Bouthier de la Tour Claire, Blanchard Laurence, Dulermo Rémi, Ludanyi Monika, Devigne Alice, Armengaud Jean, Sommer Suzanne, de Groot Arjan
1Institute for Integrative Biology of the Cell (I2BC), CEA, CNRS, Université Paris Sud, Bâtiment 409, F-91405 Orsay, France.
2CEA, DSV, IBEB, Lab Bioenerget Cellulaire, F-13108 Saint-Paul-lez-Durance, France 3CNRS, UMR 7265 Biol Veget & Microbiol Environ, F-13108 Saint-Paul-lez-Durance, France 4Aix-Marseille Université, F-13108 Saint-Paul-lez-Durance, France.
Microbiology (Reading). 2015 Dec;161(12):2410-22. doi: 10.1099/mic.0.000186. Epub 2015 Sep 18.
HU proteins have an important architectural role in nucleoid organization in bacteria. Compared with HU of many bacteria, HU proteins from Deinococcus species possess an N-terminal lysine-rich extension similar to the eukaryotic histone H1 C-terminal domain involved in DNA compaction. The single HU gene in Deinococcus radiodurans, encoding DrHU, is required for nucleoid compaction and cell viability. Deinococcus deserti contains three expressed HU genes, encoding DdHU1, DdHU2 and DdHU3. Here, we show that either DdHU1 or DdHU2 is essential in D. deserti. DdHU1 and DdHU2, but not DdHU3, can substitute for DrHU in D. radiodurans, indicating that DdHU3 may have a non-essential function different from DdHU1, DdHU2 and DrHU. Interestingly, the highly abundant DrHU and DdHU1 proteins, and also the less expressed DdHU2, are translated in Deinococcus from leaderless mRNAs, which lack a 5'-untranslated region and, hence, the Shine-Dalgarno sequence. Unexpectedly, cloning the DrHU or DdHU1 gene under control of a strong promoter in an expression plasmid, which results in leadered transcripts, strongly reduced the DrHU and DdHU1 protein level in D. radiodurans compared with that obtained from the natural leaderless gene. We also show that the start codon position for DrHU and DdHU1 should be reannotated, resulting in proteins that are 15 and 4 aa residues shorter than initially reported. The expression level and start codon correction were crucial for functional characterization of HU in Deinococcus.
HU蛋白在细菌类核组织中具有重要的结构作用。与许多细菌的HU相比，嗜热放线菌属物种的HU蛋白具有富含赖氨酸的N端延伸，类似于参与DNA压缩的真核组蛋白H1 C端结构域。嗜放射栖热菌中的单个HU基因编码DrHU，是类核压缩和细胞活力所必需的。嗜热栖热菌含有三个表达的HU基因，分别编码DdHU1、DdHU2和DdHU3。在此，我们表明DdHU1或DdHU2在嗜热栖热菌中是必不可少的。DdHU1和DdHU2，但不是DdHU3，可以替代嗜放射栖热菌中的DrHU，这表明DdHU3可能具有与DdHU1、DdHU2和DrHU不同的非必需功能。有趣的是，高度丰富的DrHU和DdHU1蛋白，以及表达较少的DdHU2，在嗜热放线菌中是从无领导mRNA翻译而来的，这些mRNA缺乏5'非翻译区，因此也缺乏Shine-Dalgarno序列。出乎意料的是，在表达质粒中强启动子的控制下克隆DrHU或DdHU1基因，这会产生有前导序列的转录本，与从天然无领导基因获得的相比，嗜放射栖热菌中DrHU和DdHU1蛋白水平大幅降低。我们还表明，DrHU和DdHU1的起始密码子位置应重新注释，导致产生的蛋白质比最初报道的短15和4个氨基酸残基。表达水平和起始密码子校正对于嗜热放线菌中HU的功能表征至关重要。
