[Establishment of a New-generation High-throughput Proteomic Profiling of Transcription Factor in Human Atrial Tissue].

ABSTRACT

OBJECTIVE: To explore for the establishment of an experimental technique for profiling transcription factors, namely transcription factor response elements (TFRE), with high throughput and efficiency using human atrial tissue. METHODS: Postoperative right atrial tissues from 2 patients, one with preoperative atrial fibrillation and the one with no preoperative atrial fibrillation, were included in the study. The nucleus protein was extracted from the human atrial tissue, and the protein concentration was then measured. A solution with a complex formed through combining magnetic beads with concatenated tandem array of the consensus transcription factor response element DNA sequence (beads-catTFRE) was prepared, and the beads-catTFREs were then used to enrich transcription factors in the nucleoprotein extraction. SDS-PAGE electrophoresis was performed after dissociating beads-catTFRE from nucleoprotein with high temperature and high salt. The gel was then cut and faded before enzymolysis by trypsin in the gels was performed. Acetonitrile was used to extract the peptides from the gels, and the peptide solution was then dried. After that, we dissolved the peptides and performed mass spectrum tests, and the data were analyzed and processed with Firmiana one-stop proteomic analysis platform. RESULTS: In this study, 220 and 181 transcription factors were identified in the normal right atrial tissue and the right atrial tissue with atrial fibrillation, respectively. A total of 241 transcription factors were identified in the two groups. Among the 241 transcription factors, 12 were in the top 10% of those transcription factors that were above the median expression level of the normal right atrial tissue, and 12 transcription factors were in the top 10% of those above the median expression level of the right atrial tissue with atrial fibrillation. CONCLUSION: The high-throughput profiling method established in this study has high coverage, and the data collected can be used to support further validation studies.