We used transgenesis to explore the requirement for downregulation of hepatocyte nuclear factor 6 (HNF6) expression in the assembly, differentiation, and function of pancreatic islets. In vivo, HNF6 expression becomes downregulated in pancreatic endocrine cells at 18. 5 days post coitum (d.p.c.), when definitive islets first begin to organize. We used an islet-specific regulatory element (pdx1(PB)) from pancreatic/duodenal homeobox (pdx1) gene to maintain HNF6 expression in endocrine cells beyond 18.5 d.p.c. Transgenic animals were diabetic. HNF6-overexpressing islets were hyperplastic and remained very close to the pancreatic ducts. Strikingly, alpha, delta, and PP cells were increased in number and abnormally intermingled with islet beta cells. Although several mature beta cell markers were expressed in beta cells of transgenic islets, the glucose transporter GLUT2 was absent or severely reduced. As glucose uptake/metabolism is essential for insulin secretion, decreased GLUT2 may contribute to the etiology of diabetes in pdx1(PB)-HNF6 transgenics. Concordantly, blood insulin was not raised by glucose challenge, suggesting profound beta cell dysfunction. Thus, we have shown that HNF6 downregulation during islet ontogeny is critical to normal pancreas formation and function: continued expression impairs the clustering of endocrine cells and their separation from the ductal epithelium, disrupts the spatial organization of endocrine cell types within the islet, and severely compromises beta cell physiology, leading to overt diabetes.

REFERENCES

Ahlgren
U.
,
Jonsson
J.
,
Jonsson
L.
,
Simu
K.
,
Edlund
H.
(
1998
)
-cell-specific inactivation of the mouse Ipf1/Pdx1 gene results in loss of the beta-cell phenotype and maturity onset diabetes.
Genes Dev
12
,
1763
1768
Ahlgren
U.
,
Pfaff
S. L.
,
Jessell
T. M.
,
Edlund
T.
,
Edlund
H.
(
1997
)
Independent requirement for ISL1 in formation of pancreatic mesenchyme and islet cells.
Nature
385
,
257
260
Ang
S. L.
,
Rossant
J.
(
1994
)
HNF-3is essential for node and notochord formation in mouse development.
Cell
78
,
561
574
Bennett
M. V.
,
Goodenough
D. A.
(
1978
)
Gap junctions, electrotonic coupling, and intercellular communication.
Neurosci. Res. Program. Bull
16
,
1
486
Bosco
D.
,
Orci
L.
,
Meda
P.
(
1989
)
Homologous but not heterologous contact increases the insulin secretion of individual pancreatic-cells.
Exp. Cell Res
184
,
72
80
Cirulli
V.
,
Baetens
D.
,
Rutishauser
U.
,
Halban
P. A.
,
Orci
L.
,
Rouiller
D. G.
(
1994
)
Expression of neural cell adhesion molecule (N-CAM) in rat islets and its role in islet cell type segregation.
J. Cell Sci
107
,
1429
1436
Cirulli
V.
,
Crisa
L.
,
Beattie
G. M.
,
Mally
M. I.
,
Lopez
A. D.
,
Fannon
A.
,
Ptasznik
A.
,
Inverardi
L.
,
Ricordi
C.
,
Deerinck
T.
, et al. 
(
1998
)
KSA antigen Ep-CAM mediates cell-cell adhesion of pancreatic epithelial cells: morphoregulatory roles in pancreatic islet development.
J. Cell Biol
140
,
1519
1534
Cockell
M.
,
Stevenson
B. J.
,
Strubin
M.
,
Hagenbuchle
O.
,
Wellauer
P. K.
(
1989
)
Identification of a cell-specific DNA-binding activity that interacts with a transcriptional activator of genes expressed in the acinar pancreas.
Mol. Cell. Biol
9
,
2464
2476
Dahl
U.
,
Sjodin
A.
,
Semb
H.
(
1996
)
Cadherins regulate aggregation of pancreatic-cells in vivo.
Development
122
,
2895
2902
Dutta
S.
,
Bonner-Weir
S.
,
Montminy
M.
,
Wright
C.
(
1998
)
Regulatory factor linked to late-onset diabetes?.
Nature
392
,
560
–.
Edlund
H.
(
1998
)
Transcribing pancreas.
Diabetes
47
,
1817
1823
Esni
F.
,
Taljedal
I. B.
,
Perl
A. K.
,
Cremer
H.
,
Christofori
G.
,
Semb
H.
(
1999
)
Neural cell adhesion molecule (N-CAM) is required for cell type segregation and normal ultrastructure in pancreatic islets.
J. Cell Biol
144
,
325
337
Gepts
W.
