Brenes-Álvarez Manuel, Ropp Halie R, Papagiannidis Dimitrios, Potel Clement M, Stein Frank, Scholz Ingeborg, Steglich Claudia, Savitski Mikhail M, Vioque Agustín, Muro-Pastor Alicia M, Hess Wolfgang R
Genetics and Experimental Bioinformatics, Faculty of Biology, University of Freiburg, Schänzlestr. 1, 79104 Freiburg, Germany.
European Molecular Biology Laboratory (EMBL), Meyerhofstr. 1, 69117 Heidelberg, Germany.
Nucleic Acids Res. 2025 Jan 24;53(3). doi: 10.1093/nar/gkae1247.
RNA-binding proteins (RBPs) are central components of gene regulatory networks. The differentiation of heterocysts in filamentous cyanobacteria is an example of cell differentiation in prokaryotes. Although multiple non-coding transcripts are involved in this process, no RBPs have been implicated thus far. Here we used quantitative mass spectrometry to analyze the differential fractionation of RNA-protein complexes after RNase treatment in density gradients yielding 333 RNA-associated proteins, while a bioinformatic prediction yielded 311 RBP candidates in Nostoc sp. PCC 7120. We validated in vivo the RNA-binding capacity of six RBP candidates. Some participate in essential physiological aspects, such as photosynthesis (Alr2890), thylakoid biogenesis (Vipp1) or heterocyst differentiation (PrpA, PatU3), but their association with RNA was unknown. Validated RBPs Asl3888 and Alr1700 were not previously characterized. Alr1700 is an RBP with two oligonucleotide/oligosaccharide-binding (OB)-fold-like domains that is differentially expressed in heterocysts and interacts with non-coding regulatory RNAs. Deletion of alr1700 led to complete deregulation of the cell differentiation process, a striking increase in the number of heterocyst-like cells, and was ultimately lethal in the absence of combined nitrogen. These observations characterize this RBP as a master regulator of the heterocyst patterning and differentiation process, leading us to rename Alr1700 to PatR.
RNA结合蛋白（RBPs）是基因调控网络的核心组成部分。丝状蓝细菌中异形胞的分化是原核生物细胞分化的一个例子。尽管多个非编码转录本参与了这一过程，但迄今为止尚未发现有RBPs与之相关。在这里，我们使用定量质谱分析法分析了核糖核酸酶处理后RNA-蛋白质复合物在密度梯度中的差异分级分离，共得到333种与RNA相关的蛋白质，同时通过生物信息学预测在集胞藻属PCC 7120中得到311个RBP候选蛋白。我们在体内验证了6个RBP候选蛋白的RNA结合能力。其中一些参与重要的生理过程，如光合作用（Alr2890）、类囊体生物发生（Vipp1）或异形胞分化（PrpA、PatU3），但它们与RNA的关联此前并不清楚。经验证的RBPs Asl3888和Alr1700此前未被鉴定。Alr1700是一种具有两个寡核苷酸/寡糖结合（OB）折叠样结构域的RBP，在异形胞中差异表达，并与非编码调控RNA相互作用。删除alr1700导致细胞分化过程完全失控，异形胞样细胞数量显著增加，最终在缺乏化合态氮的情况下致死。这些观察结果表明该RBP是异形胞模式和分化过程的主要调节因子，因此我们将Alr1700重新命名为PatR。
