Polyadenylation controls mRNA biogenesis, nucleo-cytoplasmic export, translation and decay. These processes are interdependent and coordinately regulated by poly(A)-binding proteins (PABPs), yet how PABPs are themselves regulated is not fully understood. Here, we report the discovery that human nuclear PABPN1 is phosphorylated by mitotic kinases at four specific sites during mitosis, a time when nucleoplasm and cytoplasm mix. To understand the functional consequences of phosphorylation, we generated a panel of stable cell lines inducibly over-expressing PABPN1 with point mutations at these sites. Phospho-inhibitory mutations decreased cell proliferation, highlighting the importance of PABPN1 phosphorylation in cycling cells. Dynamic regulation of poly(A) tail length and RNA stability have emerged as important modes of gene regulation. We therefore employed long-read sequencing to determine how PABPN1 phospho-site mutants affected poly(A) tails lengths and TimeLapse-seq to monitor mRNA synthesis and decay. Widespread poly(A) tail lengthening was observed for phospho-inhibitory PABPN1 mutants. In contrast, expression of phospho-mimetic PABPN1 resulted in shorter poly(A) tails with increased non-A nucleotides, in addition to increased transcription and reduced stability of a distinct cohort of mRNAs. Taken together, PABPN1 phosphorylation remodels poly(A) tails and increases mRNA turnover, supporting the model that enhanced transcriptome dynamics reset gene expression programs across the cell cycle.

中文翻译：

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有丝分裂期间核聚腺苷酸结合蛋白 (PABPN1) 的磷酸化可保护 mRNA 免于腺苷酸过度化并维持转录组动态


多聚腺苷酸化控制 mRNA 生物合成、核质输出、翻译和衰变。这些过程是相互依赖的，并由多聚腺苷酸结合蛋白（PABP）协调调节，但 PABP 本身如何调节尚不完全清楚。在此，我们报告了人核 PABPN1 在有丝分裂期间（核质和细胞质混合时）的四个特定位点被有丝分裂激酶磷酸化的发现。为了了解磷酸化的功能后果，我们生成了一组可诱导过度表达 PABPN1 的稳定细胞系，这些细胞系在这些位点具有点突变。磷酸化抑制突变降低了细胞增殖，凸显了 PABPN1 磷酸化在细胞周期中的重要性。 Poly(A) 尾长和 RNA 稳定性的动态调控已成为基因调控的重要模式。因此，我们采用长读长测序来确定 PABPN1 磷酸位点突变体如何影响 Poly(A) 尾长度，并采用 TimeLapse-seq 来监测 mRNA 合成和衰减。在磷酸抑制性 PABPN1 突变体中观察到广泛的聚 (A) 尾延长。相比之下，磷酸模拟PABPN1的表达除了导致不同mRNA群的转录增加和稳定性降低之外，还导致聚腺苷酸尾变短，非A核苷酸增加。总而言之，PABPN1 磷酸化重塑了 Poly(A) 尾并增加了 mRNA 更新，支持了增强转录组动态重置整个细胞周期基因表达程序的模型。