Monoclonal antibody 7F9 recognizes rat protein homologous to human carboxypeptidase-M in developing and adult rat lung.
OBJECTIVE
The purpose of this study was to obtain an antibody that would be useful for investigating the yet unclear molecular mechanism underlying the differentiation of lung alveolar type I and II cells.


METHODS
Monoclonal antibodies were raised against membrane proteins from embryonal day 18.5 rat lungs and characterized by immunoblotting on rat lung lysates at various developmental stages to select an appropriate clone. The antigen of the selected antibody was purified by serial column chromatography and immunoprecipitation and identified by mass spectrometry.


RESULTS
7F9 antibody recognizes a 65-kDa protein that is expressed most prominently from embryonal day 20.5 to postnatal day 1. This protein was identified as a rat protein that is similar to 5730456K23Rik protein. The protein is homologous to human carboxypeptidase-M. Although human carboxypeptidase-M is known as a marker of type I cells, the expression of this rat protein was detected in columnar epithelial cells expressing type II cell markers, SP-C and a lamellar body protein ABCA3, in developing lung. Its expression was detected in alveolar cells lacking T1alpha, a type I cell marker protein, in adult lung. It was also expressed in RLE-6TN cells derived from type II cells. The expression in RLE-6TN cells was down-regulated by transforming growth factor-beta1 and up-regulated by Wnt3a.


CONCLUSIONS
7F9 antibody detects a protein in rat lung cells expressing type II markers. The antibody is a useful tool for studying signalling triggered by transforming growth factor-beta1 and Wnt3a in rat type II cells.
Fujiwara
Naoyuki
N
Department of Medical Biochemistry, Graduate School of Medicine, Tokyo Medical and Dental University, Tokyo, Japan.
Ikeda
Mitsunobu
M
Hirabayashi
Susumu
S
Mori
Hiroki
H
Shirasawa
Madoka
M
Kansaku
Ai
A
Sunamori
Makoto
M
Hata
Yutaka
Y
D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology*; D058851:GPI-Linked Proteins; D018507:Gene Expression Regulation, Developmental / Q000502:physiology; D006801:Humans; D015151:Immunoblotting; D007150:Immunohistochemistry; D008168:Lung / Q000254:growth & development* / Q000276:immunology / Q000378:metabolism; D013058:Mass Spectrometry; D008666:Metalloendopeptidases / Q000276:immunology*; D051381:Rats; D053773:Transforming Growth Factor beta1 / Q000096:biosynthesis / Q000235:genetics; D051153:Wnt Proteins / Q000096:biosynthesis / Q000235:genetics; D060508:Wnt3 Protein; D060509:Wnt3A Protein
D000911:Antibodies, Monoclonal; D058851:GPI-Linked Proteins; D053773:Transforming Growth Factor beta1; C555510:WNT3A protein, human; D051153:Wnt Proteins; D060508:Wnt3 Protein; D060509:Wnt3A Protein; C058479:carboxypeptidase M; D008666:Metalloendopeptidases