Yeast poly(A)-binding protein (Pab1) controls translation initiation in vivo primarily by blocking mRNA decapping and decay

Poly(A)-binding protein (Pab1 in yeast) is involved in mRNA decay and translation initiation, but its molecular functions are incompletely understood. In this study, we employed a comprehensive multiomics strategy encompassing Ribosome profiling, spike-in normalized RNA-seq, SMPAT-seq, and CAGE-seq to gain mechanistic insight into Pab1's role in Saccharomyces cerevisiae. We found that auxin-induced degradation of Pab1 reduced bulk mRNA and polysome abundance in a manner suppressed by deleting the catalytic subunit of decapping enzyme (dcp2Δ), demonstrating that enhanced decapping/degradation is the major driver of reduced mRNA abundance and protein synthesis at limiting Pab1 levels. An increased median poly(A) tail length conferred by Pab1 depletion was also nullified by dcp2Δ, suggesting that mRNA isoforms with shorter tails are preferentially decapped/degraded at limiting Pab1. In contrast to findings on mammalian cells, the translational efficiencies (TEs) of many mRNAs were altered by Pab1 depletion; however, these changes were broadly diminished by dcp2∆, suggesting that reduced mRNA abundance is a major driver of translational reprogramming at limiting Pab1. Thus, assembly of the closed-loop mRNP via PABP-eIF4G interaction appears to be dispensable for normal translation of most yeast mRNAs in vivo at normal mRNA levels. Interestingly, histone mRNAs and proteins are preferentially diminished on Pab1 depletion dependent on Dcp2, accompanied by activation of internal cryptic promoters in the manner expected for reduced nucleosome occupancies, revealing a new layer of post-transcriptional control of histone gene expression.聚(A)结合蛋白（Poly(A)-binding protein，酵母中同源蛋白为Pab1）参与mRNA降解与翻译起始过程，但其分子功能尚未被完全阐明。本研究采用涵盖核糖体谱（Ribosome profiling）、spike-in归一化RNA测序（spike-in normalized RNA-seq）、SMPAT-seq以及Cap分析基因表达测序（CAGE-seq）的综合性多组学策略，以解析酿酒酵母（Saccharomyces cerevisiae）中Pab1的作用机制。研究发现，生长素诱导的Pab1降解会降低整体mRNA与多聚核糖体丰度，这一效应可通过敲除脱帽酶（decapping enzyme）的催化亚基dcp2Δ得以抑制，表明在Pab1水平受限的条件下，增强的脱帽/降解过程是mRNA丰度与蛋白质合成降低的主要驱动因素。由Pab1缺失所导致的poly(A)尾长度中位数升高，同样可被dcp2Δ抵消，这提示在Pab1水平受限的情况下，带有较短poly(A)尾的mRNA亚型会被优先脱帽/降解。与哺乳动物细胞的已有研究结果不同，许多mRNA的翻译效率（translational efficiencies, TEs）会因Pab1缺失而发生改变；但这类变化可被dcp2Δ广泛削弱，表明在Pab1水平受限的情况下，mRNA丰度降低是翻译重编程的主要驱动因素。由此可见，通过PABP-eIF4G相互作用形成的闭环信使核糖核蛋白复合物（messenger ribonucleoprotein complex, mRNP）组装，对于正常mRNA水平下酿酒酵母多数mRNA的体内正常翻译并非必需。值得注意的是，组蛋白mRNA与蛋白质会因Pab1缺失而优先减少，且该过程依赖于Dcp2；同时伴随内部隐蔽启动子（internal cryptic promoters）的激活，这与核小体占据率降低所预期的效应一致，该发现揭示了组蛋白基因表达转录后调控的新层面。