Mouse models of two missense mutations in actin-binding domain 1 of dystrophin associated with duchenne or becker muscular dystrophy

Jackie L. McCourt, Dana M Talsness, Angus Lindsay, Robert W Arpke, Paul D. Chatterton, D'anna M. Nelson, Christopher M Chamberlain, John T. Olthoff, Joseph J Belanto, Preston M. McCourt, Michael Kyba, Dawn A Lowe, James M Ervasti

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

Missense mutations in the dystrophin protein can cause Duchenne muscular dystrophy (DMD) or Becker muscular dystrophy (BMD) through an undefined pathomechanism. In vitro studies suggest that missense mutations in the N-terminal actinbinding domain (ABD1) cause protein instability, and cultured myoblast studies reveal decreased expression levels that can be restored to wild-type with proteasome inhibitors. To further elucidate the pathophysiology of missense dystrophin in vivo, we generated two transgenic mdx mouse lines expressing L54R or L172H mutant dystrophin, which correspond to missense mutations identified in human patients with DMD or BMD, respectively. Our biochemical, histologic and physiologic analysis of the L54R and L172H mice show decreased levels of dystrophin which are proportional to the phenotypic severity. Proteasome inhibitors were ineffective in both the L54R and L172H mice, yet mice homozygous for the L172H transgene were able to express even higher levels of dystrophin which caused further improvements in muscle histology and physiology. Given that missense dystrophin is likely being degraded by the proteasome but whole body proteasome inhibition was not possible, we screened for ubiquitin-conjugating enzymes involved in targeting dystrophin to the proteasome. A myoblast cell line expressing L54R mutant dystrophin was screened with an siRNA library targeting E1, E2 and E3 ligases which identified Amn1, FBXO33, Zfand5 and Trim75. Our study establishes new mouse models of dystrophinopathy and identifies candidate E3 ligases that may specifically regulate dystrophin protein turnover in vivo.

Original languageEnglish (US)
Pages (from-to)451-462
Number of pages12
JournalHuman molecular genetics
Volume27
Issue number3
DOIs
StatePublished - Feb 1 2018

Fingerprint

Dystrophin
Duchenne Muscular Dystrophy
Missense Mutation
Actins
Proteasome Endopeptidase Complex
Proteasome Inhibitors
Ubiquitin-Protein Ligases
Myoblasts
Ubiquitin-Conjugating Enzymes
Inbred mdx Mouse
Proteins
Transgenes
Small Interfering RNA
Transgenic Mice
Histology
Cell Line
Muscles

PubMed: MeSH publication types

  • Journal Article
  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

Cite this

Mouse models of two missense mutations in actin-binding domain 1 of dystrophin associated with duchenne or becker muscular dystrophy. / McCourt, Jackie L.; Talsness, Dana M; Lindsay, Angus; Arpke, Robert W; Chatterton, Paul D.; Nelson, D'anna M.; Chamberlain, Christopher M; Olthoff, John T.; Belanto, Joseph J; McCourt, Preston M.; Kyba, Michael; Lowe, Dawn A; Ervasti, James M.

In: Human molecular genetics, Vol. 27, No. 3, 01.02.2018, p. 451-462.

Research output: Contribution to journalArticle

McCourt, JL, Talsness, DM, Lindsay, A, Arpke, RW, Chatterton, PD, Nelson, DM, Chamberlain, CM, Olthoff, JT, Belanto, JJ, McCourt, PM, Kyba, M, Lowe, DA & Ervasti, JM 2018, 'Mouse models of two missense mutations in actin-binding domain 1 of dystrophin associated with duchenne or becker muscular dystrophy', Human molecular genetics, vol. 27, no. 3, pp. 451-462. https://doi.org/10.1093/hmg/ddx414
McCourt, Jackie L. ; Talsness, Dana M ; Lindsay, Angus ; Arpke, Robert W ; Chatterton, Paul D. ; Nelson, D'anna M. ; Chamberlain, Christopher M ; Olthoff, John T. ; Belanto, Joseph J ; McCourt, Preston M. ; Kyba, Michael ; Lowe, Dawn A ; Ervasti, James M. / Mouse models of two missense mutations in actin-binding domain 1 of dystrophin associated with duchenne or becker muscular dystrophy. In: Human molecular genetics. 2018 ; Vol. 27, No. 3. pp. 451-462.
@article{109eccafb5734fc8bec09f8991d0791d,
title = "Mouse models of two missense mutations in actin-binding domain 1 of dystrophin associated with duchenne or becker muscular dystrophy",
abstract = "Missense mutations in the dystrophin protein can cause Duchenne muscular dystrophy (DMD) or Becker muscular dystrophy (BMD) through an undefined pathomechanism. In vitro studies suggest that missense mutations in the N-terminal actinbinding domain (ABD1) cause protein instability, and cultured myoblast studies reveal decreased expression levels that can be restored to wild-type with proteasome inhibitors. To further elucidate the pathophysiology of missense dystrophin in vivo, we generated two transgenic mdx mouse lines expressing L54R or L172H mutant dystrophin, which correspond to missense mutations identified in human patients with DMD or BMD, respectively. Our biochemical, histologic and physiologic analysis of the L54R and L172H mice show decreased levels of dystrophin which are proportional to the phenotypic severity. Proteasome inhibitors were ineffective in both the L54R and L172H mice, yet mice homozygous for the L172H transgene were able to express even higher levels of dystrophin which caused further improvements in muscle histology and physiology. Given that missense dystrophin is likely being degraded by the proteasome but whole body proteasome inhibition was not possible, we screened for ubiquitin-conjugating enzymes involved in targeting dystrophin to the proteasome. A myoblast cell line expressing L54R mutant dystrophin was screened with an siRNA library targeting E1, E2 and E3 ligases which identified Amn1, FBXO33, Zfand5 and Trim75. Our study establishes new mouse models of dystrophinopathy and identifies candidate E3 ligases that may specifically regulate dystrophin protein turnover in vivo.",
author = "McCourt, {Jackie L.} and Talsness, {Dana M} and Angus Lindsay and Arpke, {Robert W} and Chatterton, {Paul D.} and Nelson, {D'anna M.} and Chamberlain, {Christopher M} and Olthoff, {John T.} and Belanto, {Joseph J} and McCourt, {Preston M.} and Michael Kyba and Lowe, {Dawn A} and Ervasti, {James M}",
year = "2018",
month = "2",
day = "1",
doi = "10.1093/hmg/ddx414",
language = "English (US)",
volume = "27",
pages = "451--462",
journal = "Human Molecular Genetics",
issn = "0964-6906",
publisher = "Oxford University Press",
number = "3",

