During hair growth, cortical cells emerging from the proliferative follicle bulb rapidly undergo a differentiation program and synthesise large amounts of hair keratin proteins. To identify some of the controls that specify expression of hair genes we have defined the minimal promoter of the wool keratin intermediate filament gene K2.10. The region of this gene spanning nucleotides −350 to +53 was sufficient to direct expression of the lacZ gene to the follicle cortex of transgenic mice but deletion of nucleotides −350 to −150 led to a complete loss of promoter activity. When a four base substitution mutation was introduced into the minimal functional promoter at the binding site for lymphoid enhancer factor 1 (LEF-1), promoter activity in transgenic mice was decreased but specificity was not affected. To investigate the interaction of trans-acting factors within the minimal K2.10 promoter we performed DNase I footprinting analyses and electrophoretic mobility shift assays. In addition to LEF-1, Sp1, AP2-like and NF1-like proteins bound to the promoter. The Sp1 and AP2-like proteins bound sequences flanking the LEF-1 binding site whereas the NF1-like proteins bound closer to the transcription start site. We conclude that the LEF-1 binding site is an enhancer element of the K2.10 promoter in the hair follicle cortex and that factors other than LEF-1 regulate promoter tissue- and differentiation-specificity.

REFERENCES

Adams
A. D.
,
Choate
D. M.
,
Thompson
M. A.
(
1995
).
NF1-L is the DNA-binding component of the protein complex at the peripherin negative regulatory element.
J. Biol. Chem
270
,
6975
6983
Bawden
C. S.
,
Powell
B. C.
,
Walker
S. K.
,
Rogers
G. E.
(
1998
).
Expression of a wool intermediate filament keratin transgene in sheep alters fibre structure.
Transgenic Research
7
,
1
15
Briggs
M. R.
,
Kadonaga
J. T.
,
Bell
S. P.
,
Tjian
R.
(
1986
).
Purification and biochemical characterization of the promoter-specific transcription factor, Sp1.
Science
234
,
47
52
Chaudhry
A. Z.
,
Lyons
G. E.
,
Gronostajski
R. M.
(
1997
).
Expression patterns of the four nuclear factor 1 genes during mouse embryogenesis indicate a potential role in development.
Dev. Dynam
208
,
313
325
Dillon
N.
,
Grosveld
F.
(
1993
).
Transcriptional regulation of multigene loci: multilevel control.
Trends Genet
9
,
134
137
Eckert
R. L.
,
Crish
J. F.
,
Banks
E. B.
,
Welter
J. F.
(
1997
).
The epidermis: genes on—genes off.
J. Invest. Dermatol
109
,
501
509
Fire
A.
(
1992
).
Histochemical techniques for locating Escherichia coli -galactosidase activity in transgenic organisms.
Genet. Anal. Tech. Appl
9
,
151
158
Giese
K.
,
Grosschedl
R.
(
1993
).
LEF-1 contains an activation domain that stimulates transcription only in a specific context of factor-binding sites.
EMBO J
12
,
4667
4676
Giese
K.
,
Kingsley
C.
,
Kirshner
J. R.
,
Grosschedl
R.
(
1995
).
Assembly and function of a TCRenhancer complex is dependent on LEF-1-induced DNA bending and multiple protein-protein interactions.
Genes Dev
9
,
995
1008
Gottschalk
L. R.
,
Leiden
J. M.
(
1990
).
Identification and functional characterization of the human T-cell receptorgene transcriptional enhancer: common nuclear proteins interact with the transcriptional regulatory elements of the T-cell receptor andgenes.
Mol. Cell. Biol
10
,
5486
5495
Grosschedl
R.
,
Giese
K.
,
Pagel
J.
(
1994
).
HMG domain proteins: Architectural elements in the assembly of nucleoprotein structures.
Trends Genet
10
,
94
100
Haynes
T. L.
,
Thomas
M. B.
,
Dusing
M. R.
,
Valerius
M. T.
,
Potter
S. S.
,
Wiginton
D. A.
(
1996
).
An enhancer LEF-1/TCF-1 site is essential for insertion site-independent transgene expression in thymus.
Nucl. Acids Res
24
,
5034
5044
Jahroudi
N.
,
Ardekani
A. M.
,
Greenberger
J. S.
(
1996
).
An NF1-like protein functions as a repressor of the von Willebrand factor promoter.
J. Biol. Chem
271
,
21413
21421
Jerkins
A. A.
,
Liu
W. R.
,
Lee
S.
,
Sul
H. S.
(
1995
).
Characterization of the murine mitochondrial glycerol-3-phosphate acyltransferase promoter.
J. Biol. Chem
270
,
1416
1421
Kaytes
P.
,
McNab
A. R.
,
Rea
T. J.
,
Groppi
V.
