Chromatin-associated α-satellite RNA maintains chromosome stability by reestablishing SAF-A in the mitotic cell cycle

α-Satellite is the largest class of tandem repeats and is located on all human chromosome centromeres. Non-coding α-satellite RNAs have been observed in various cell types and are known to play crucial roles in maintaining genome stability. In this study, we demonstrated that α-satellite RNAs are dynamically expressed, heterogeneous transcripts that are regulated by Aurora kinases and closely associated with centromere chromatin throughout the mitotic cell cycle. We identified scaffold attachment factor A (SAF-A) as a previously uncharacterized α-satellite RNA binding protein. Depletion of either α-satellite RNA or SAF-A resulted in chromosome missegregation, revealing that their concerted action is essential for preserving genome integrity during the mitotic cell cycle. Our result demonstrated that SAF-A is excluded from the chromatin genome-wide during Mitosis, and α-satellite RNAs are required for the recruitment of SAF-A upon mitotic exit. Both α-satellite RNAs and SAF-A are essential in safeguarding the human genome against chromosomal instability during Mitosis. Moreover, α-satellite RNAs and SAF-A aid in the reassembly of the nuclear lamina. Our results provide novel insights into the features, regulations, and functional roles of α-satellite RNAs and propose a model for the dismantling and reformation of the SAF-A nuclear scaffold during Mitosis.