}

TY - JOUR

T1 - Mouse models of two missense mutations in actin-binding domain 1 of dystrophin associated with duchenne or becker muscular dystrophy

AU - McCourt, Jackie L.

AU - Talsness, Dana M

AU - Lindsay, Angus

AU - Arpke, Robert W

AU - Chatterton, Paul D.

AU - Nelson, D'anna M.

AU - Chamberlain, Christopher M

AU - Olthoff, John T.

AU - Belanto, Joseph J

AU - McCourt, Preston M.

AU - Kyba, Michael

AU - Lowe, Dawn A

AU - Ervasti, James M

PY - 2018/2/1

Y1 - 2018/2/1

N2 - Missense mutations in the dystrophin protein can cause Duchenne muscular dystrophy (DMD) or Becker muscular dystrophy (BMD) through an undefined pathomechanism. In vitro studies suggest that missense mutations in the N-terminal actinbinding domain (ABD1) cause protein instability, and cultured myoblast studies reveal decreased expression levels that can be restored to wild-type with proteasome inhibitors. To further elucidate the pathophysiology of missense dystrophin in vivo, we generated two transgenic mdx mouse lines expressing L54R or L172H mutant dystrophin, which correspond to missense mutations identified in human patients with DMD or BMD, respectively. Our biochemical, histologic and physiologic analysis of the L54R and L172H mice show decreased levels of dystrophin which are proportional to the phenotypic severity. Proteasome inhibitors were ineffective in both the L54R and L172H mice, yet mice homozygous for the L172H transgene were able to express even higher levels of dystrophin which caused further improvements in muscle histology and physiology. Given that missense dystrophin is likely being degraded by the proteasome but whole body proteasome inhibition was not possible, we screened for ubiquitin-conjugating enzymes involved in targeting dystrophin to the proteasome. A myoblast cell line expressing L54R mutant dystrophin was screened with an siRNA library targeting E1, E2 and E3 ligases which identified Amn1, FBXO33, Zfand5 and Trim75. Our study establishes new mouse models of dystrophinopathy and identifies candidate E3 ligases that may specifically regulate dystrophin protein turnover in vivo.

AB - Missense mutations in the dystrophin protein can cause Duchenne muscular dystrophy (DMD) or Becker muscular dystrophy (BMD) through an undefined pathomechanism. In vitro studies suggest that missense mutations in the N-terminal actinbinding domain (ABD1) cause protein instability, and cultured myoblast studies reveal decreased expression levels that can be restored to wild-type with proteasome inhibitors. To further elucidate the pathophysiology of missense dystrophin in vivo, we generated two transgenic mdx mouse lines expressing L54R or L172H mutant dystrophin, which correspond to missense mutations identified in human patients with DMD or BMD, respectively. Our biochemical, histologic and physiologic analysis of the L54R and L172H mice show decreased levels of dystrophin which are proportional to the phenotypic severity. Proteasome inhibitors were ineffective in both the L54R and L172H mice, yet mice homozygous for the L172H transgene were able to express even higher levels of dystrophin which caused further improvements in muscle histology and physiology. Given that missense dystrophin is likely being degraded by the proteasome but whole body proteasome inhibition was not possible, we screened for ubiquitin-conjugating enzymes involved in targeting dystrophin to the proteasome. A myoblast cell line expressing L54R mutant dystrophin was screened with an siRNA library targeting E1, E2 and E3 ligases which identified Amn1, FBXO33, Zfand5 and Trim75. Our study establishes new mouse models of dystrophinopathy and identifies candidate E3 ligases that may specifically regulate dystrophin protein turnover in vivo.

UR - http://www.scopus.com/inward/record.url?scp=85042617108&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=85042617108&partnerID=8YFLogxK

U2 - 10.1093/hmg/ddx414

DO - 10.1093/hmg/ddx414

M3 - Article

C2 - 29194514

AN - SCOPUS:85042617108

VL - 27

SP - 451

EP - 462

JO - Human Molecular Genetics

JF - Human Molecular Genetics

SN - 0964-6906

IS - 3

ER -