The lupus autoantigen La is an Xist-binding RNA chaperone involved in Xist folding and cloud formation

Using the programmable RNA-sequence binding domain of the Pumilio protein, we FLAG-tagged Xist (inactivated X chromosome specific transcript) in live cells. Affinity pulldown coupled to mass spectrometry was employed to identify a list of 138 candidate Xist-binding proteins, from which, the lupus autoantigen La (encoding gene Ssb) was validated as a protein functionally critical for X chromosome inactivation (XCI). Extensive XCI defects were detected in Ssb knockdown cells, including chromatin compaction, death of female ES cells during in vitro differentiation and chromosome-wide monoallelic gene expression pattern. Live-cell imaging of Xist RNA reveals the defining XCI defect: Xist cloud formation. La is a ubiquitous and versatile RNA-binding protein with RNA chaperone activity. Functional dissection of La shows that the RNA chaperon domain plays critical roles in XCI. In mutant cells, Xist transcripts are misfolded and unstable. These results show that La is involved in XCI as an RNA chaperone for Xist.