Repression by the H-NS/YmoA histone-like protein complex enables IscR dependent regulation of the Yersinia T3SS.

The type III secretion system (T3SS) is an appendage used by many bacterial pathogens, such as pathogenic Yersinia, to subvert host defenses. However, because the T3SS is energetically costly and immunogenic, it must be tightly regulated in response to environmental cues to enable survival in the host. Here we show that expression of the Yersinia Ysc T3SS master regulator, LcrF, is orchestrated by the opposing activities of the repressive H-NS/YmoA histone-like protein complex and induction by the iron and oxygen-regulated IscR transcription factor. While deletion of iscR or ymoA has been shown to decrease and increase LcrF expression and type III secretion, respectively, the role of H-NS in this system has not been definitively established because hns is an essential gene in Yersinia. Using CRISPRi knockdown of hns, we show that hns depletion causes derepression of lcrF. Furthermore, we find that while YmoA is dispensable for H-NS binding to the lcrF promoter, YmoA binding to H-NS is important for H-NS repressive activity. We bioinformatically identified three H-NS binding regions within the lcrF promoter and demonstrate binding of H-NS to these sites in vivo using chromatin immunoprecipitation. Using promoter truncation and binding site mutation analysis, we show that two of these H-NS binding regions are important for H-NS/YmoA-mediated repression of the lcrF promoter. Surprisingly, we find that IscR is dispensable for lcrF transcription in the absence of H-NS/YmoA. Indeed, IscR-dependent regulation of LcrF and type III secretion in response to changes in oxygen, such as those Yersinia is predicted to experience during host infection, only occurs in the presence of an H-NS/YmoA complex. These data suggest that, in the presence of host tissue cues that drive sufficient IscR expression, IscR can act as a roadblock to H-NS/YmoA-dependent repression of RNA polymerase at the lcrF promoter to turn on T3SS expression.

Iron availability and oxygen tension regulate the Yersinia Ysc type III secretion system to enable disseminated infection.

The enteropathogen Yersinia pseudotuberculosis and the related plague agent Y. pestis require the Ysc type III secretion system (T3SS) to subvert phagocyte defense mechanisms and cause disease. Yet type III secretion (T3S) in Yersinia induces growth arrest and innate immune recognition, necessitating tight regulation of the T3SS. Here we show that Y. pseudotuberculosis T3SS expression is kept low under anaerobic, iron-rich conditions, such as those found in the intestinal lumen where the Yersinia T3SS is not required for growth. In contrast, the Yersinia T3SS is expressed under aerobic or anaerobic, iron-poor conditions, such as those encountered by Yersinia once they cross the epithelial barrier and encounter phagocytic cells. We further show that the [2Fe-2S] containing transcription factor, IscR, mediates this oxygen and iron regulation of the T3SS by controlling transcription of the T3SS master regulator LcrF. IscR binds directly to the lcrF promoter and, importantly, a mutation that prevents this binding leads to decreased disseminated infection of Y. pseudotuberculosis but does not perturb intestinal colonization. Similar to E. coli, Y. pseudotuberculosis uses the Fe-S cluster occupancy of IscR as a readout of oxygen and iron conditions that impact cellular Fe-S cluster homeostasis. We propose that Y. pseudotuberculosis has coopted this system to sense entry into deeper tissues and induce T3S where it is required for virulence. The IscR binding site in the lcrF promoter is completely conserved between Y. pseudotuberculosis and Y. pestis. Deletion of iscR in Y. pestis leads to drastic disruption of T3S, suggesting that IscR control of the T3SS evolved before Y. pestis split from Y. pseudotuberculosis.

Motivated exploratory behaviour in the rat: the role of hippocampus and the histaminergic neurotransmission.

Exploration is one of most basic adaptive behavioural responses, giving the animal an important evolutionary advantage to survive in a changing environment. Inspection of novel environments might be come with motivated exploratory behaviour. In spite that this type of exploration in the rat is known for many years, little attention has been given to the intrinsic mechanisms or the brain structures that are involved in. In the present work the hippocampus, the neurotransmitter histamine, and the geometrical features of novel objects were examined in a model of conflictive and non-conflictive exploration in the rat which evaluates incentive-motivated exploration. Young adult intact rats were tested in a neutral non-conflictive behavioural activity detector (OVM), with (eOVM) or without (sOVM) novel objects. Three different objects were used: a box, a toy duck, and a tower. Results show that animals decrease its general motor activity (horizontal, ambulatory and non-ambulatory activity) in favor to exploration of the objects. Motivated exploration was not the same for all three objects. Rats explored significantly more the Tower and the Box objects than the Duck item. Behavioral patterns of hippocampus-implanted rats showed decreased scores in motor activity but maintained the difference in the relation of "without/with objects" exploration. When hippocampus-implanted rats were tested in a conflictive exploration device (the elevated asymmetric plus-maze), exploration of the No Wall arm, considered the most fear-inducing environment, was significantly more explored by the animal when the tower object was positioned at its end than when it was absent. Microinjection into the ventral hippocampus of histamine abolished this motivated exploratory response. Pre-treatment with pyrilamine, and not with ranitidine, was effective to block the inhibitory effect of histamine on the object motivated exploration. Results confirm that the hippocampus is involved on incentive motivated exploration, and suggest that histamine is part of an analyzing neuronal circuit of novelty incentivating behavioural responses in rats.