Release of paused RNA polymerase II (Pol II) requires incorporation of the positive transcription elongation factor b (P-TEFb) into the super elongation complex (SEC), thus resulting in rapid yet synchronous transcriptional activation. However, the mechanism underlying dynamic transition of P-TEFb from inactive to active state remains unclear. Here, we found that the SEC components are able to compartmentalize and concentrate P-TEFb via liquid-liquid phase separation from the soluble inactive HEXIM1 containing the P-TEFb complex. Specifically, ENL or its intrinsically disordered region is sufficient to initiate the liquid droplet formation of SEC. AFF4 functions together with ENL in fluidizing SEC droplets. SEC droplets are fast and dynamically formed upon serum exposure and required for rapid transcriptional induction. We also found that the fusion of ENL with MLL can boost SEC phase separation. In summary, our results suggest a critical role of multivalent phase separation of SEC in controlling transcriptional pause release.

Background: Accurate classification of different Burkholderia cepacia complex (BCC) species is essential for therapy, prognosis assessment and research. The taxonomic status of BCC remains problematic and an improved knowledge about the classification of BCC is in particular needed. Methods: We compared phylogenetic trees of BCC based on 16S rRNA, recA, hisA and MLSA (multilocus sequence analysis). Using the available whole genome sequences of BCC, we inferred a species tree based on estimated single-copy orthologous genes and demarcated species of BCC using dDDH/ANI clustering. Results: We showed that 16S rRNA, recA, hisA and MLSA have limited resolutions in the taxonomic study of closely related bacteria such as BCC. Our estimated species tree and dDDH/ANI clustering clearly separated 116 BCC strains into 36 clusters. With the appropriate reclassification of misidentified strains, these clusters corresponded to 22 known species as well as 14 putative novel species. Conclusions: This is the first large-scale and systematic study of the taxonomic status of the BCC and could contribute to further insights into BCC taxonomy. Our study suggested that conjunctive use of core phylogeny based on single-copy orthologous genes, as well as pangenome-based dDDH/ANI clustering would provide a preferable framework for demarcating closely related species. Reviewer: This article was reviewed by Dr. Xianwen Ren.

BackgroundSome microorganisms can produce pigments such as melanin, which has been associated with virulence in the host and with a survival advantage in the environment. In Vibrio cholerae, studies have shown that pigment-producing mutants are more virulent than the parental strain in terms of increased UV resistance, production of major virulence factors, and colonization. To date, almost all of the pigmented V. cholerae strains investigated have been induced by chemicals, culture stress, or transposon mutagenesis. However, during our cholera surveillance, some nontoxigenic serogroup O139 strains and one toxigenic O1 strain, which can produce pigment steadily under the commonly used experimental growth conditions, were obtained in different years and from different areas. The genes VC1344 to VC1347, which correspond to the El Tor strain N16961 genome and which comprise an operon in the tyrosine catabolic pathway, have been confirmed to be associated with a pigmented phenotype. In the present study, we investigated the mechanism of pigment production in these strains.ResultsSequencing of the VC1344, VC1345, VC1346, and VC1347 genes in these pigmented strains suggested that a deletion mutation in the homogentisate oxygenase gene (VC1345) may be associated with the pigmented phenotype, and gene complementation confirmed the role of this gene in pigment production. An identical 15-bp deletion was found in the VC1345 gene of all six O139 pigment-producing strains examined, and a 10-bp deletion was found in the VC1345 gene of the O1 strain. Strict sequence conservation in the VC1344 gene but higher variance in the other three genes of this operon were observed, indicating the different stress response functions of these genes in environmental adaption and selection. On the basis of pulsed-field gel electrophoresis typing, the pigment-producing O139 strains showed high clonality, even though they were isolated in different years and from different regions. Additionally all these O139 strains belong to the rb4 ribotype, which contains the O139 strains isolated from diarrheal patients, although these strains are cholera toxin negative.ConclusionDysfunction of homogentisate oxygenase (VC1345) causes homogentisate accumulation and pigment formation in naturally pigmented strains of V. cholerae. The high clonality of these strains may correlate to an environmental survival advantage in the V. cholerae community due to their pigment production, and may imply a potential protective function of melanin in environmental survival of such strains.

In the process of host–pathogen interactions, bacterial pathogens always employ some special genes, e.g., virulence factors (VFs) to interact with host and cause damage or diseases to host. A number of VFs have been identified in bacterial pathogens that confer upon bacterial pathogens the ability to cause various types of damage or diseases. However, it has been clarified that some of the identified VFs are also encoded in the genomes of nonpathogenic bacteria, and this finding gives rise to considerable controversy about the definition of virulence factor. Here 1988 virulence factors of 51 sequenced pathogenic bacterial genomes from the virulence factor database (VFDB) were collected, and an orthologous comparison to a non-pathogenic bacteria protein database was conducted using the reciprocal-best-BLAST-hits approach. Six hundred and twenty pathogen-specific VFs and 1368 common VFs (present in both pathogens and nonpathogens) were identified, which account for 31.19% and 68.81% of the total VFs, respectively. The distribution of pathogen-specific VFs and common VFs in pathogenicity islands (PAIs) was systematically investigated, and pathogen-specific VFs were more likely to be located in PAIs than common VFs. The function of the two classes of VFs were also analyzed and compared in depth. Our results indicated that most but not all T3SS proteins are pathogen-specific. T3SS effector proteins tended to be distributed in pathogen-specific VFs, whereas T3SS translocation proteins, apparatus proteins, and chaperones were inclined to be distributed in common VFs. We also observed that exotoxins were located in both pathogen-specific and common VFs. In addition, the architecture of the two classes of VFs was compared, and the results indicated that common VFs had a higher domain number and lower domain coverage value, revealed that common VFs tend to be more complex and less compact proteins.

Vibrio cholerae O1 El Tor biotype strains are responsible for three multiyear epidemics of cholera in China during the seventh ongoing pandemic. The presence of the integrative conjugative element SXT is strongly correlated with resistance to nalidixic acid, tetracycline, and trimethoprim-sulfamethoxazole in these strains. Here, we sequenced the conserved genes of the SXT element, including eex, setR, and int, from 59 V. cholerae O1 El Tor strains and extracted and assembled the intact SXT sequences from the 11 genome sequenced strains. These elements had characteristics distinct from those of previously reported integrative conjugative elements (ICEs). They could be clearly divided into two types based on the clustering of conserved genes and gene structures of the elements, showing their possibly independent derivation and evolution. These two types were present before and after 2005, respectively, demonstrating the type substitution that occurred in 2005. Four to six antibiotic-resistant genes were found on the SXT elements, including genes resistant to tetracycline, trimethoprim-sulfamethoxazole, and multiple drugs. In summary, our findings demonstrated the roles of the SXT element in the emergence of multidrug resistance in epidemic O1 El Tor V. cholerae strains in China.