Ressenyes

Obra ressenyada: Edward W. SOJA, Seeking Spatial Justice.Minneapolis : University of Minnesota Press, 2010

Structural, functional, and genetic analyses of the actinobacterial transcription factor RbpA

Gene expression is highly regulated at the step of transcription initiation, and transcription activators play a critical role in this process. RbpA, an actinobacterial transcription activator that is essential in Mycobacterium tuberculosis (Mtb), binds selectively to group 1 and certain group 2 σ-factors. To delineate the molecular mechanism of RbpA, we show that the Mtb RbpA σ-interacting domain (SID) and basic linker are sufficient for transcription activation. We also present the crystal structure of the Mtb RbpA-SID in complex with domain 2 of the housekeeping σ-factor, σ(A). The structure explains the basis of σ-selectivity by RbpA, showing that RbpA interacts with conserved regions of σ(A) as well as the nonconserved region (NCR), which is present only in housekeeping σ-factors. Thus, the structure is the first, to our knowledge, to show a protein interacting with the NCR of a σ-factor. We confirm the basis of selectivity and the observed interactions using mutagenesis and functional studies. In addition, the structure allows for a model of the RbpA-SID in the context of a transcription initiation complex. Unexpectedly, the structural modeling suggests that RbpA contacts the promoter DNA, and we present in vivo and in vitro studies supporting this finding. Our combined data lead to a better understanding of the mechanism of RbpA function as a transcription activator

Structural and Biophysical Studies on Two Promoter Recognition Domains of the Extra-cytoplasmic Function σ Factor σC from Mycobacterium tuberculosis

σ factors are transcriptional regulatory proteins that bind to the RNA polymerase and dictate gene expression. The extracytoplasmic function σ factors (ECF) govern the environment dependent regulation of transcription. ECF σ factors have two domains σ(2) and σ(4) that recognize the −10 and −35 promoter elements. However, unlike the primary σ factor σ(A), the ECF σ factors lack σ(3), a region that helps in the recognition of the extended −10 element and σ(1.1), a domain involved in the auto-inhibition of σ(A) in the absence of core RNA polymerase. Mycobacterium tuberculosis σ(C) is an ECF σ factor that is essential for the pathogenesis and virulence of M. tuberculosis in the mouse and guinea pig models of infection. However, unlike other ECF σ factors, σ(C) does not appear to have a regulatory anti-σ factor located in the same operon. We also note that Mycobacterium tuberculosis σ(C) differs from the canonical ECF σ factors as it has an N-terminal domain comprising of 126 amino acids that precedes the σ(C)(2) and σ(C)(4) domains. In an effort to understand the regulatory mechanism of this protein, the crystal structures of the σ(C)(2) and σ(C)(4) domains of σ(C) were determined. These promoter recognition domains are structurally similar to the corresponding domains of σ(A) despite the low sequence similarity. Fluorescence experiments using the intrinsic tryptophan residues of σ(C)(2) as well as surface plasmon resonance measurements reveal that the σ(C)(2) and σ(C)(4) domains interact with each other. Mutational analysis suggests that the Pribnow box binding region of σ(C)(2) is involved in this inter-domain interaction. Interaction between the promoter recognition domains in M. tuberculosis σ(C) are thus likely to regulate the activity of this protein even in the absence of an anti-σ factor

Bacterial Stress Responses: What Doesn't Kill Them Can Make Them Stronger

Identifying specific mechanisms that contribute to microbial survival under rapidly changing conditions could provide insight into stress response systems across life forms

The Structural Basis for Promoter −35 Element Recognition by the Group IV σ Factors

The control of bacterial transcription initiation depends on a primary σ factor for housekeeping functions, as well as alternative σ factors that control regulons in response to environmental stresses. The largest and most diverse subgroup of alternative σ factors, the group IV extracytoplasmic function σ factors, directs the transcription of genes that regulate a wide variety of responses, including envelope stress and pathogenesis. We determined the 2.3-Å resolution crystal structure of the −35 element recognition domain of a group IV σ factor, Escherichia coli σ(E) (4), bound to its consensus −35 element, GGAACTT. Despite similar function and secondary structure, the primary and group IV σ factors recognize their −35 elements using distinct mechanisms. Conserved sequence elements of the σ(E) −35 element induce a DNA geometry characteristic of AA/TT-tract DNA, including a rigid, straight double-helical axis and a narrow minor groove. For this reason, the highly conserved AA in the middle of the GGAACTT motif is essential for −35 element recognition by σ(E) (4), despite the absence of direct protein–DNA interactions with these DNA bases. These principles of σ(E) (4)/−35 element recognition can be applied to a wide range of other group IV σ factors

