Instituto de Bioquímica Vegetal y Fotosíntesis, Consejo Superior de Investigaciones Científicas and Universidad de Sevilla, Sevilla, Spain.
Genetics and Experimental Bioinformatics, Faculty of Biology, University of Freiburg, Freiburg, Germany.
FEBS J. 2021 Mar;288(5):1614-1629. doi: 10.1111/febs.15516. Epub 2020 Sep 2.
NsiR3 (nitrogen stress-inducible RNA 3) is a small noncoding RNA strongly conserved in heterocyst-forming cyanobacteria. In Nostoc sp. PCC 7120, transcription of NsiR3 is induced by nitrogen starvation and depends on the global nitrogen regulator NtcA. A conserved NtcA-binding site is centered around position -42.5 with respect to the transcription start site of NsiR3 homologs, and NtcA binds in vitro to a DNA fragment containing this sequence. In the absence of combined nitrogen, NsiR3 expression is induced in all cells along the Nostoc filament but much more strongly in heterocysts, differentiated cells devoted to nitrogen fixation. Co-expression analysis of transcriptomic data obtained from microarrays hybridized with RNA obtained from Nostoc wild-type or mutant strains grown in the presence of ammonium or in the absence of combined nitrogen revealed that the expression profile of gene putA (proline oxidase) correlates negatively with that of NsiR3. Using a heterologous system in Escherichia coli, we show that NsiR3 binds to the 5'-UTR of putA mRNA, resulting in reduced expression of a reporter gene. Overexpression of NsiR3 in Nostoc resulted in strong reduction of putA mRNA accumulation, further supporting the negative regulation of putA by NsiR3. The higher expression of NsiR3 in heterocysts versus vegetative cells of the N -fixing filament could contribute to the previously described absence of putA mRNA and of the catabolic pathway to produce glutamate from arginine via proline specifically in heterocysts. Post-transcriptional regulation by NsiR3 represents an indirect NtcA-operated regulatory mechanism of putA expression. DATABASE: Microarray data are available in GEO database under accession numbers GSE120377 and GSE150191.
NsiR3（氮胁迫诱导 RNA3）是一种在异形胞形成蓝藻中高度保守的小非编码 RNA。在 Nostoc sp. PCC 7120 中，NsiR3 的转录受氮饥饿诱导，并依赖于全局氮调节因子 NtcA。一个保守的 NtcA 结合位点位于转录起始位点的-42.5 左右，NsiR3 同源物的位置，并且 NtcA 在体外与包含该序列的 DNA 片段结合。在没有结合氮的情况下，NsiR3 在 Nostoc 丝状体的所有细胞中都被诱导表达，但在异形胞中表达更强，异形胞是专门用于固氮的分化细胞。用微阵列获得的转录组数据的共表达分析与在铵存在或没有结合氮的情况下生长的 Nostoc 野生型或突变株获得的 RNA 杂交的微阵列，表明基因 putA（脯氨酸氧化酶）的表达谱与 NsiR3 呈负相关。使用大肠杆菌中的异源系统，我们表明 NsiR3 与 putA mRNA 的 5'-UTR 结合，导致报告基因的表达减少。NsiR3 在 Nostoc 中的过表达导致 putA mRNA 积累的强烈减少，进一步支持 NsiR3 对 putA 的负调控。与固氮丝状体的营养细胞相比，异形胞中 NsiR3 的高表达可能导致先前描述的 putA mRNA 缺失和从精氨酸通过脯氨酸产生谷氨酸的分解代谢途径缺失，特别是在异形胞中。NsiR3 的转录后调节代表了 putA 表达的一种间接的 NtcA 操作调节机制。数据库：微阵列数据可在 GEO 数据库中获得，注册号为 GSE120377 和 GSE150191。
