| Salmonella are Gram-negative bacterial pathogens that a capable of infecting a wide range of animals including mice, young cows, chicken and pig, however they are also able to colonize poultry and adult cattle without symptoms. In humans, infection with Salmonella results in a self-limiting gastroenteritis involving fever, diarrhea, and abdominal pain. The virulence of Salmonella is determined by flagella, fimbriae, and Salmonella pathogenicity islands (SPI) genes. There have been five SPIs identified in S. enterica serovar Typhimurium LT2, and invasion is mediated by a type III secretion system encoded on SPI-1. The SPI-1type III secretion system forms a needle-like complex that is responsible for the injection of bacterial effector proteins into the host cell cytosol. SPI-1effector proteins elicit several physiological changes in the host cell, including actin rearrangement leading to engulfment of the bacterium. After invasion of intestinal epithelial cells, Salmonella are able to disseminate to any tissue in the body, where the bacteria propagate inside macrophages, a process that requires a second type III secretion system encoded on SPI-2. The effectors facilitate Salmonella survival and replication in host cells by altering a variety of cell properties including its cytoskeletal structure, signal transduction pathways, and vesicular trafficking.The nucleoid-associated proteins Fis is a global regulator in bacteria, it could involve in maintaining genomic structure and gene regulation. Fis is a homodimer protein, and the intracellular level of Fis protein is growth-dependent and changes from less than100copies in stationary phase to more than50,000copies per cell in log phase. Fis represses transcription initiation either by imposing a blockade at the target promoter and excluding RNA polymerase or by a more subtle mechanism, in which it modulates the RNA polymerase-mediated isomerization of the closed transcription complex to an open complex. Fis can also be a conventional transcription activator, making physical contact with RNA polymerase. In addition, another mechanism involves the displacement of DNA twisting from an upstream site to the target promoter by Fis-mediated DNA bending.In the previously report, by using microarray technic, Fis was detected to regulate hundreds of genes transcription in S. enterica Typhimurium SL1344, including SPI genes. In order to obtain the regulation of global gene transcription by Fis and the regulatory mechanism, especially the regulatory mechanism on SPI genes, we used the high-throughput sequencing technic to perform ChIP-seq and RNA-seq in this study. We clarified the genome-wide distribution of binding regions and global regulation pattern of Fis in S. enterica Typhimurium LT2.In mid-exponential phase, Fis could up-regulate989genes and down-regulated657genes in LT2. And in the same time, Fis has895binding sites on genome-wide, spreading on943genes. Combined the two datasets, we find Fis regulates and binds on317genes, including207Fis up-regulated genes, and110Fis down-regulated genes. It shows that317Fis-regulated and Fis-binding genes are probably regulated by Fis directly, and1329Fis-regulated genes without Fis-binding are probably regulated by Fis indirectly. Since Fis regulates the transcriptions of several transcriptional factors genes, by using gene knockout and RT-PCR, we proved Fis controls the transcription of many genes by regulating corresponding transcriptional factors.We detect that several SPI genes are Fis-regulated genes and several SPI-genes are also Fis-binding genes. In this study, we firstly conclude the existence of several modes of Fis regulatory mechanism on SPI genes, and provide a regulatory network of Fis on SPI genes, including:1) Fis binds and regulates SPI genes directly;2) Fis binds to and activates the SPI positive regulator genes;3) Fis binds to the ORF region of SPI genes to mediate transcriptional activation of these genes and those downstream genes;4) Fis positively influences the SPI regulators by binding to and activating ompR.We also performed RNA-seq in early-exponential phase, and found that Fis could up-regulate509genes and down-regulated989genes in LT2. Fis is a repressor in early-exponential phase which is opposite with as an activator in mid-exponential phase. Compared the Fis-regulated genes in both phases, it shows that278genes have same Fis-regulatory tendency, whereas398genes have opposite Fis-regulatory tendency.We forecasted50sRNA from RNA-seq, and many of which are Fis-regulated. |
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