Cancer Epigenetics Program, Fox Chase Cancer Center, Temple University Health System, Philadelphia, PA 19111, USA.
Department of Cell and Developmental Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA 19104, USA
J Cell Sci. 2020 May 27;133(10):jcs240416. doi: 10.1242/jcs.240416.
A large fraction of epigenetically silent heterochromatin is anchored to the nuclear periphery via 'tethering proteins' that function to bridge heterochromatin and the nuclear membrane or nuclear lamina. We previously identified a human tethering protein, PRR14, that binds heterochromatin through an N-terminal domain, but the mechanism and regulation of nuclear lamina association remained to be investigated. Here we identify an evolutionarily conserved PRR14 nuclear lamina binding domain (LBD) that is both necessary and sufficient for positioning of PRR14 at the nuclear lamina. We show that PRR14 associates dynamically with the nuclear lamina, and provide evidence that such dynamics are regulated through phosphorylation and dephosphorylation of the LBD. Furthermore, we identify a PP2A phosphatase recognition motif within the evolutionarily conserved C-terminal Tantalus domain of PRR14. Disruption of this motif affects PRR14 localization to the nuclear lamina. The overall findings demonstrate a heterochromatin anchoring mechanism whereby the PRR14 tether simultaneously binds heterochromatin and the nuclear lamina through two separable modular domains. Our findings also describe an optimal PRR14 LBD fragment that could be used for efficient targeting of fusion proteins to the nuclear lamina.
大量的表观遗传沉默异染色质通过“连接蛋白”锚定在核周，这些连接蛋白的功能是连接异染色质和核膜或核纤层。我们之前鉴定了一种人类连接蛋白 PRR14，它通过一个 N 端结构域结合异染色质，但核纤层结合的机制和调控仍有待研究。在这里，我们确定了一个进化上保守的 PRR14 核纤层结合域（LBD），它对于 PRR14 在核纤层上的定位既是必需的，也是充分的。我们表明 PRR14 与核纤层动态相关，并提供证据表明这种动力学通过 LBD 的磷酸化和去磷酸化来调节。此外，我们在 PRR14 的进化上保守的 C 端 Tantalus 结构域内鉴定了一个 PP2A 磷酸酶识别基序。破坏该基序会影响 PRR14 向核纤层的定位。总的来说，这些发现展示了一种异染色质锚定机制，其中 PRR14 连接蛋白通过两个可分离的模块化结构域同时结合异染色质和核纤层。我们的研究结果还描述了一个优化的 PRR14 LBD 片段，可用于将融合蛋白有效地靶向核纤层。
