A Betacellulin Mutant Promotes Differentiation of Pancreatic Acinar AR42J Cells into Insulin-Producing Cells with Low Affinity of Binding to ErbB1

Tadahiro Nagaoka, Takayuki Fukuda, Toshihiro Hashizume, Tomoko Nishiyama, Hiroko Tada, Hidenori Yamada, David S. Salomon, Satoko Yamada, Itaru Kojima, Masaharu Seno

Research output: Contribution to journalArticle

10 Citations (Scopus)

Abstract

Betacellulin (BTC) is one of the members of the epidermal growth factor (EGF) ligand family of ErbB receptor tyrosine kinases. It is a differentiation factor as well as a potent mitogen. BTC promotes the differentiation of pancreatic acinar-derived AR42J cells into insulin-producing cells. It independently and preferentially binds to two type I tyrosine kinase receptors, the EGF receptor (ErbB1) and ErbB4. However, the physiochemical characteristics of BTC that are responsible for its preferential binding to these two receptors have not been fully defined. In this study, to investigate the essential amino acid residues of BTC for binding to the two receptors, we introduced point mutations into the EGF domain of BTC employing error-prone PCR. The receptor binding abilities of 190 mutants expressed in Escherichia coli were assessed by enzyme immunoassay. Replacement of the glutamic acid residue at position 88 with a lysine residue in BTC was found to produce a significant loss of affinity for binding to ErbB1, while the affinity of binding to ErbB4 was unchanged. In addition, the mutant of BTC-E/88/K showed less growth-promoting activity on BALB/c 3T3 cells compared with that of the wild-type BTC protein. Interestingly, the BTC mutant protein promoted differentiation of pancreatic acinar AR42J cells at a high frequency into insulin-producing cells compared with AR42J cells that were treated with wild-type BTC protein. These results indicate the possibility of designing BTC mutants, which have an activity of inducing differentiation only, without facilitating growth promotion.

Original languageEnglish
Pages (from-to)83-94
Number of pages12
JournalJournal of Molecular Biology
Volume380
Issue number1
DOIs
Publication statusPublished - Jun 27 2008

Fingerprint

Acinar Cells
Insulin
Epidermal Growth Factor
Betacellulin
BALB 3T3 Cells
Essential Amino Acids
Receptor Protein-Tyrosine Kinases
Mutant Proteins
Immunoenzyme Techniques
Mitogens
Point Mutation
Epidermal Growth Factor Receptor
Protein-Tyrosine Kinases
Lysine
Glutamic Acid
Proteins
Escherichia coli
Ligands

Keywords

  • betacellulin
  • cell differentiation
  • ErbB1
  • ErbB4
  • receptor-ligand interaction

ASJC Scopus subject areas

  • Virology

Cite this

A Betacellulin Mutant Promotes Differentiation of Pancreatic Acinar AR42J Cells into Insulin-Producing Cells with Low Affinity of Binding to ErbB1. / Nagaoka, Tadahiro; Fukuda, Takayuki; Hashizume, Toshihiro; Nishiyama, Tomoko; Tada, Hiroko; Yamada, Hidenori; Salomon, David S.; Yamada, Satoko; Kojima, Itaru; Seno, Masaharu.

In: Journal of Molecular Biology, Vol. 380, No. 1, 27.06.2008, p. 83-94.

Research output: Contribution to journalArticle

Nagaoka, Tadahiro ; Fukuda, Takayuki ; Hashizume, Toshihiro ; Nishiyama, Tomoko ; Tada, Hiroko ; Yamada, Hidenori ; Salomon, David S. ; Yamada, Satoko ; Kojima, Itaru ; Seno, Masaharu. / A Betacellulin Mutant Promotes Differentiation of Pancreatic Acinar AR42J Cells into Insulin-Producing Cells with Low Affinity of Binding to ErbB1. In: Journal of Molecular Biology. 2008 ; Vol. 380, No. 1. pp. 83-94.
@article{ec4d61bc813a48fd9833e00d198e4ea3,
title = "A Betacellulin Mutant Promotes Differentiation of Pancreatic Acinar AR42J Cells into Insulin-Producing Cells with Low Affinity of Binding to ErbB1",
abstract = "Betacellulin (BTC) is one of the members of the epidermal growth factor (EGF) ligand family of ErbB receptor tyrosine kinases. It is a differentiation factor as well as a potent mitogen. BTC promotes the differentiation of pancreatic acinar-derived AR42J cells into insulin-producing cells. It independently and preferentially binds to two type I tyrosine kinase receptors, the EGF receptor (ErbB1) and ErbB4. However, the physiochemical characteristics of BTC that are responsible for its preferential binding to these two receptors have not been fully defined. In this study, to investigate the essential amino acid residues of BTC for binding to the two receptors, we introduced point mutations into the EGF domain of BTC employing error-prone PCR. The receptor binding abilities of 190 mutants expressed in Escherichia coli were assessed by enzyme immunoassay. Replacement of the glutamic acid residue at position 88 with a lysine residue in BTC was found to produce a significant loss of affinity for binding to ErbB1, while the affinity of binding to ErbB4 was unchanged. In addition, the mutant of BTC-E/88/K showed less growth-promoting activity on BALB/c 3T3 cells compared with that of the wild-type BTC protein. Interestingly, the BTC mutant protein promoted differentiation of pancreatic acinar AR42J cells at a high frequency into insulin-producing cells compared with AR42J cells that were treated with wild-type BTC protein. These results indicate the possibility of designing BTC mutants, which have an activity of inducing differentiation only, without facilitating growth promotion.",
keywords = "betacellulin, cell differentiation, ErbB1, ErbB4, receptor-ligand interaction",
author = "Tadahiro Nagaoka and Takayuki Fukuda and Toshihiro Hashizume and Tomoko Nishiyama and Hiroko Tada and Hidenori Yamada and Salomon, {David S.} and Satoko Yamada and Itaru Kojima and Masaharu Seno",
year = "2008",
month = "6",
day = "27",
doi = "10.1016/j.jmb.2008.03.054",
language = "English",
volume = "380",
pages = "83--94",
journal = "Journal of Molecular Biology",
issn = "0022-2836",
publisher = "Academic Press Inc.",
number = "1",

