Furthermore, vaccination of mice with the ΔyscN mutant provided some level of protection against a s.c. challenge (the equivalent of ~90LD50) with the wild-type strain for even the group vaccinated with the lowest mutant dose. Following two vaccinations with varying doses of the ΔyscN mutant, quantitative anti-F1 and anti-LcrV ELISA were performed with sera collected from the vaccinated mice. As expected for a yscN mutant, no increase in the immune response to LcrV was determined. Variability in the quantitative anti-F1 ELISA titers as demonstrated by the high standard deviations was reflected somewhat in the flattened survival results and may be

the result of testing only three mice per dosage group. Variation in antibody titers has also been reported by others

using live mutant Y. pestis vaccine strains (Okan et al., 2010; Ion Channel Ligand Library clinical trial Oyston et al., 2010). These results may suggest that with this live vaccine strain, anti-F1 titers may not be solely protective and that other bacterial antigens or cytokine-mediated immunity (Kummer et al., 2008) may also play a concerted role in protection. The humoral immune response against Y. pestis is directed against multiple proteins, many encoded by genes on the virulence plasmids (Benner et al., 1999). Among them, the acquired immunity to F1 and LcrV is sufficient to typically protect against plague (Powell et al., 2005). However, the emergence of atypical F1 mutants fully virulent in humans and with natural heterogeneity to Y. pestis LcrV highlights the limits Protease Inhibitor Library cell line of the current rF1-V fusion vaccine (Quenee et al., 2008). In conclusion, future work with use of the ΔyscN mutant as a live vaccine should proceed. The current study provides initial steps toward this goal. To further characterize the use of this strain as a potential vaccine, many other studies would need to be completed, such as histopathological analysis

of the vaccinated mice. In addition, testing for protection tetracosactide against pneumonic plague would need to be explored. It is not uncommon for mutant strains of Y. pestis to be attenuated in bubonic models but still retain virulence in pneumonic challenges (Friedlander et al., 1995; Welkos et al., 1995, 1997; Worsham & Roy, 2003; Cathelyn et al., 2006; Bozue et al., 2011). We thank Brad Stiles and Susan Welkos for review of this manuscript, and Diane Fisher for completing the statistical analysis of this study. This work was funded by the Defense Threat Reduction Agency (project 2.10019_08_RD_B to W.S.). Research was conducted in compliance with the Animal Welfare Act and other federal statutes and regulations relevant to animals and experiments using animals and complies with all principles stated in the Guide for the Care and Use of Laboratory Animals (National Research Council, 1996). The research facility used is fully accredited by the Association for Assessment and Accreditation of Laboratory Care International.

Here, we demonstrate a new link between plasmid carriage, biofilm formation, and eDNA for P. putida KT2440. The potential universality and molecular mechanism by which CDK inhibitor TOL carriage results in excess eDNA remains, so far, unresolved but do not appear to be related to enhanced cell lysis, and suggest secretion. Additional studies will be required to examine the exact mechanism of eDNA release and the nature of the released eDNA associated with TOL carriage in P. putida KT22440. This study was supported by an EC-FP6 Marie Curie Excellence Grant (MEXT-CT-2005-024004) to B.F.S. and the Villum Kan Rasmussen Center for Environmental and

Agricultural Microbiology. We thank N. Kroer and S. Molin for providing strains and plasmids, B.S. Lauritzen for plasmid tagging, and N. El Azhari for initial flow cell observations. Fig. S1. Observation of little and abundant pellicle formation in 5-day-old static cultures of Pseudomonas putida