,
Kawabe
T. T.
,
Buhl
A. E.
,
Bertolino
A. P.
,
Hatzenbuhler
N. T.
,
Vogeli
G.
(
1991
).
Hair-specific keratins: characterization and expression of a mouse type I keratin gene.
J. Invest. Dermatol
97
,
835
842
Keough
R. A.
,
Powell
B. C.
,
Rogers
G. E.
(
1995
).
Targeted expression of SV-40 T antigen in the hair follicle of transgenic mice produces an aberrant phenotype.
J. Cell Sci
108
,
957
966
Kratochwil
K.
,
Dull
M.
,
Farinas
I.
,
Galceran
J.
,
Grosschedl
R.
(
1996
).
Lef1 expression is activated by BMP-4 and regulates inductive tissue interactions in tooth and hair development.
Genes Dev
10
,
1382
1394
Leask
A.
,
Byrne
C.
,
Fuchs
E.
(
1991
).
Transcription factor AP2 and its role in epidermal-specific gene expression.
Proc. Nat. Acad. Sci. USA
88
,
7948
7952
Leegwater
P. A. J.
,
van Driel
W.
,
van der Vliet
P. C.
(
1985
).
Recognition site of nuclear factor 1, a sequence-specific DNA-binding protein from HeLa cells that stimulates adenovirus DNA replication.
EMBO J
4
,
1515
1521
McNab
A. R.
,
Wood
L.
,
Theriault
N.
,
Gierman
T.
,
Vogeli
G.
(
1989
).
An ultra-high sulphur keratin gene is expressed specifically during hair growth.
J. Invest. Dermatol
92
,
263
266
McNab
A. R.
,
Andrus
P.
,
Wagner
T. E.
,
Buhl
A. E.
,
Waldon
D. J.
,
Kawabe
T. T.
,
Rea
T. J.
,
Groppi
V.
,
Vogeli
G.
(
1990
).
Hair-specific expression of chloramphenicol acetyltransferase in transgenic mice under the control of an ultra-high-sulphur keratin promoter.
Proc. Nat. Acad. Sci. USA
87
,
6848
6852
O'Connor
M.
,
Bernard
H.-U.
(
1995
).
Oct-1 activates the epithelial-specific enhancer of human papillomavirus type 16 via a synergistic interaction with NF1 at a conserved composite regulatory element.
Virology
207
,
77
88
Ohtsuki
M.
,
Flanagan
S.
,
Freedberg
I. M.
,
Blumenberg
M.
(
1993
).
A cluster of five nuclear proteins regulates keratin gene transcription.
Gene Expression
3
,
201
213
Oosterwegel
M.
,
van de Wetering
M.
,
Timmerman
J.
,
Kruisbeek
A.
,
Destree
O.
,
Meijlink
F.
,
Clevers
H.
(
1993
).
Differential expression of the HMG box factors TCF-1 and LEF-1 during murine embryogenesis.
Development
118
,
439
448
Pascal
E.
,
Tjian
R.
(
1991
).
Different activation domains of Sp1 govern formation of multimers and mediate transcriptional synergism.
Genes Dev
5
,
1646
1656
Persengiev
S. P.
,
Saffer
J. D.
,
Kilpatrick
D. L.
(
1995
).
An alternatively spliced form of the transcription factor Sp1 containing only a single glutamine-rich transactivation domain.
Proc. Nat. Acad. Sci. USA
92
,
9107
9111
Phi-Van
L.
,
von Kries
J. P.
,
Osterag
W.
,
Stratling
W. H.
(
1990
).
The chicken lysozyme 5matrix attachment region increases transcription from a heterologous promoter in heterologous cells and dampens position effects on the expression of transfected genes.
Mol. Cell. Biol
10
,
2302
2307
Powell
B. C.
,
Rogers
G. E.
(
1990
).
Cyclic hair-loss and regrowth in transgenic mice overexpressing an intermediate filament gene.
EMBO J
9
,
1485
1493
Powell
B. C.
,
Nesci
A.
,
Rogers
G. E.
(
1991
).
Regulation of keratin gene expression in hair follicle differentiation.
Ann. NY Acad. Sci
642
,
1
20
Powell
B.
,
Crocker
L. A.
,
Rogers
G. E.
(
1992
).
Hair follicle differentiation: expression, structure and evolutionary conservation of the hair type II keratin intermediate filament gene family.
Development
114
,
417
434
Redondo
J. M.
,
Hata
S.
,
Brockelhurst
C.
,
Krangel
M. S.
(
1990
).
A T cell-specific transcriptional enhancer within the human T cell receptor delta locus.
Science
247
,
1225
1229
Rogers
G. E.
,
Martinet
N.
,
Steinert
P.
,
Wynn
P.
,
Roop
D.
,
Kilkenny
A.