}

TY - JOUR

T1 - A Betacellulin Mutant Promotes Differentiation of Pancreatic Acinar AR42J Cells into Insulin-Producing Cells with Low Affinity of Binding to ErbB1

AU - Nagaoka, Tadahiro

AU - Fukuda, Takayuki

AU - Hashizume, Toshihiro

AU - Nishiyama, Tomoko

AU - Tada, Hiroko

AU - Yamada, Hidenori

AU - Salomon, David S.

AU - Yamada, Satoko

AU - Kojima, Itaru

AU - Seno, Masaharu

PY - 2008/6/27

Y1 - 2008/6/27

N2 - Betacellulin (BTC) is one of the members of the epidermal growth factor (EGF) ligand family of ErbB receptor tyrosine kinases. It is a differentiation factor as well as a potent mitogen. BTC promotes the differentiation of pancreatic acinar-derived AR42J cells into insulin-producing cells. It independently and preferentially binds to two type I tyrosine kinase receptors, the EGF receptor (ErbB1) and ErbB4. However, the physiochemical characteristics of BTC that are responsible for its preferential binding to these two receptors have not been fully defined. In this study, to investigate the essential amino acid residues of BTC for binding to the two receptors, we introduced point mutations into the EGF domain of BTC employing error-prone PCR. The receptor binding abilities of 190 mutants expressed in Escherichia coli were assessed by enzyme immunoassay. Replacement of the glutamic acid residue at position 88 with a lysine residue in BTC was found to produce a significant loss of affinity for binding to ErbB1, while the affinity of binding to ErbB4 was unchanged. In addition, the mutant of BTC-E/88/K showed less growth-promoting activity on BALB/c 3T3 cells compared with that of the wild-type BTC protein. Interestingly, the BTC mutant protein promoted differentiation of pancreatic acinar AR42J cells at a high frequency into insulin-producing cells compared with AR42J cells that were treated with wild-type BTC protein. These results indicate the possibility of designing BTC mutants, which have an activity of inducing differentiation only, without facilitating growth promotion.

AB - Betacellulin (BTC) is one of the members of the epidermal growth factor (EGF) ligand family of ErbB receptor tyrosine kinases. It is a differentiation factor as well as a potent mitogen. BTC promotes the differentiation of pancreatic acinar-derived AR42J cells into insulin-producing cells. It independently and preferentially binds to two type I tyrosine kinase receptors, the EGF receptor (ErbB1) and ErbB4. However, the physiochemical characteristics of BTC that are responsible for its preferential binding to these two receptors have not been fully defined. In this study, to investigate the essential amino acid residues of BTC for binding to the two receptors, we introduced point mutations into the EGF domain of BTC employing error-prone PCR. The receptor binding abilities of 190 mutants expressed in Escherichia coli were assessed by enzyme immunoassay. Replacement of the glutamic acid residue at position 88 with a lysine residue in BTC was found to produce a significant loss of affinity for binding to ErbB1, while the affinity of binding to ErbB4 was unchanged. In addition, the mutant of BTC-E/88/K showed less growth-promoting activity on BALB/c 3T3 cells compared with that of the wild-type BTC protein. Interestingly, the BTC mutant protein promoted differentiation of pancreatic acinar AR42J cells at a high frequency into insulin-producing cells compared with AR42J cells that were treated with wild-type BTC protein. These results indicate the possibility of designing BTC mutants, which have an activity of inducing differentiation only, without facilitating growth promotion.

KW - betacellulin

KW - cell differentiation

KW - ErbB1

KW - ErbB4

KW - receptor-ligand interaction

UR - http://www.scopus.com/inward/record.url?scp=44649143126&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=44649143126&partnerID=8YFLogxK

U2 - 10.1016/j.jmb.2008.03.054

DO - 10.1016/j.jmb.2008.03.054

M3 - Article

C2 - 18508082

AN - SCOPUS:44649143126

VL - 380

SP - 83

EP - 94

JO - Journal of Molecular Biology

JF - Journal of Molecular Biology

SN - 0022-2836

IS - 1

ER -