Department of Plant Biology, University of Illinois at Urbana-Champaign, Urbana, Illinois, 61801, USA.
College of Life Sciences, Nanjing Normal University, Nanjing, 210023, China.
J Integr Plant Biol. 2021 Dec;63(12):2075-2092. doi: 10.1111/jipb.13169. Epub 2021 Nov 2.
Translational control of gene expression, including recruitment of ribosomes to messenger RNA (mRNA), is particularly important during the response to stress. Purification of ribosome-associated mRNAs using translating ribosome affinity purification (TRAP) followed by RNA-sequencing facilitates the study of mRNAs undergoing active transcription and better proxies the translatome, or protein response, to stimuli. To identify plant responses to Magnesium (Mg) deficiency at the translational level, we combined transcriptome and translatome analyses. Excitingly, we found 26 previously unreported Mg-responsive genes that were only regulated at the translational level and not the transcriptional level, during the early response to Mg deficiency. In addition, mutants of the transcription factor ELONGATED HYPOCOTYL 5 (HY5), the H /CATION EXCHANGER 1 and 3 (CAX1 and CAX3), and UBIQUITIN 11 (UBQ11) exhibited early chlorosis phenotype under Mg deficiency, supporting their functional involvement in ion homeostasis. Overall, our study strongly supports that TRAP-seq combined with RNA-seq followed by phenotype screening could facilitate the identification of novel players during stress responses.
基因表达的翻译调控，包括核糖体向信使 RNA（mRNA）的募集，在应激反应中尤为重要。使用翻译核糖体亲和纯化（TRAP）对核糖体相关的 mRNA 进行纯化，然后进行 RNA 测序，这有助于研究正在进行活跃转录的 mRNA，并更好地代表对刺激的核糖体组或蛋白质反应。为了在翻译水平上鉴定植物对镁（Mg）缺乏的反应，我们结合了转录组和翻译组分析。令人兴奋的是，我们发现了 26 个以前未报道的 Mg 响应基因，它们在 Mg 缺乏的早期反应中仅在翻译水平而不是转录水平上受到调节。此外，转录因子 ELONGATED HYPOCOTYL 5（HY5）、H / CATION EXCHANGER 1 和 3（CAX1 和 CAX3）以及泛素 11（UBQ11）的突变体在 Mg 缺乏下表现出早期黄化表型，这支持它们在离子稳态中的功能参与。总的来说，我们的研究强烈支持 TRAP-seq 与 RNA-seq 相结合，然后进行表型筛选，可促进在应激反应中鉴定新的参与者。