,
Morgan
D.
,
Yuspa
S.
(
1987
).
Cultivation of murine hair follicles as organoids in a collagen matrix.
J. Invest. Dermatol
89
,
369
379
Rogers
M. A.
,
Winter
H.
,
Langbein
L.
,
Krieg
T.
,
Schweizer
J.
(
1996
).
Genomic characterization of the human type I cuticular hair keratin hHa2 and identification of an adjacent novel type I hair keratin gene hHa5.
J. Invest. Dermatol
107
,
633
638
Rogers
M. A.
,
Langbein
L.
,
Praetzel
S.
,
Moll
I.
,
Krieg
T.
,
Winter
H.
,
Schweizer
J.
(
1997
).
Sequences and differential expression of three novel human type II hair keratins.
Differentiation
61
,
187
194
Roy
R. J.
,
Gosslin
P.
,
Anzivino
M. J.
,
Moore
D. D.
,
Guerin
S. L.
(
1992
).
Binding of a nuclear protein to the rat growth hormone silencer element.
Nucl. Acids Res
20
,
401
408
Sanes
J. R.
,
Rubenstein
J. L. R.
,
Nicolas
J.-F.
(
1986
).
Use of a recombinant retrovirus to study post-implantation cell lineage in mouse embryos.
EMBO J
5
,
3133
3142
Sawada
S.
,
Littman
D. R.
(
1991
).
Identification and characterization of a T-cell-specific enhancer adjacent to the murine CD4 gene.
Mol. Cell. Biol
11
,
5506
5515
Schneider
R.
,
Gander
I.
,
Muller
V.
,
Mertz
R.
,
Winnacker
E. L.
(
1986
).
A sensitive and rapid gel retention assay for nuclear factor I and other DNA-binding proteins in crude nuclear extracts.
Nucl. Acids Res
14
,
1303
1317
Sheridan
P. L.
,
Sheline
C. T.
,
Cannon
K.
,
Voz
M. L.
,
Pazin
M. J.
,
Kadonaga
J. T.
,
Jones
K. A.
(
1995
).
Activation of the HIV-1 enhancer by the LEF-1 HMG protein on nucleosome-assembled DNA in vitro.
Genes Dev
9
,
2090
2104
Steinert
P. M.
,
Roop
D. R.
(
1988
).
Molecular and cellular biology of intermediate filaments.
Annu. Rev. Biochem
57
,
593
625
Tobiasch
E.
,
Schweizer
J.
,
Winter
H.
(
1992
).
Structure and site of expression of a murine type II hair keratin.
Mol. Biol. Rep
16
,
39
47
Travis
A.
,
Amsterdam
A.
,
Belanger
C.
,
Grosschedl
R.
(
1991
).
LEF-1, a gene encoding a lymphoid-specific protein with an HMG domain, regulates T-cell receptorenhancer function.
Genes Dev
5
,
880
894
van Genderen
C.
,
Okamura
R. M.
,
Farinas
I.
,
Quo
R.-G.
,
Parslow
T. G.
,
Bruhn
L.
,
Grosschedl
R.
(
1994
).
Development of several organs that require inductive epithelial-mesenchymal interactions is impaired in LEF-1-deficient mice.
Genes Dev
8
,
2691
2703
Walters
M. C.
,
Magis
W.
,
Fiering
S.
,
Eidemiller
J.
,
Scalzo
D.
,
Groudine
M.
,
Martin
D. I. K.
(
1996
).
Transcriptional enhancers act in cis to suppress position-effect variegation.
Genes Dev
10
,
185
195
Waterman
M. L.
,
Fischer
W. H.
,
Jones
K. A.
(
1991
).
A thymus-specific member of the HMG protein family regulates the human T cell receptor Cenhancer.
Genes Dev
5
,
656
669
Williams
T.
,
Admon
A.
,
Luscher
B.
,
Tjian
R.
(
1988
).
Cloning and expression of AP-2 a cell-type-specific transcription factor that activates inducible enhancer elements.
Genes Dev
2
,
1557
1569
Winter
H.
,
Siry
P.
,
Tobiasch
E.
,
Schweizer
J.
(
1994
).
Sequence and expression of murine type I hair keratins mHa2 and mHa3.
Exp. Cell Res
212
,
190
200
Wu
R.-L.
,
Chen
T.-T.
,
Sun
T.-T.
(
1994
).
Functional importance of an Sp1-and an NFkB-related nuclear protein in a keratinocyte-specific promoter of rabbit K3 keratin gene.
J. Biol. Chem
269
,
28450
28459
Zhou
P.
,
Byrne
C.
,
Jacobs
J.
,
Fuchs
E.
(
1995
).
Lymphoid enhancer factor 1 directs hair follicle patterning and epithelial cell fate.
Genes Dev
9
,
570
583
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