Molecular cloning of a heart antigen that cross-reacts with a neutralizing antibody to coxsackievirus B4

Kirk Beisel, J. Srinivasappa, B. S. Prabhakar

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

A panel of coxsackievirus B4 (CVB4) neutralizing monoclonal antibodies was tested against a panel of normal mouse tissues. One mAb, 356-1, reacted specifically with murine heart tissue. Examination of the reactivity of 356-1 with CVB4 polypeptides using Western immunoblotting revealed that 356-1 binds to the VP-1 capsid protein. Immunohistochemical studies revealed an A band pattern of staining of the heart by this antibody. Western immunoblotting of sequential differential extracts of heart showed that 356-1 predominantly reacted with the murine cardiac myosin heavy chain. A rather weak cross-reaction was found with actin. These observations were confirmed by the binding of 356-1 to purified cardiac myosin and actin. This antibody showed a higher affinity for murine cardiac muscle myosin than for skeletal muscle myosin. The monoclonal antibody 356-1 was then used to screen a λgt11 CD1 mouse heart cDNA expression library. Forty-eight positive plaques were obtained, 14 of which reacted strongly with the antibody and were selected for additional studies. In 13/14 clones, inserts were amplified using the polymerase chain reaction (PCR). The PCR products ranged size from ~ 150 to 1400 bp. Northern hybridization using these inserts demonstrated that 10/13 recognized a ~ 6.5 kb message in mouse heart total RNA and not in liver total RNA. These amplified inserts were sequenced and were found to contain sequences which encode ~ 1/3 of the amino-terminal end of light meromyosin. By comparison with known sequences of rat alpha and beta cardiac myosin heavy chains, we were able to identify sequences specific for alpha chain as potential cross-reactive autoantigenic epitope. This putative antigenic site is located between amino-acid residues 1304 and 1647 of the alpha cardiac myosin encoded by the cDNAs. These studies imply that molecular mimicry is one mechanism by which autoimmunity might develop during CVB4-induced myocarditis.

Original languageEnglish
Pages (from-to)60-64
Number of pages5
JournalEuropean Heart Journal
Volume12
Issue numberSUPPL. D
StatePublished - 1991
Externally publishedYes

Fingerprint

Enterovirus
Molecular Cloning
Neutralizing Antibodies
Cardiac Myosins
Antigens
Antibodies
Actins
Western Blotting
Monoclonal Antibodies
Skeletal Muscle Myosins
RNA
Molecular Mimicry
Viral Structural Proteins
Ventricular Myosins
Myosin Subfragments
Polymerase Chain Reaction
Myosin Heavy Chains
Cross Reactions
Myocarditis
Capsid Proteins

All Science Journal Classification (ASJC) codes

  • Cardiology and Cardiovascular Medicine

Cite this

Molecular cloning of a heart antigen that cross-reacts with a neutralizing antibody to coxsackievirus B4 . / Beisel, Kirk; Srinivasappa, J.; Prabhakar, B. S.

In: European Heart Journal, Vol. 12, No. SUPPL. D, 1991, p. 60-64.

Research output: Contribution to journalArticle

@article{6fcb3522aaff4ac7a45865e1e26b3f44,
title = "Molecular cloning of a heart antigen that cross-reacts with a neutralizing antibody to coxsackievirus B4",
abstract = "A panel of coxsackievirus B4 (CVB4) neutralizing monoclonal antibodies was tested against a panel of normal mouse tissues. One mAb, 356-1, reacted specifically with murine heart tissue. Examination of the reactivity of 356-1 with CVB4 polypeptides using Western immunoblotting revealed that 356-1 binds to the VP-1 capsid protein. Immunohistochemical studies revealed an A band pattern of staining of the heart by this antibody. Western immunoblotting of sequential differential extracts of heart showed that 356-1 predominantly reacted with the murine cardiac myosin heavy chain. A rather weak cross-reaction was found with actin. These observations were confirmed by the binding of 356-1 to purified cardiac myosin and actin. This antibody showed a higher affinity for murine cardiac muscle myosin than for skeletal muscle myosin. The monoclonal antibody 356-1 was then used to screen a λgt11 CD1 mouse heart cDNA expression library. Forty-eight positive plaques were obtained, 14 of which reacted strongly with the antibody and were selected for additional studies. In 13/14 clones, inserts were amplified using the polymerase chain reaction (PCR). The PCR products ranged size from ~ 150 to 1400 bp. Northern hybridization using these inserts demonstrated that 10/13 recognized a ~ 6.5 kb message in mouse heart total RNA and not in liver total RNA. These amplified inserts were sequenced and were found to contain sequences which encode ~ 1/3 of the amino-terminal end of light meromyosin. By comparison with known sequences of rat alpha and beta cardiac myosin heavy chains, we were able to identify sequences specific for alpha chain as potential cross-reactive autoantigenic epitope. This putative antigenic site is located between amino-acid residues 1304 and 1647 of the alpha cardiac myosin encoded by the cDNAs. These studies imply that molecular mimicry is one mechanism by which autoimmunity might develop during CVB4-induced myocarditis.",
author = "Kirk Beisel and J. Srinivasappa and Prabhakar, {B. S.}",
year = "1991",
language = "English",
volume = "12",
pages = "60--64",
journal = "European Heart Journal",
issn = "0195-668X",
publisher = "Oxford University Press",
number = "SUPPL. D",

