we have generated transgenic lines expressing a tetracycline-controlled transactivator protein (tTA), with specific expression patterns during embryonic and larval development. These lines were used to direct expression of a tTA-responsive promoter fused to the coding region of either the beta-galactosidase or the homeotic protein Antennapedia (ANTP), under various conditions of tetracycline treatment. We found that expression of beta-galactosidase can be efficiently inhibited in embryos and larvae with tetracycline provided in the food, and that a simple removal of the larvae from tetracycline exposure results in the induction of the enzyme in a time- and concentration-dependent manner. Similar treatments can be used to prevent the lethality associated with the ectopic expression of ANTP in embryos and, subsequently, to control the timing of expression of the homeoprotein ANTP specifically in the antennal imaginal disc. Our results show that the expression of a gene placed under the control of a tetracycline-responsive promoter can be tightly controlled, both spatially by the regulatory sequences driving the expression of tTA and temporally by tetracycline. This provides the basis of a versatile binary system for controlling gene expression in Drosophila, with an additional level of regulation as compared to the general method using the yeast transcription factor GAL4.
REFERENCES
REFERENCES
Brand
A. H.
, Perrimon
N.
(1993
) Targeted gene expression as a mean of altering cell fates and generating dominant phenotypes.
Development
118
, 401
–415
Calleja
M.
, Moreno
E.
, Pelaz
S.
, Morata
G.
(1996
) Visualization of gene expression in living adult Drosophila.
Science
274
, 252
–255
Capdevila
J.
, Guerrero
I.
(1994
) Targeted expression of the signaling molecule decapentaplegic induces pattern duplications and growth alterations in Drosophila wings.
EMBO J
13
, 4459
–4468
Chrast-Balz
J.
, Hooft van Huijsduijnen
R.
(1996
) Bi-directional gene switching with the tetracycline repressor and a novel tetracycline antagonist.
Nucleic Acids Res
24
, 2900
–2904
Condie
J. M.
, Mustard
J. A.
, Bower
D. L.
(1991
) Generation of anti- Antennapedia monoclonal antibodies and Antennapedia protein expression in imaginal discs.
Drosophila Inf. Service
70
, 52
–54
Efrat
S.
, Fusco-DeMane
D.
, Lemberg
H.
, Al Emran
O.
, Wang
X.
(1995
) Conditional transformation of a pancreatic-cell line derived from transgenic mice expressing a tetracycline-regulated oncogene.
Proc. Nat. Acad. Sci. USA
92
, 3576
–3580
Ewald
D.
, Li
M.
, Efrat
S.
, Auer
G.
, Wall
R. J.
, Furth
P. A.
, Hennighausen
L.
(1996
) Time-sensitive reversal of hyperplasia in transgenic mice expressing SV40 T antigen.
Science
273
, 1384
–1386
Ferver
J. F.
, Störtkhuhl
K. F.
, Stocker
R. F.
, Greenspan
R. J.
(1995
) Genetic feminization of brain structures and changed sexual orientation in male Drosophila.
Science
267
, 902
–905
Freeman
M.
(1996
) Reiterative use of the EGF receptor triggers differentiation of all cell types in the Drosophila eye.
Cell
87
, 651
–660
Gibson
G.
, Gehring
W. J.
(1988
) Head and thoracic transformations caused by ectopic expression of Antennapedia during Drosophila development.
Development
102
, 657
–675
Gibson
G.
, Schier
A.
, LeMotte
P.
, Gehring
W. J.
(1990
) The specificities of Sex combs reduced and Antennapedia are defined by a distinct portion of each protein that includes the homeodomain.
Cell
62
, 1087
–1103
Gossen
M.
, Bujard
H.
(1992
) Tight control of gene expression in mammalian cells by tetracycline-responsive promoters.
Proc. Nat. Acad. Sci. USA
89
, 5547
–5551
Gossen
M.
, Bujard
H.
(1993
) Anhydrotetracycline, a novel effector for tetracycline controlled gene expression systems in eukaryotic cell.
Nucleic Acids Res
21
, 4411
–4412
Gossen
M.
, Freundlieb
S.
, Bender
G.
, Muller
G.
, Hillen
W.
, Bujard
H.
(1995
) Transcriptional activation by tetracyclines in mammalian cells.
Science
268
, 1766
–1769
Haerry
T. H.
, Gehring
W. J.
(1996
) Intron of the mouse Hoxa-7 gene contains conserved homeodomain binding sites that can function as an enhancer element in Drosophila.
Proc. Nat. Acad. Sci. USA
93
, 13884
–13889
Halder
G.
, Callaerts
P.
, Gehring
W. J.
(1995
) Induction of ectopic eyes by targeted expression of the eyeless gene in Drosophila.
Science
267
, 1788
–1792
Henninghausen
L.
, Wall
R. J.
, Tillmann
U.
, Li
M.
, Furth
P. A.
(1995
) Conditional gene expression in secretory tissues and skin of transgenic mice using the MMTV-LTR and the tetracycline responsive system.
