Structural insights into the assembly and polyA signal recognition mechanism of the human CPSF complex.
Structural insights into the assembly and polyA signal recognition mechanism of the human CPSF complex.
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DOI：
10.7554/elife.33111
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发表时间：
2017-12-23
期刊：
影响因子：
7.7
通讯作者：
Jinek M
中科院分区：
文献类型：
--
作者：
Clerici M;Faini M;Aebersold R;Jinek M
3 '聚腺苷酸化是真核生物mRNA生物发生的关键步骤。在哺乳动物细胞中，这一过程依赖于切割和聚腺苷化特异性因子（CPSF）复合物对mrna前聚腺苷化信号中六核苷酸AAUAAA基序的识别。由CPSF160、WDR33、CPSF30和Fip1组成的核心CPSF复合物足以识别AAUAAA基序，但其组装的分子相互作用和PAS识别机制尚不清楚。基于交联耦合质谱，CPSF160-WDR33亚复合物的晶体结构和生化分析，我们确定了核心人类CPSF复合物的分子结构，确定了参与亚基间相互作用的特定结构域。除了CPSF30中的锌指结构域外，我们通过定量rna结合分析发现WDR33中n端赖氨酸/精氨酸丰富的基序是特异性AAUAAA基序识别的关键决定因素。总之，这些结果揭示了CPSF在介导pas依赖的RNA切割和聚腺苷化中的功能。
3’ polyadenylation is a key step in eukaryotic mRNA biogenesis. In mammalian cells, this process is dependent on the recognition of the hexanucleotide AAUAAA motif in the pre-mRNA polyadenylation signal by the cleavage and polyadenylation specificity factor (CPSF) complex. A core CPSF complex comprising CPSF160, WDR33, CPSF30 and Fip1 is sufficient for AAUAAA motif recognition, yet the molecular interactions underpinning its assembly and mechanism of PAS recognition are not understood. Based on cross-linking-coupled mass spectrometry, crystal structure of the CPSF160-WDR33 subcomplex and biochemical assays, we define the molecular architecture of the core human CPSF complex, identifying specific domains involved in inter-subunit interactions. In addition to zinc finger domains in CPSF30, we identify using quantitative RNA-binding assays an N-terminal lysine/arginine-rich motif in WDR33 as a critical determinant of specific AAUAAA motif recognition. Together, these results shed light on the function of CPSF in mediating PAS-dependent RNA cleavage and polyadenylation.