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author:("Niu, guolin")
1.  GouR, a TetR Family Transcriptional Regulator, Coordinates the Biosynthesis and Export of Gougerotin in Streptomyces graminearus 
Gougerotin is a peptidyl nucleoside antibiotic. It functions as a specific inhibitor of protein synthesis by binding ribosomal peptidyl transferase and exhibits a broad spectrum of biological activities. gouR, situated in the gougerotin biosynthetic gene cluster, encodes a TetR family transcriptional regulatory protein. Gene disruption and genetic complementation revealed that gouR plays an important role in the biosynthesis of gougerotin. Transcriptional analysis suggested that GouR represses the transcription of the gouL-to-gouB operon consisting of 11 structural genes and activates the transcription of the major facilitator superfamily (MFS) transporter gene (gouM). Electrophoresis mobility shift assays (EMSAs) and DNase I footprinting experiments showed that GouR has specific DNA-binding activity for the promoter regions of gouL, gouM, and gouR. Our data suggested that GouR modulates gougerotin production by coordinating its biosynthesis and export in Streptomyces graminearus.
doi:10.1128/AEM.03003-13
PMCID: PMC3911114  PMID: 24242236
2.  Novel nikkomycin analogues generated by mutasynthesis in Streptomyces ansochromogenes 
Background
Nikkomycins are competitive inhibitors of chitin synthase and inhibit the growth of filamentous fungi, insects, acarids and yeasts. The gene cluster responsible for biosynthesis of nikkomycins has been cloned and the biosynthetic pathway was elucidated at the genetic, enzymatic and regulatory levels.
Results
Streptomyces ansochromogenes ΔsanL was constructed by homologous recombination and the mutant strain was fed with benzoic acid, 4-hydroxybenzoic acid, nicotinic acid and isonicotinic acid. Two novel nikkomycin analogues were produced when cultures were supplemented with nicotinic acid. These two compounds were identified as nikkomycin Px and Pz by electrospray ionization mass spectrometry (ESI-MS) and nuclear magnetic resonance (NMR). Bioassays against Candida albicans and Alternaria longipes showed that nikkomycin Px and Pz exhibited comparatively strong inhibitory activity as nikkomycin X and Z produced by Streptomyces ansochromogenes 7100 (wild-type strain). Moreover, nikkomycin Px and Pz were found to be more stable than nikkomycin X and Z at different pH and temperature conditions.
Conclusions
Two novel nikkomycin analogues (nikkomycin Px and Pz) were generated by mutasynthesis with the sanL inactivated mutant of Streptomyces ansochromogenes 7100. Although antifungal activities of these two compounds are similar to those of nikkomycin X and Z, their stabilities are much better than nikkomycin X and Z under different pHs and temperatures.
doi:10.1186/1475-2859-13-59
PMCID: PMC4021061  PMID: 24751325
Novel analogues; Mutasynthesis; sanL mutant; Streptomyces ansochromogenes
3.  Molecular Regulation of Antibiotic Biosynthesis in Streptomyces 
SUMMARY
Streptomycetes are the most abundant source of antibiotics. Typically, each species produces several antibiotics, with the profile being species specific. Streptomyces coelicolor, the model species, produces at least five different antibiotics. We review the regulation of antibiotic biosynthesis in S. coelicolor and other, nonmodel streptomycetes in the light of recent studies. The biosynthesis of each antibiotic is specified by a large gene cluster, usually including regulatory genes (cluster-situated regulators [CSRs]). These are the main point of connection with a plethora of generally conserved regulatory systems that monitor the organism's physiology, developmental state, population density, and environment to determine the onset and level of production of each antibiotic. Some CSRs may also be sensitive to the levels of different kinds of ligands, including products of the pathway itself, products of other antibiotic pathways in the same organism, and specialized regulatory small molecules such as gamma-butyrolactones. These interactions can result in self-reinforcing feed-forward circuitry and complex cross talk between pathways. The physiological signals and regulatory mechanisms may be of practical importance for the activation of the many cryptic secondary metabolic gene cluster pathways revealed by recent sequencing of numerous Streptomyces genomes.
doi:10.1128/MMBR.00054-12
PMCID: PMC3591988  PMID: 23471619
4.  Identification of a Novel Bacteriocin Regulatory System in Streptococcus mutans 
Molecular microbiology  2010;78(6):1431-1447.