J. Cell Biochem
59
, 463
–472
Hinrichs
W.
, Kisker
C.
, Duvel
M.
, Muller
A.
, Tovar
K.
, Hillen
W.
, Saenger
W.
(1994
) Structure of the tet repressor-tetracycline complex and regulation of antibiotic resistance.
Science
264
, 418
–420
Hinz
U.
, Giebel
B.
, Campos-Ortega
J. A.
(1994
) The basic-helix-loop-helix domain of Drosophila lethal of scute protein is sufficient for proneural function and activates neurogenic genes.
Cell
76
, 77
–87
Kistner
A.
, Gossen
M.
, Zimmermann
F.
, Jerecic
J.
, Ullmer
C.
, Lubbert
H.
, Bujard
H.
(1996
) Doxycycline-mediated quantitative and tissue-specific control of gene expression in transgenic mice.
Proc. Nat. Acad. Sci. USA
93
, 10933
–10938
Larsen
E.
, Lee
T.
, Glickman
N.
(1996
) Antenna to leg transformation: dynamics of developmental competence.
Dev. Genet
19
, 333
–339
Mayford
M.
, Bach
M. E.
, Huang
Y. Y.
, Wang
L.
, Hawkins
R. D.
, Kandel
E. R.
(1996
) Control of memory formation through regulated expression of a CaMKII transgene.
Science
274
, 1678
–1683
Morimura
S.
, Maves
L.
, Chen
Y.
, Hoffmann
F. M.
(1996
) decapentaplegic overexpression affects Drosophila wing and leg imaginal disc development and wingless expression.
Dev. Biol
177
, 136
–151
Pignoni
F.
, Zipursky
S. L.
(1997
) Induction of Drosophila eye development by Decapentaplegic.
Development
124
, 271
–278
Quiring
R.
, Walldorf
U.
, Kloter
U.
, Gehring
W. J.
(1994
) Homology of the eyeless gene of Drosophila to the Small eye gene in mice and Aniridia in Humans.
Science
265
, 785
–789
Ready
D. F.
, Hanson
T. E.
, Benzer
S.
(1976
) Development of the Drosophila retina, a neurocrystalline lattice.
Dev. Biol
53
, 217
–240
Rimmington
G.
, Dalby
B.
, Glover
D. M.
(1994
) Expression of an N-terminally truncated cyclin B in the Drosophila larval brain leads to mitotic delay at late anaphase.
J. Cell. Sci
107
, 2729
–2738
Scanga
S.
, Manoukian
A.
, Larsen
E.
(1995
) Time-and concentration-dependent response of the Drosophila antenna imaginal disc to Antennapedia.
Dev. Biol
169
, 673
–682
Shockett
P.
, Difilippantonio
M.
, Hellman
N.
, Schatz
D. G.
(1995
) A modified tetracycline-regulated system provides autoregulatory, inducible gene expression in cultured cells and transgenic mice.
Proc. Nat. Acad. Sci. USA
92
, 6522
–6526
Shockett
P. E.
, Schatz
D. G.
(1996
) Diverse strategies for tetracycline-regulated inducible gene expression.
Proc. Nat. Acad. Sci. USA
93
, 5173
–5176
Speicher
S. A.
, Thomas
U.
, Hinz
U.
, Knust
E.
(1994
) The Serrate locus of Drosophila and its role in morphogenesis of the wing imaginal discs: control of cell proliferation.
Development
140
, 535
–544
St-Onge
L.
, Furth
P. A.
, Gruss
P.
(1996
) Temporal control of the Cre recombinase in transgenic mice by a tetracycline responsive promoter.
Nucleic Acids Res
24
, 3875
–3877
Struhl
G.
, Basler
K.
(1993
) Organising activity of the wingless protein in Drosophila.
Cell
72
, 527
–540
Thummel
C. S.
, Pirrotta
V.
(1992
) New pCaSpeR P element vectors.
Drosophila Inf. Service
71
, 150
–.Wagner-Bernholz
J. T.
, Wilson
C.
, Gibson
G.
, Schuh
R.
, Gehring
W. J.
(1991
) Identification of target genes of the homeotic gene Antennapedia by enhancer detection.
Genes Dev
5
, 2467
–2480
Wharton
K. A.
, Crews
S. T.
(1993
) CNS midline enhancers of the Drosophila split and toll genes.
Mech. Dev
40
, 141
–154
Wilson
C.
, Bellen
H. J.
, Gehring
W. J.
(1990
) Position effects on eukaryotic gene expression.
Annu. Rev. Cell Biol
6
, 679
–714
Wirz
J.
, Fessler
L. I.
, Gehring
W. J.
(1986
) Localization of the Antennapedia protein in Drosophila embryos and imaginal discs.
EMBO J
12
, 3327
–3334
Zink
D.
, Paro
R.
(1995
) Drosophila Polycomb-group regulated chromatin inhibits the accessibility of a trans-activator to its target DNA.
EMBO J
14
, 5660
–5671
This content is only available via PDF.
© 1998 by Company of Biologists
1998
