2024-03-28T10:28:38Zhttp://repisalud.isciii.es/oai/requestoai:repisalud.isciii.es:20.500.12105/103212022-10-06T11:20:07Zcom_20.500.12105_2145com_20.500.12105_2051com_20.500.12105_2144col_20.500.12105_2146
Repisalud
author
Rivas-Pardo, Jaime Andrés
author
Li, Yong
author
Mártonfalvi, Zsolt
author
Tapia-Rojo, Rafael
author
Unger, Andreas
author
Fernández-Trasancos, Ángel
author
Herrero-Galan, Elas
author
Velazquez-Carreras, Diana
author
Fernández, Julio M
author
Linke, Wolfgang A
author
Alegre-Cebollada, Jorge
funder
Ministerio de Ciencia e Innovación (España)
funder
Comunidad de Madrid (España)
funder
Instituto de Salud Carlos III
funder
Fundación ProCNIC
funder
Deutsche Forschungsgemeinschaft (Alemania)
funder
Hungarian Academy of Sciences
funder
European Molecular Biology Organization
funder
Boehringer Ingelheim Fonds
2020-06-10T09:54:47Z
2020-06-10T09:54:47Z
2020-04-28
Nat Commun. 2020; 11(1):2060
http://hdl.handle.net/20.500.12105/10321
32345978
10.1038/s41467-020-15465-9
2041-1723
Nature communications
Single-molecule methods using recombinant proteins have generated transformative hypotheses on how mechanical forces are generated and sensed in biological tissues. However, testing these mechanical hypotheses on proteins in their natural environment remains inaccesible to conventional tools. To address this limitation, here we demonstrate a mouse model carrying a HaloTag-TEV insertion in the protein titin, the main determinant of myocyte stiffness. Using our system, we specifically sever titin by digestion with TEV protease, and find that the response of muscle fibers to length changes requires mechanical transduction through titin's intact polypeptide chain. In addition, HaloTag-based covalent tethering enables examination of titin dynamics under force using magnetic tweezers. At pulling forces < 10 pN, titin domains are recruited to the unfolded state, and produce 41.5 zJ mechanical work during refolding. Insertion of the HaloTag-TEV cassette in mechanical proteins opens opportunities to explore the molecular basis of cellular force generation, mechanosensing and mechanotransduction.
eng
A HaloTag-TEV genetic cassette for mechanical phenotyping of proteins from tissues.
journal article
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
URL
https://repisalud.isciii.es/bitstream/20.500.12105/10321/1/AHaloTag-TEVGeneticCassette_2020.pdf
File
MD5
c22d945c9b60062e9fdcd42895f66f22
4497001
application/pdf
AHaloTag-TEVGeneticCassette_2020.pdf
URL
https://repisalud.isciii.es/bitstream/20.500.12105/10321/2/AHaloTag-TEVGeneticCassette_2020_MOESM1_ESM.mpg
File
MD5
5c12e42931d7b89fc67a756469c6bd9e
4261888
video/mpeg
AHaloTag-TEVGeneticCassette_2020_MOESM1_ESM.mpg
URL
https://repisalud.isciii.es/bitstream/20.500.12105/10321/3/AHaloTag-TEVGeneticCassette_2020_MOESM2_ESM.pdf
File
MD5
53b3a13275d7d473b71e4c06794cde65
1909349
application/pdf
AHaloTag-TEVGeneticCassette_2020_MOESM2_ESM.pdf
URL
https://repisalud.isciii.es/bitstream/20.500.12105/10321/4/AHaloTag-TEVGeneticCassette_2020_MOESM3_ESM.pdf
File
MD5
1bd6c54673e8031b451b01aa86adcd09
78251
application/pdf
AHaloTag-TEVGeneticCassette_2020_MOESM3_ESM.pdf
URL
https://repisalud.isciii.es/bitstream/20.500.12105/10321/5/AHaloTag-TEVGeneticCassette_2020_MOESM4_ESM.pdf
File
MD5
0382aaeffe35213157cec715c6569dc6
258406
application/pdf
AHaloTag-TEVGeneticCassette_2020_MOESM4_ESM.pdf
URL
https://repisalud.isciii.es/bitstream/20.500.12105/10321/6/AHaloTag-TEVGeneticCassette_2020_MOESM5_ESM.pdf
File
MD5
dfac4eae3babf869993a54e40e6fdb13
787386
application/pdf
AHaloTag-TEVGeneticCassette_2020_MOESM5_ESM.pdf
URL
https://repisalud.isciii.es/bitstream/20.500.12105/10321/7/AHaloTag-TEVGeneticCassette_2020_MOESM6_ESM.xlsx
File
MD5
40a7dbf775e27a0d7dff8bcdaff38153
69464
application/vnd.openxmlformats-officedocument.spreadsheetml.sheet
AHaloTag-TEVGeneticCassette_2020_MOESM6_ESM.xlsx
URL
https://repisalud.isciii.es/bitstream/20.500.12105/10321/20/AHaloTag-TEVGeneticCassette_2020.pdf.txt
File
MD5
f3134005f530fdcda080f38c6d142339
82937
text/plain
AHaloTag-TEVGeneticCassette_2020.pdf.txt
URL
https://repisalud.isciii.es/bitstream/20.500.12105/10321/22/AHaloTag-TEVGeneticCassette_2020_MOESM2_ESM.pdf.txt
File
MD5
7aee66846ae50667df2932653ccade9d
26901
text/plain
AHaloTag-TEVGeneticCassette_2020_MOESM2_ESM.pdf.txt
URL
https://repisalud.isciii.es/bitstream/20.500.12105/10321/24/AHaloTag-TEVGeneticCassette_2020_MOESM3_ESM.pdf.txt
File
MD5
1233307d437a642d21a5271a13b4b060
402
text/plain
AHaloTag-TEVGeneticCassette_2020_MOESM3_ESM.pdf.txt
URL
https://repisalud.isciii.es/bitstream/20.500.12105/10321/26/AHaloTag-TEVGeneticCassette_2020_MOESM4_ESM.pdf.txt
File
MD5
9c07c94ecc1874cf8f7a364f031769d6
3519
text/plain
AHaloTag-TEVGeneticCassette_2020_MOESM4_ESM.pdf.txt
URL
https://repisalud.isciii.es/bitstream/20.500.12105/10321/28/AHaloTag-TEVGeneticCassette_2020_MOESM5_ESM.pdf.txt
File
MD5
001ff063a5d840f9f0920672fd4ee96e
57005
text/plain
AHaloTag-TEVGeneticCassette_2020_MOESM5_ESM.pdf.txt