Összes szerző

Mártonfalvi Zsolt

az alábbi absztraktok szerzői között szerepel:

Balog-Molnár Andrea
Phosporylation alters the structure of surface bound titin molecules

Aug 29 - kedd

09:45 – 10:00

Molekuláris biofizika

E05

Phosporylation alters the structure of surface bound titin molecules

Andrea Balogh-Molnár, Zsolt Mártonfalvi

Semmelweis University, Faculty of Medicine, Department of Biophysics and Radiation Biology

During muscle stretch, elastic or "passive" force develops which is mainly determined by the giant protein titin that forms the third filament system of muscle sarcomeres. The magnitude of this force is mainly dependent on the elasticity of the titin filaments, which is primarily determined by the structure of the polypeptide chain. However, it is suggested that post-translational modifications of titin, such as phosphokinase activity, regulate the sarcomeric passive force development. Mechanical studies on single myofibrils revealed that different protein kinases alter the passive tension of muscle, surprisingly, in opposite ways. This suggests that phosphorylation of sarcomeric proteins by various kinases is an essential regulatory mechanism of passive force. However, the extrapolation of these findings to titin’s phosphorylation has not been studied so far at the single- molecule level. To reveal titin’s structural alterations due to phosphorylation, single-molecule experiments must be carried out on individual titin molecules, where the effect of phosphorylation can be tested individually. In our work, we isolated individual titin molecules from rabbit and mouse skeletal muscle. The isolated native titin molecules showed high levels of phosphorylation, when stained with phosphoprotein gel stain. The isolated molecules were treated with lambda protein phosphatase to decrease the in situ phosphorylation level of the polymer. To investigate the effect of the treatment on titin's structure, we visualized surface-bound titin molecules by atomic force microscope and found that the terminal regions of dephosphorylated titins collapse into a compact, coiled structure. Our findings suggest that the structure of both I-band and terminal A-band region of titin is highly sensitive for phospohrylation which might play a role in controlling protein-titin interactions.

Balogh Anna
The FINmaj mutation results in hypophosporilation of the sarcomeric titin molecules

Aug 29 - kedd

15:30 – 17:00

I. Poszterszekció

P02

The FINmaj mutation results in hypophosporilation of the sarcomeric titin molecules

Anna Balogh, Andrea Balogh-Molnár, Zsolt Mártonfalvi

Semmelweis University, Faculty of Medicine, Department of Biophysics and Radiation Biology

The giant sarcomeric protein titin has central mechanistic and developmental roles in striated muscle. Mutations in the TTN gene cause a wide spectrum of skeletal and cardiac myopathies. The dominant tibial muscular dystrophy (TMD) and recessive limb griddle muscular dystrophy (LGMD2J) in Finnish patients are caused by the FINmaj mutation, changing four amino acids (EVTW > VKEK) in the most C-terminal Ig-domain M10 of titin, in the sarcomeric M-line, which domain harbors binding sites for obscurin/obscurin-like 1 and myospryn. Overall, the FINmaj induces a pathological cleavage and degradation of a 70–80 kDa portion from the titin C-terminus, soon after its synthesis.

To investigate the effect of the mutation on the sarcomeric structure of titin we isolated titin molecules from FINmaj murine skeletal muscle samples and from skeletal muscle biopsies of TMD/LGMD2J patients homozygous for the FINmaj mutation. The isolated titin molecules were visualized using atomic force microscopy. We found that the isolated titin molecules maintained their oligomeric structures, in which the overlapping C-termini of titins from neighboring sarcomeres remain bound. The oligomers showed a collapsed structure with an M‑band complex that both increased in area and topographical height. Overall, the appearance of the FINmaj oligomers resembled the structure of hypophosphorylated titin molecules.

Our findings suggest that the FINmaj mutation does not destruct the M-band complex in titin oligomers but decreases the level of phosphorylation in the A-band segment of titin.

Acknowledgment

We thank for the FINmaj samples for prof. Udd Bjarne at University of Helsinki

Sulea Cristina
Fibrillin-1 microfibrils in Marfan syndrome: nanoscale structural characterization using atomic force microscopy

Aug 29 - kedd

15:30 – 17:00

I. Poszterszekció

P27

Fibrillin-1 microfibrils in Marfan syndrome: nanoscale structural characterization using atomic force microscopy

Cristina M. Șulea1,2,3, Zsolt Mártonfalvi1, Csilla Csányi1, Dóra Haluszka1, Miklós Pólos2,3, Kálmán Benke2,3, Zoltán Szabolcs2,3 and Miklós S. Z. Kellermayer1

1 Department of Biophysics and Radiation Biology, Semmelweis University, 1094 Budapest, Hungary

2 Heart and Vascular Center, Semmelweis University, 1122 Budapest, Hungary

3 Hungarian Marfan Foundation, 1122 Budapest, Hungary

Fibrillin-1 microfibrils are essential elements of the extracellular matrix serving as a scaffold for the deposition of elastin and endowing connective tissues with tensile strength and elasticity. Mutations in the fibrillin-1 gene (FBN1) are linked to Marfan syndrome (MFS), a systemic connective tissue disorder that usually manifests in life-threatening aortic complications. The aortic involvement may be explained by a dysregulation in microfibrillar function and, conceivably, alterations in the microfibrils’ supramolecular structure.

The aim of the study was to perform a nanoscale structural characterization of fibrillin-1 microfibrils isolated from human aortic samples with different FBN1 gene mutations and to compare them with microfibrillar assemblies purified from non-MFS human aortic tissue.

Aortic wall samples were obtained from patients undergoing specific cardiovascular surgical interventions. Fibrillin-rich microfibrils were extracted by bacterial collagenase digestion and purified by size-exclusion chromatography. Atomic force microscopy was employed to visualize and study the microfibrillar assemblies.

Fibrillin-1 microfibrils displayed a characteristic “beads-on-a-string” appearance. The microfibrillar assemblies were investigated for bead geometry (height, length, and width), interbead region height, and periodicity. MFS fibrillin-1 microfibrils had a slightly higher mean bead height, but the bead length and width, as well as the interbead height, were significantly smaller in the MFS group. The mean periodicity varied around 50–52 nm among samples.

In conclusion, the data suggest an overall thinner and presumably more frail structure for the MFS fibrillin-1 microfibrils, which may play a role in the development of MFS-specific aortic symptomatology.

Acknowledgment

Funding sources: NRDI Office (ÚNKP-22-3-I-SE-49 to C.M.Ș.; K135360 to M.S.Z.K.; TKP2021-EGA-23), European Union (RRF-2.3.1-21-2022-00003 – National Cardiovascular Laboratory).