Summary
Recently, we described the function of an uncharacterized two-gene regulatory system consisting of a LytTR family transcription regulator and a putative membrane protein, which we referred to as the hdrRM operon. In this study, we determined that the HdrRM system controls the expression of an analogous uncharacterized regulatory system annotated as SMU.2080 and SMU.2081. Like hdrRM, the SMU.2080–2081 operon encodes a LytTR family transcription regulator and putative membrane protein, which we now refer to as BrsR and BrsM, respectively. Examination of the regulatory mechanism of the BrsRM system suggests that BrsM serves to antagonize the function of the transcription regulator BrsR. Further analyses of the regulatory role of BrsR determined that it functions as a transcription activator for a variety of bacteriocins and bacteriocin-related genes. In vitro electromobility shift assays confirmed that BrsR binds to the promoter regions of several bacteriocin genes and requires the presence of a LytTR family consensus direct repeat in order to stably bind DNA. In addition, we identified a novel regulatory scheme in which both the HdrRM and BrsRM systems coregulate each other and ultimately determine whether bacteriocin production will inhibit competitor organisms or result in lethality to the producer.
doi:10.1111/j.1365-2958.2010.07417.x
PMCID: PMC3059113  PMID: 21143316
bacteriocin; LytTR; oral bacteria; Streptococcus; cell death
5.  The cia Operon of Streptococcus mutans Encodes a Unique Component Required for Calcium-Mediated Autoregulation 
Molecular microbiology  2008;70(1):112-126.
Streptococcus mutans is a primary pathogen for dental caries in humans. CiaR and CiaH of S. mutans comprise a two-component signal transduction system (TCS) involved in regulating various virulent factors. However, the signal that triggers the CiaRH response remains unknown. In this study, we show that calcium is a signal for regulation of the ciaRH operon, and that a double-glycine-containing small peptide encoded within the ciaRH operon (renamed ciaX) mediates this regulation. CiaX contains a serine-aspartate (SD) domain that is shared by calcium-binding proteins. A markerless in-frame deletion of ciaX reduced ciaRH operon expression and diminished the calcium repression of operon transcription. Point mutations of the SD-domain resulted in the same phenotype as the in-frame deletion, indicating that the SD-domain is required for CiaX function. Further characterization of ciaX demonstrated that it is involved in calcium mediated biofilm formation. Furthermore, inactivation of ciaR or ciaH led to the same phenotype as the in-frame deletion of ciaX, suggesting that all three genes are involved in the same regulatory pathway. Sequence analysis and real-time RT-PCR identified a putative CiaR binding site upstream of ciaX. We conclude that the ciaXRH operon is a three-component, self-regulatory system modulating cellular functions in response to calcium.
doi:10.1111/j.1365-2958.2008.06390.x
PMCID: PMC2955730  PMID: 18681938
6.  The hdrRM Operon of Streptococcus mutans Encodes a Novel Regulatory System for Coordinated Competence Development and Bacteriocin Production▿  
Journal of Bacteriology  2010;192(7):1844-1852.
The Streptococcus mutans hdrRM operon encodes a novel two-gene regulatory system induced by high cell density. Previous studies identified hdrM as the only known negative regulator of competence development in S. mutans. In the present study, we demonstrated that the HdrRM system bypasses the prototypical competence gene regulators ComC and ComDE in the transcriptional regulation of the competence-specific sigma factor comX and the late competence genes. Similarly, the HdrRM system can abrogate the requirement for ComE to produce the bacteriocin mutacin IV. To further probe the regulatory mechanism of hdrRM, we created an hdrR overexpression strain and showed that it could reproduce each of the hdrM competence and mutacin phenotypes, indicating that HdrM acts as a negative regulator of HdrR activity. Using a mutacin IV-luciferase reporter, we also demonstrated that the hdrRM system utilizes the same promoter elements recognized by ComE and thus appears to comprise a novel regulatory pathway parallel to ComCDE.
doi:10.1128/JB.01667-09
PMCID: PMC2838059  PMID: 20118256
7.  The Streptococcus mutans IrvR Repressor Is a CI-Like Regulator That Functions through Autocleavage and Clp-Dependent Proteolysis▿  
Journal of Bacteriology  2009;192(6):1586-1595.