KT2440 and KT2440. Fig. S2. Micrographs of 1–7-day-old click here Pseudomonas putida KT2440 pellicles, with and without TOL plasmid, grown in presence or absence of DNase I. Please note: Wiley-Blackwell is not responsible for the content or functionality of any supporting materials supplied by the authors. Any queries (other than missing material) should be directed to the corresponding author for the article. “
“Of the five dd-carboxypeptidases in Escherichia coli, only PBP5 demonstrates its physiological significance by maintaining cell shape and intrinsic beta-lactam resistance. DacD can partially compensate for the lost beta-lactam resistance in PBP5 mutant, although its biochemical reason is unclear. To understand the mechanism(s) underlying such behaviour, we constructed soluble DacD (sDacD) and compared its biophysical and (-)-p-Bromotetramisole Oxalate biochemical properties with those of sPBP5, in vitro. Unlike sPBP6, sDacD can deacylate Bocillin significantly, which is very similar to sPBP5. sDacD shows weak dd-carboxypeptidase activity, although lower than that of sPBP5. Bioinformatics analyses reveal a similar architecture of sPBP5 and sDacD. Therefore, based on the obtained results we can infer that biochemically

DacD and PBP5 are more closely related to each other than to PBP6, enabling DacD and PBP5 to play a nearly similar physiological function in terms of recovering the lost beta-lactam resistance. Of the five dd-carboxypeptidases (DD-CPases) in Escherichia coli, PBP4, PBP5, PBP6, DacD and AmpH (Holtje, 1998; Ghosh et al., 2008; Sauvage et al., 2008; Gonzalez-Leiza et al., 2011), only PBP5 has been studied thoroughly concerning enzymology, structure and physiological aspects (Nelson & Young, 2000; Nelson & Young, 2001; Chowdhury et al., 2010; Sarkar et al., 2010, 2011). However, other DD-CPases are mostly characterized on the basis of their kinetic properties in vitro (Korat et al., 1991; Chowdhury et al., 2010; Gonzalez-Leiza et al.

Using a fourfold cut-off,

we observed that 237 genes were differentially expressed (data not shown). To reduce reporting of false positives, we chose the higher cut-off, where the expression patterns of biological replicates (from the two animals) were similar (Fig. 1), suggesting that the differences observed are the representative of expression in vivo. Thirteen of the 44 genes encode proteins of unknown function. This is not surprising, as 31% of the coding sequences in the M. hemolytica A1 genome were annotated as hypothetical proteins (Gioia et al., 2006). In the family Pasteurellaceae, a large proportion of genes that were differentially expressed in other published microarray studies do not have a prescribed function, thus their products have been annotated as hypothetical proteins (Boyce et al., 2002, 2004; Melnikow et al., 2005; Deslandes et al., 2007, 2010). Interestingly, the majority of the Enzalutamide clinical trial genes (13/17) showing higher expression in vivo encode proteins of unknown function. A similar result was reported for Actinobacillus pleuropneumoniae grown under in vitro iron-restricted conditions PS-341 solubility dmso (Deslandes et al., 2007). In Helicobacter pylori, 10 of 14 genes encoding hypothetical proteins were transcribed in vivo and not in vitro (Graham et al., 2002). Two of the 11 hypothetical proteins (MHA_0428 and MHA_2589) are unique to M. hemolytica A1 but their

roles in bovine pneumonic pasteurellosis are not known. The challenge that most array-based studies have to face is identifying and characterizing genes of interest from a large number of genes encoding proteins of uncharacterized function. In this study, the hypothetical BCKDHA proteins identified show a comparatively high level of

expression in vivo (8- to 37-fold), 6 days after challenge. Three genes encoding components of the Mu-like bacteriophage, discovered in strain ATCC BAA-410 (Gioia et al., 2006), were up-regulated in vivo. Two bacteriophage-associated genes were also differentially expressed in an in vivo study of A. pleuropneumoniae (Deslandes et al., 2010). These genes are as follows: a putative lipoprotein gene (MHA_2737) showing identity to an A. pleuropneumoniae gene (ZP_00134432) and an Actinobacillus minor gene (ZP_03612071). More than 12% of the M. hemolytica A1 genome has been annotated as bacteriophage-related (Gioia et al., 2006). The Mu-related prophage sequence is incomplete in the draft genome sequence and mapped at the end of a scaffold. At a less stringent cut-off, we observed increased expression of many other genes from this phage in vivo (data not shown) suggesting that the entire sequence may represent a complete and potentially active prophage. We observed a 12-fold increase in the expression of a gene coding for a putative lipoprotein with a predicted molecular mass of approximately 22 kDa. The amino acid sequence for the putative lipoprotein has identity to a predicted periplasmic or secreted proteins in A.