}

TY - JOUR

T1 - Molecular cloning of a heart antigen that cross-reacts with a neutralizing antibody to coxsackievirus B4

AU - Beisel, Kirk

AU - Srinivasappa, J.

AU - Prabhakar, B. S.

PY - 1991

Y1 - 1991

N2 - A panel of coxsackievirus B4 (CVB4) neutralizing monoclonal antibodies was tested against a panel of normal mouse tissues. One mAb, 356-1, reacted specifically with murine heart tissue. Examination of the reactivity of 356-1 with CVB4 polypeptides using Western immunoblotting revealed that 356-1 binds to the VP-1 capsid protein. Immunohistochemical studies revealed an A band pattern of staining of the heart by this antibody. Western immunoblotting of sequential differential extracts of heart showed that 356-1 predominantly reacted with the murine cardiac myosin heavy chain. A rather weak cross-reaction was found with actin. These observations were confirmed by the binding of 356-1 to purified cardiac myosin and actin. This antibody showed a higher affinity for murine cardiac muscle myosin than for skeletal muscle myosin. The monoclonal antibody 356-1 was then used to screen a λgt11 CD1 mouse heart cDNA expression library. Forty-eight positive plaques were obtained, 14 of which reacted strongly with the antibody and were selected for additional studies. In 13/14 clones, inserts were amplified using the polymerase chain reaction (PCR). The PCR products ranged size from ~ 150 to 1400 bp. Northern hybridization using these inserts demonstrated that 10/13 recognized a ~ 6.5 kb message in mouse heart total RNA and not in liver total RNA. These amplified inserts were sequenced and were found to contain sequences which encode ~ 1/3 of the amino-terminal end of light meromyosin. By comparison with known sequences of rat alpha and beta cardiac myosin heavy chains, we were able to identify sequences specific for alpha chain as potential cross-reactive autoantigenic epitope. This putative antigenic site is located between amino-acid residues 1304 and 1647 of the alpha cardiac myosin encoded by the cDNAs. These studies imply that molecular mimicry is one mechanism by which autoimmunity might develop during CVB4-induced myocarditis.

AB - A panel of coxsackievirus B4 (CVB4) neutralizing monoclonal antibodies was tested against a panel of normal mouse tissues. One mAb, 356-1, reacted specifically with murine heart tissue. Examination of the reactivity of 356-1 with CVB4 polypeptides using Western immunoblotting revealed that 356-1 binds to the VP-1 capsid protein. Immunohistochemical studies revealed an A band pattern of staining of the heart by this antibody. Western immunoblotting of sequential differential extracts of heart showed that 356-1 predominantly reacted with the murine cardiac myosin heavy chain. A rather weak cross-reaction was found with actin. These observations were confirmed by the binding of 356-1 to purified cardiac myosin and actin. This antibody showed a higher affinity for murine cardiac muscle myosin than for skeletal muscle myosin. The monoclonal antibody 356-1 was then used to screen a λgt11 CD1 mouse heart cDNA expression library. Forty-eight positive plaques were obtained, 14 of which reacted strongly with the antibody and were selected for additional studies. In 13/14 clones, inserts were amplified using the polymerase chain reaction (PCR). The PCR products ranged size from ~ 150 to 1400 bp. Northern hybridization using these inserts demonstrated that 10/13 recognized a ~ 6.5 kb message in mouse heart total RNA and not in liver total RNA. These amplified inserts were sequenced and were found to contain sequences which encode ~ 1/3 of the amino-terminal end of light meromyosin. By comparison with known sequences of rat alpha and beta cardiac myosin heavy chains, we were able to identify sequences specific for alpha chain as potential cross-reactive autoantigenic epitope. This putative antigenic site is located between amino-acid residues 1304 and 1647 of the alpha cardiac myosin encoded by the cDNAs. These studies imply that molecular mimicry is one mechanism by which autoimmunity might develop during CVB4-induced myocarditis.

UR - http://www.scopus.com/inward/record.url?scp=0026049738&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=0026049738&partnerID=8YFLogxK

M3 - Article

C2 - 1717274

AN - SCOPUS:0026049738

VL - 12

SP - 60

EP - 64

JO - European Heart Journal

JF - European Heart Journal

SN - 0195-668X

IS - SUPPL. D

ER -