Previous work has shown that irvR is required for the proper regulation of genetic competence and dextran-dependent aggregation due to its ability to repress the transcription regulator irvA. In this study, we determined the mechanism used to relieve the repression of irvA. We demonstrate that IrvR is a “LexA-like” protein with four conserved amino acid residues likely required for IrvR autocleavage activity. Furthermore, recombinant IrvR protein purified from Escherichia coli was competent to undergo autocleavage in vitro. Using several truncated IrvR constructs, we show that the amino acids adjacent to the autocleavage site are essential for relieving irvA repression and engaging the irvA-dependent regulatory pathway primarily through the ClpXP and ClpCP proteases. By extending the IrvR C terminus with an epitope derived from the autocleavage site, we were also able to create a constitutive Clp-dependent degradation of the full-length IrvR protein. This suggests that the derepression of irvA occurs through a two-step mechanism involving the initial autocleavage of IrvR and exposure of a proteolytic degradation sequence followed by Clp-dependent degradation of the IrvR DNA binding domain. Thus, irvA derepression is highly analogous to the genetic switch mechanism used to regulate lysogeny in bacteriophages.
doi:10.1128/JB.01261-09
PMCID: PMC2832535  PMID: 20038591
8.  Autoaggregation Response of Fusobacterium nucleatum▿ †  
Applied and Environmental Microbiology  2009;75(24):7725-7733.
Fusobacterium nucleatum is a gram-negative oral bacterial species associated with periodontal disease progression. This species is perhaps best known for its ability to adhere to a vast array of other bacteria and eukaryotic cells. Numerous studies of F. nucleatum have examined various coaggregation partners and inhibitors, but it is largely unknown whether these interactions induce a particular genetic response. We tested coaggregation between F. nucleatum ATCC strain 25586 and various species of Streptococcus in the presence of a semidefined growth medium containing saliva. We found that this condition could support efficient coaggregation but, surprisingly, also stimulated a similar degree of autoaggregation. We further characterized the autoaggregation response, since few reports have examined this in F. nucleatum. After screening several common coaggregation inhibitors, we identified l-lysine as a competitive inhibitor of autoaggregation. We performed a microarray analysis of the planktonic versus autoaggregated cells and found nearly 100 genes that were affected after only about 60 min of aggregation. We tested a subset of these genes via real-time reverse transcription-PCR and confirmed the validity of the microarray results. Some of these genes were also found to be inducible in cell pellets created by centrifugation. Based upon these data, it appears that autoaggregation activates a genetic program that may be utilized for growth in a high cell density environment, such as the oral biofilm.
doi:10.1128/AEM.00916-09
PMCID: PMC2794088  PMID: 19837836
9.  Characterization of irvR, a Novel Regulator of the irvA-Dependent Pathway Required for Genetic Competence and Dextran-Dependent Aggregation in Streptococcus mutans▿  
Journal of Bacteriology  2008;190(21):7268-7274.
Previous studies identified irvA as a normally repressed but highly inducible transcription regulator capable of repressing mutacin I gene expression in Streptococcus mutans. In this study, we aimed to identify and characterize the regulator(s) responsible for repressing the expression of irvA. An uncharacterized open reading frame (SMU.1398) located immediately adjacent to irvA and annotated as a putative transcription repressor was identified as a likely candidate. The results of mutation studies confirmed that the expression of irvA was greatly increased in the SMU.1398 background. Mutation of SMU.1398 (“irvR”) abolished genetic competence and reduced the expression of the late competence genes/operons comEA, comY, and dprA without affecting the expression of the known competence regulators comC, comED, or comX. In addition, irvR was found to be a potent negative regulator of dextran-dependent aggregation (DDAG) and gbpC expression. Each of these irvR mutant phenotypes could be rescued with a double mutation of irvA or complemented by introducing a wild-type copy of irvR on a shuttle vector. These data indicate that the repression of irvA is critically dependent upon irvR and that irvA repression is essential for the development of genetic competence and the proper control of DDAG in S. mutans.
doi:10.1128/JB.00967-08
PMCID: PMC2580701  PMID: 18757533

Results 1-9 (9)