,
Lecompte
P. M.
(
1981
)
The pancreatic islets in diabetes.
Am. J. Med
70
,
105
115
Gittes
G. K.
,
Rutter
W. J.
(
1992
)
Onset of cell-specific gene expression in the developing mouse pancreas.
Proc. Natl. Acad. Sci. USA
89
,
1128
1132
Gu
D.
,
Sarvetnick
N.
(
1993
)
Epithelial cell proliferation and islet neogenesis in IFN-transgenic mice.
Development
118
,
33
46
Guillam
M. T.
,
Hummler
E.
,
Schaerer
E.
,
Yeh
J. I.
,
Birnbaum
M. J.
,
Beermann
F.
,
Schmidt
A.
,
Deriaz
N.
,
Thorens
B.
,
Wu
J. Y.
(
1997
)
Early diabetes and abnormal postnatal pancreatic islet development in mice lacking Glut-2.
Nat. Genet
17
,
327
330
Halban
P. A.
,
Wollheim
C. B.
,
Blondel
B.
,
Meda
P.
,
Niesor
E. N.
,
Mintz
D. H.
(
1982
)
The possible importance of contact between pancreatic islet cells for the control of insulin release.
Endocrinology
111
,
86
94
Henquin
J. C.
(
1987
)
Regulation of insulin release by ionic and electrical events incells.
Horm. Res
27
,
168
178
Herrera
P. L.
,
Huarte
J.
,
Sanvito
F.
,
Meda
P.
,
Orci
L.
,
Vassalli
J. D.
(
1991
)
Embryogenesis of the murine endocrine pancreas; early expression of pancreatic polypeptide gene.
Development
113
,
1257
1265
Inoue
K.
,
Cetkovic-Cvrlje
M.
,
Eizirik
D. L.
,
Grill
V.
(
1994
)
Irreversible loss of normal-cell regulation by glucose in neonatally streptozotocin diabetic rats.
Diabetologia
37
,
351
357
Jansson
L.
,
Eizirik
D. L.
,
Pipeleers
D. G.
,
Borg
L. A.
,
Hellerstrom
C.
,
Andersson
A.
(
1995
)
Impairment of glucose-induced insulin secretion in human pancreatic islets transplanted to diabetic nude mice.
J. Clin. Invest
96
,
721
726
Jonsson
J.
,
Carlsson
L.
,
Edlund
T.
,
Edlund
H.
(
1994
)
Insulin-promoter-factor 1 is required for pancreas development in mice.
Nature
371
,
606
609
Krapp
A.
,
Knofler
M.
,
Frutiger
S.
,
Hughes
G. J.
,
Hagenbuchle
O.
,
Wellauer
P. K.
(
1996
)
The p48 DNA-binding subunit of transcription factor PTF1 is a new exocrine pancreas-specific basic helix-loop-helix protein.
EMBO J
15
,
4317
4329
Krapp
A.
,
Knofler
M.
,
Ledermann
B.
,
Burki
K.
,
Berney
C.
,
Zoerkler
N.
,
Hagenbuchle
O.
,
Wellauer
P. K.
(
1998
)
The bHLH protein PTF1-p48 is essential for the formation of the exocrine and the correct spatial organization of the endocrine pancreas.
Genes Dev
12
,
3752
3763
Landry
C.
,
Clotman
F.
,
Hioki
T.
,
Oda
H.
,
Picard
J. J.
,
Lemaigre
F. P.
,
Rousseau
G. G.
(
1997
)
HNF-6 is expressed in endoderm derivatives and nervous system of the mouse embryo and participates to the cross-regulatory network of liver-enriched transcription factors.
Dev. Biol
192
,
247
257
Larsson
L. I.
,
Madsen
O. D.
,
Serup
P.
,
Jonsson
J.
,
Edlund
H.
(
1996
)
Pancreatic-duodenal homeobox 1—role in gastric endocrine patterning.
Mech. Dev
60
,
175
184
Leahy
J. L.
,
Bonner-Weir
S.
,
Weir
G. C.
(
1992
)
-cell dysfunction induced by chronic hyperglycemia. Current ideas on mechanism of impaired glucose-induced insulin secretion.
Diabetes Care
15
,
442
455
Lemaigre
F. P.
,
Durviaux
S. M.
,
Truong
O.
,
Lannoy
V. J.
,
Hsuan
J. J.
,
Rousseau
G. G.
(
1996
)
Hepatocyte nuclear factor 6, a transcription factor that contains a novel type of homeodomain and a single cut domain.
Proc. Natl. Acad. Sci. USA
93
,
9460
9464
Madsen
O. D.
,
Jensen
J.
,
Petersen
H. V.
,
Pedersen
E. E.
,
Oster
A.
