İskelet Kası Hücrelerinde Desmin'in Nükleer Protein Ortaklarının Araştırılması

Abstract

Desmin is a type III intermediate filament protein involved in myofibrillar alignment and force transmission in muscle cells. Its presence in the nucleus suggests that it may participate in intranuclear processes in addition to its structural functions. This thesis aimed to characterize the nuclear interaction network of Desmin in skeletal muscle cells and investigate its potential association with chromatin organization. Human LHCN-M2 and mouse C2C12 cells were analyzed using RNA-sequencing data analysis, polymerase chain reaction, cellular fractionation, Western blotting, immunoprecipitation, and mass spectrometry. Because targeted approaches investigating the associations of Desmin with Nkx2.5 and SMYD1 yielded limited results, the study was redirected toward proteomic characterization of the nuclear Desmin interaction network. Desmin was confirmed to be present in the nuclear fraction and to co-precipitate with nuclear protein partners. In immunoprecipitation–mass spectrometry analysis, Desmin and Vimentin were enriched 47.5-fold and 159-fold, respectively. Candidate proteins were evaluated using a composite score integrating log₂FC, statistical significance, peptide-spectrum matches, unique peptide count, IP/IgG ratio, and evidence from independent datasets; TMPO, RANBP2, HNRNPU, NUP153, DDX5, and EEF1A1 were prioritized. Reciprocal IP–WB analyses demonstrated the co-precipitation of TMPO and DDX5 with Desmin, supporting their possible presence within the same protein complexes. The association of these candidates with the nuclear envelope, nuclear pore complex, and RNA metabolism suggests that Desmin may participate in intranuclear protein networks. RNase A treatment produced no statistically significant changes; however, EEF1A1 and HNRNPA2B1 showed decreasing trends consistent with possible RNA-mediated associations. Overall, this study characterized the nuclear interaction network of Desmin and provided evidence supporting its association with nuclear architecture, nucleocytoplasmic transport, and RNA-related protein networks.