Xist Repeat A coordinates an assembly of SR proteins to recruit SPEN and induce gene silencing

Trotman, J. B.; Porrello, A.; Schactler, S. A.; DeLeon, L. E.; Eberhard, Q. E.; Boyson, S. P.; Zhang, Z.; Lee, D. M.; Kirik, S. E.; Nguyen, S. N.; Beltejar, M.-C. G.; Shinn, M. K.; Ong, S.-E.; Gonzalez-Perez, M. P.; Shaffer, S. A.; Dominguez, D.; Shechner, D. M.; Calabrese, M.

doi:10.1101/2025.05.21.655143

bioRxiv · 10.1101/2025.05.21.655143

Xist Repeat A coordinates an assembly of SR proteins to recruit SPEN and induce gene silencing

Trotman, J. B.·Porrello, A.·Schactler, S. A.·DeLeon, L. E.·Eberhard, Q. E.·Boyson, S. P.·Zhang, Z.·Lee, D. M.·Kirik, S. E.·Nguyen, S. N.·Beltejar, M.-C. G.·Shinn, M. K.·Ong, S.-E.·Gonzalez-Perez, M. P.·Shaffer, S. A.·Dominguez, D.·Shechner, D. M.·Calabrese, M.

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Abstract

The lncRNA Xist represents a paradigm to understand the mechanisms of RNA-mediated gene silencing in mammals, which remain largely unresolved. To induce silencing, Xist recruits the RNA-binding protein SPEN through its 5'-proximal Repeat A domain. Yet, how Repeat A recruits SPEN and how SPEN coordinates silencing remain unclear. We report that sequences in Repeat A critical for SPEN recruitment directly bind SR-rich splicing factors. SRSF1, one such factor, is required for optimal SPEN recruitment and its RS-domain recruits SPEN when tethered to Xist. SPEN and SR-protein-binding motifs promote Repeat As association with many proteins, including the m6A machinery and elongating RNA polymerase II. SPEN also represses autosomal genes where its recruitment coincides with SR-protein binding. Our results reveal an unexpectedly essential role for splicing factors in coordinating silencing by Xist and suggest that the sensing of SR-protein-rich assemblies is a general mechanism through which SPEN targets genes for repression.

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Trotman, J. B., Porrello, A., Schactler, S. A., DeLeon, L. E., Eberhard, Q. E., Boyson, S. P., Zhang, Z., Lee, D. M., Kirik, S. E., Nguyen, S. N., Beltejar, M.-C. G., Shinn, M. K., Ong, S.-E., Gonzalez-Perez, M. P., Shaffer, S. A., Dominguez, D., Shechner, D. M., Calabrese, M.. 2025-05-26. Xist Repeat A coordinates an assembly of SR proteins to recruit SPEN and induce gene silencing. https://doi.org/10.1101/2025.05.21.655143

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