,
Andersen
F. G.
,
Jorgensen
M. C.
,
Jensen
P. B.
,
Larsson
L. I.
,
Serup
P.
(
1997
)
Transcription factors contributing to the pancreatic-cell phenotype.
Horm. Metab. Res
29
,
265
270
Marynissen
G.
,
Leclercq-Meyer
V.
,
Sener
A.
,
Malaisse
W. J.
(
1990
)
Perturbation of pancreatic islet function in glucose-infused rats.
Metabolism
39
,
87
95
Meissner
H. P.
(
1976
)
Electrophysiological evidence for coupling betweencells of pancreatic islets.
Nature
262
,
502
504
Naya
F. J.
,
Huang
H. P.
,
Qiu
Y.
,
Mutoh
H.
,
DeMayo
F. J.
,
Leiter
A. B.
,
Tsai
M. J.
(
1997
)
Diabetes, defective pancreatic morphogenesis, and abnormal enteroendocrine differentiation in BETA2/neuroD-deficient mice.
Genes Dev
11
,
2323
2334
Naya
F. J.
,
Stellrecht
C. M.
,
Tsai
M. J.
(
1995
)
Tissue-specific regulation of the insulin gene by a novel basic helix-loop-helix transcription factor.
Genes Dev
9
,
1009
1019
Offield
M. F.
,
Jetton
T. L.
,
Labosky
P. A.
,
Ray
M.
,
Stein
R. W.
,
Magnuson
M. A.
,
Hogan
B. L.
,
Wright
C. V.
(
1996
)
PDX1 is required for pancreatic outgrowth and differentiation of the rostral duodenum.
Development
122
,
983
995
Orci
L.
,
Thorens
B.
,
Ravazzola
M.
,
Lodish
H. F.
(
1989
)
Localization of the pancreaticcell glucose transporter to specific plasma membrane domains.
Science
245
,
295
297
Oster
A.
,
Jensen
J.
,
Serup
P.
,
Galante
P.
,
Madsen
O. D.
,
Larsson
L. I.
(
1998
).
Rat endocrine pancreatic development in relation to two homeobox gene products (Pdx-1 and Nkx 6.1).
J. Histochem. Cytochem
46
,
707
715
Peshavaria
M.
,
Gamer
L.
,
Henderson
E.
,
Teitelman
G.
,
Wright
C. V.
,
Stein
R.
(
1994
)
XIHbox 8, an endoderm-specific Xenopus homeodomain protein, is closely related to a mammalian insulin gene transcription factor.
Mol. Endocrinol
8
,
806
816
Pierreux
C. E.
,
Stafford
J.
,
Demonte
D.
,
Scott
D. K.
,
Vandenhaute
J.
,
O'Brien
R. M.
,
Granner
D. K.
,
Rousseau
G. G.
,
Lemaigre
F. P.
(
1999
)
Antiglucocorticoid activity of hepatocyte nuclear factor-6.
Proc. Natl. Acad. Sci. USA
96
,
8961
8966
Rausa
F.
,
Samadani
U.
,
Ye
H.
,
Lim
L.
,
Fletcher
C. F.
,
Jenkins
N. A.
,
Copeland
N. G.
,
Costa
R. H.
(
1997
)
The cut-homeodomain transcriptional activator HNF-6 is coexpressed with its target gene HNF-3in the developing murine liver and pancreas.
Dev. Biol
192
,
228
246
Rouiller
D. G.
,
Cirulli
V.
,
Halban
P. A.
(
1990
)
Differences in aggregation properties and levels of the neural cell adhesion molecule (NCAM) between islet cell types.
Exp. Cell Res
191
,
305
312
Samadani
U.
,
Costa
R. H.
(
1996
)
The transcriptional activator hepatocyte nuclear factor 6 regulates liver gene expression.
Mol. Cell Biol
16
,
6273
6284
Samols
E.
,
Stagner
J. I.
,
Ewart
R. B.
,
Marks
V.
(
1988
)
The order of islet microvascular cellular perfusion is B—A—D in the perfused rat pancreas.
J. Clin. Invest
82
,
350
353
Sander
M.
,
Neubuser
A.
,
Kalamaras
J.
,
Ee
H. C.
,
Martin
G. R.
,
German
M. S.
(
1997
)
Genetic analysis reveals that PAX6 is required for normal transcription of pancreatic hormone genes and islet development.
Genes Dev
11
,
1662
1673
Song
S. Y.
,
Gannon
M.
,
Washington
M. K.
,
Scoggins
C. R.
,
Meszoely
I. M.
,
Goldenring
J. R.
,
Marino
C. R.
,
Sandgren
E. P.
,
Coffey
R. J.
Jr.
