TY - JOUR
T1 - The effects of PKCα phosphorylation on the extensibility of titin's PEVK element
AU - Anderson, Brian R.
AU - Bogomolovas, Julius
AU - Labeit, Siegfried
AU - Granzier, Henk
N1 - Funding Information:
We thank Carlos Hidalgo for preparation of phosphorylated samples. Brian Anderson received support from NIH training Grant GM084905 . Supported by NIH HL062881 to H.G. and by the DFG ( La668/13-1 ) and the NAR Initiative University of Heidelberg to S.L.
PY - 2010/5
Y1 - 2010/5
N2 - Post-translational modifications, along with isoform splicing, of titin determine the passive tension development of stretched sarcomeres. It was recently shown that PKCα phosphorylates two highly-conserved residues (S26 and S170) of the PEVK region in cardiac titin, resulting in passive tension increase. To determine how each phosphorylated residue affects myocardial stiffness, we generated three recombinant mutant PEVK fragments (S26A, S170A and S170A/S26A), each flanked by Ig domains. Single-molecule force spectroscopy shows that PKCα decreases the PEVK persistence length (from 0.99 to 0.68. nm); the majority of this decrease is attributable to phosphorylation of S26. Before PKCα, all three mutant PEVK fragments showed at least 40% decrease in persistence length compared to wildtype. Furthermore, Ig domain unfolding force measurements indicate that PEVK's flanking Ig domains are relatively unstable compared to other titin Ig domains. We conclude that phosphorylation of S26 is the primary mechanism through which PKCα modulates cardiac stiffness.
AB - Post-translational modifications, along with isoform splicing, of titin determine the passive tension development of stretched sarcomeres. It was recently shown that PKCα phosphorylates two highly-conserved residues (S26 and S170) of the PEVK region in cardiac titin, resulting in passive tension increase. To determine how each phosphorylated residue affects myocardial stiffness, we generated three recombinant mutant PEVK fragments (S26A, S170A and S170A/S26A), each flanked by Ig domains. Single-molecule force spectroscopy shows that PKCα decreases the PEVK persistence length (from 0.99 to 0.68. nm); the majority of this decrease is attributable to phosphorylation of S26. Before PKCα, all three mutant PEVK fragments showed at least 40% decrease in persistence length compared to wildtype. Furthermore, Ig domain unfolding force measurements indicate that PEVK's flanking Ig domains are relatively unstable compared to other titin Ig domains. We conclude that phosphorylation of S26 is the primary mechanism through which PKCα modulates cardiac stiffness.
KW - Passive tension
KW - Post-translational modifications
KW - Single molecule mechanics
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U2 - 10.1016/j.jsb.2010.02.002
DO - 10.1016/j.jsb.2010.02.002
M3 - Article
C2 - 20149875
AN - SCOPUS:77951977209
SN - 1047-8477
VL - 170
SP - 270
EP - 277
JO - Journal of Structural Biology
JF - Journal of Structural Biology
IS - 2
ER -