Adiponectin, diabetes and ischemic heart failure: a challenging relationship

Abstract Background Several peptides, named adipokines, are produced by the adipose tissue. Among those, adiponectin (AD) is the most abundant. AD promotes peripheral insulin sensitivity, inhibits liver gluconeogenesis and displays anti-atherogenic and anti-inflammatory properties. Lower levels of AD are related to a higher risk of myocardial infarction and a worse prognosis in patients with coronary artery disease. However, despite a favorable clinical profile, AD increases in relation to worsening heart failure (HF); in this context, higher adiponectinemia is reliably related to poor prognosis. There is still little knowledge about how certain metabolic conditions, such as diabetes mellitus, modulate the relationship between AD and HF. We evaluated the level of adiponectin in patients with ischemic HF, with and without type 2 diabetes, to elucidate whether the metabolic syndrome was able to influence the relationship between AD and HF. Results We demonstrated that AD rises in patients with advanced HF, but to a lesser extent in diabetics than in non-diabetics. Diabetic patients with reduced systolic performance orchestrated a slower rise of AD which began only in face of overt HF. The different behavior of AD in the presence of diabetes was not entirely explained by differences in body mass index. In addition, NT-proBNP, the second strongest predictor of AD, did not differ significantly between diabetic and non-diabetic patients. These data indicate that some other mechanisms are involved in the regulation of AD in patients with type 2 diabetes and coronary artery disease. Conclusions AD rises across chronic heart failure stages but this phenomenon is less evident in type 2 diabetic patients. In the presence of diabetes, the progressive increase of AD in relation to the severity of LV dysfunction is hampered and becomes evident only in overt HF.</p

Stringent promoter recognition and autoregulation by the group 3 σ-factor SigF in the cyanobacterium Synechocystis sp. strain PCC 6803

The cyanobacteirum Synechocystis sp. strain PCC 6803 possesses nine species of the sigma (σ)-factor gene for RNA polymerase (RNAP). Here, we identify and characterize the novel-type promoter recognized by a group 3 σ-factor, SigF. SigF autoregulates its own transcription and recognizes the promoter of pilA1 that acts in pilus formation and motility in PCC 6803. The pilA1 promoter (PpilA1-54) was recognized only by SigF and not by other σ-factors in PCC 6803. No PpilA1-54 activity was observed in Escherichia coli cells that possess RpoF (σ28) for fragellin and motility. Studies of in vitro transcription for PpilA1-54 identified the region from −39 to −7 including an AG-rich stretch and a core promoter with TAGGC (−32 region) and GGTAA (−12 region) as important for transcription. We also confirmed the unique PpilA1-54 architecture and further identified two novel promoters, recognized by SigF, for genes encoding periplasmic and phytochrome-like phototaxis proteins. These results and a phylogenetic analysis suggest that the PCC 6803 SigF is distinct from the E. coli RpoF or RpoD (σ70) type and constitutes a novel eubacterial group 3 σ-factor. We discuss a model case of stringent promoter recognition by SigF. Promoter types of PCC 6803 genes are also summarized

RinA controls phage-mediated packaging and transfer of virulence genes in Gram-positive bacteria

Phage-mediated transfer of microbial genetic elements plays a crucial role in bacterial life style and evolution. In this study, we identify the RinA family of phage-encoded proteins as activators required for transcription of the late operon in a large group of temperate staphylococcal phages. RinA binds to a tightly regulated promoter region, situated upstream of the terS gene, that controls expression of the morphogenetic and lysis modules of the phage, activating their transcription. As expected, rinA deletion eliminated formation of functional phage particles and significantly decreased the transfer of phage and pathogenicity island encoded virulence factors. A genetic analysis of the late promoter region showed that a fragment of 272 bp contains both the promoter and the region necessary for activation by RinA. In addition, we demonstrated that RinA is the only phage-encoded protein required for the activation of this promoter region. This region was shown to be divergent among different phages. Consequently, phages with divergent promoter regions carried allelic variants of the RinA protein, which specifically recognize its own promoter sequence. Finally, most Gram-postive bacteria carry bacteriophages encoding RinA homologue proteins. Characterization of several of these proteins demonstrated that control by RinA of the phage-mediated packaging and transfer of virulence factor is a conserved mechanism regulating horizontal gene transfer