,
Wright
C. V.
, et al. 
(
1999
)
Expansion of Pdx1-expressing pancreatic epithelium and islet neogenesis in transgenic mice overexpressing transforming growth factor alpha.
Gastroenterology
117
,
1416
1426
Sosa-Pineda
B.
,
Chowdhury
K.
,
Torres
M.
,
Oliver
G.
,
Gruss
P.
(
1997
)
The Pax4 gene is essential for differentiation of insulin-producingcells in the mammalian pancreas.
Nature
386
,
399
402
St-Onge
L.
,
Sosa-Pineda
B.
,
Chowdhury
K.
,
Mansouri
A.
,
Gruss
P.
(
1997
)
Pax6 is required for differentiation of glucagon-producing-cells in mouse pancreas.
Nature
387
,
406
409
Stoffers
D. A.
,
Ferrer
J.
,
Clarke
W. L.
,
Habener
J. F.
(
1997
)
Early-onset type-II diabetes mellitus (MODY4) linked to IPF1.
Nat. Genet
17
,
138
139
Stoffers
D. A.
,
Zinkin
N. T.
,
Stanojevic
V.
,
Clarke
W. L.
,
Habener
J. F.
(
1997
)
Pancreatic agenesis attributable to a single nucleotide deletion in the human IPF1 gene coding sequence.
Nat. Genet
15
,
106
110
Stoffers
D. A.
,
Stanojevic
V.
,
Habener
J. F.
(
1998
)
Insulin promoter factor-1 gene mutation linked to early-onset type 2 diabetes mellitus directs expression of a dominant negative isoprotein.
J Clin Invest
102
,
232
241
Tal
M.
,
Liang
Y.
,
Najafi
H.
,
Lodish
H. F.
,
Matschinsky
F. M.
(
1992
)
Expression and function of GLUT-1 and GLUT-2 glucose transporter isoforms in cells of cultured rat pancreatic islets.
J. Biol. Chem
267
,
17241
17247
Tattersall
R.
(
1998
)
Maturity-onset diabetes of the young: a clinical history.
Diabet. Med
15
,
11
14
Tattersall
R. B.
(
1974
)
Mild familial diabetes with dominant inheritance.
Q. J. Med
43
,
339
357
Teitelman
G.
,
Alpert
S.
,
Polak
J. M.
,
Martinez
A.
,
Hanahan
D.
(
1993
)
Precursor cells of mouse endocrine pancreas coexpress insulin, glucagon and the neuronal proteins tyrosine hydroxylase and neuropeptide Y, but not pancreatic polypeptide.
Development
118
,
1031
1039
Thorens
B.
,
Sarkar
H. K.
,
Kaback
H. R.
,
Lodish
H. F.
(
1988
)
Cloning and functional expression in bacteria of a novel glucose transporter present in liver, intestine, kidney, and beta-pancreatic islet cells.
Cell
55
,
281
290
Valera
A.
,
Solanes
G.
,
Fernandez-Alvarez
J.
,
Pujol
A.
,
Ferrer
J.
,
Asins
G.
,
Gomis
R.
,
Bosch
F.
(
1994
)
Expression of GLUT-2 antisense RNA incells of transgenic mice leads to diabetes.
J. Biol. Chem
269
,
28543
28546
Wang
Z.
,
Gleichmann
H.
(
1995
)
Glucose transporter 2 expression:prevention of streptozotocin-induced reduction in-cells with 5-thio-D-glucose.
Exp. Clin. Endocrinol. Diabetes
103
,
83
97
Weinstein
D. C.
,
Ruiz i Altaba
A.
,
Chen
W. S.
,
Hoodless
P.
,
Prezioso
V. R.
,
Jessell
T. M.
,
Darnell
J. E.
Jr
(
1994
)
The winged-helix transcription factor HNF-3is required for notochord development in the mouse embryo.
Cell
78
,
575
588
Wu
K. L.
,
Gannon
M.
,
Peshavaria
M.
,
Offield
M. F.
,
Henderson
E.
,
Ray
M.
,
Marks
A.
,
Gamer
L. W.
,
Wright
C. V.
,
Stein
R.
(
1997
)
Hepatocyte nuclear factor 3is involved in pancreatic beta-cell-specific transcription of the pdx-1 gene.
Mol. Cell Biol
17
,
6002
6013
Zangen
D. H.
,
Bonner-Weir
S.
,
Lee
C. H.
,
Latimer
J. B.
,
Miller
C. P.
,
Habener
J. F.
,
Weir
G. C.
(
1997
)
Reduced insulin, GLUT2, and IDX-1 in-cells after partial pancreatectomy.
Diabetes
46
,
258
264
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