Attack rates among Dutch travelers to developing regions declined for hepatitis

A, shigellosis, and typhoid fever. Region-specific trends in attack rates of shigellosis resembled trends of hepatitis A and typhoid fever. Declining attack rates of the three fecal-orally transmitted diseases correlated Antidiabetic Compound Library with improvements in socioeconomic, sanitary, and water supply conditions of the local population at travel destination. Conclusions. These findings suggest that improved hygienic standards at travel destination strongly contributed to the overall decline in attack rates of fecal-orally transmitted diseases among visiting travelers. In industrialized countries, the incidence of fecal-orally transmitted infections, such as hepatitis A, typhoid fever, and shigellosis, has declined substantially.1–4 Currently, most cases in these countries arise from visits to non-industrialized countries.2 A few studies have addressed trends in hepatitis A or typhoid fever among international travelers, BIBF 1120 finding that, over the past decades, their risk of acquiring

hepatitis A or typhoid fever has decreased.5–7 This decline is often attributed to pretravel vaccination and improvements in hygienic and sanitary conditions at travel destinations. However, their absolute or relative contributions are unknown, given the lack of studies on the influence of hygienic factors on the incidence of fecal-orally transmitted diseases. This study analyzes region-specific trends in attack rates of hepatitis A, typhoid fever, and shigellosis among Dutch travelers, combining Dutch travelers’ statistics with information from the national infectious diseases notification system. All three diseases are transmitted through fecally contaminated water or food. Hepatitis A virus (HAV) infection causes an acute viral liver disease and confers lifelong immunity.

It is a common childhood disease in developing countries, but the prevalence of hepatitis A antibodies is low in developed regions.1,8 In the Netherlands, an inactivated HAV vaccine is available HSP90 for risk groups, such as travelers, and is almost 100% effective.9 Typhoid fever is a bacterial systemic infection caused by Salmonella enterica serotype Typhi.3,10 Immunity following infection is limited and can be overcome.9 Two parenteral capsular polysaccharide vaccines are available in the Netherlands, and studies report their efficacy at 35% to 70%.11 Shigellosis is a bacterial enteric disease, caused by one of the four serogroups of Shigella. Immunity following infection is type-specific and probably limited.4 No vaccine is available. To study if the attack rates of fecal-orally transmitted diseases in travelers are influenced by improvements in hygienic standards at travel destinations, we compared trends in vaccine-preventable hepatitis A and typhoid fever with trends in non-vaccine-preventable shigellosis.

Attack rates among Dutch travelers to developing regions declined for hepatitis

A, shigellosis, and typhoid fever. Region-specific trends in attack rates of shigellosis resembled trends of hepatitis A and typhoid fever. Declining attack rates of the three fecal-orally transmitted diseases correlated selleck chemicals with improvements in socioeconomic, sanitary, and water supply conditions of the local population at travel destination. Conclusions. These findings suggest that improved hygienic standards at travel destination strongly contributed to the overall decline in attack rates of fecal-orally transmitted diseases among visiting travelers. In industrialized countries, the incidence of fecal-orally transmitted infections, such as hepatitis A, typhoid fever, and shigellosis, has declined substantially.1–4 Currently, most cases in these countries arise from visits to non-industrialized countries.2 A few studies have addressed trends in hepatitis A or typhoid fever among international travelers, Selleckchem PI3K inhibitor finding that, over the past decades, their risk of acquiring

hepatitis A or typhoid fever has decreased.5–7 This decline is often attributed to pretravel vaccination and improvements in hygienic and sanitary conditions at travel destinations. However, their absolute or relative contributions are unknown, given the lack of studies on the influence of hygienic factors on the incidence of fecal-orally transmitted diseases. This study analyzes region-specific trends in attack rates of hepatitis A, typhoid fever, and shigellosis among Dutch travelers, combining Dutch travelers’ statistics with information from the national infectious diseases notification system. All three diseases are transmitted through fecally contaminated water or food. Hepatitis A virus (HAV) infection causes an acute viral liver disease and confers lifelong immunity.

It is a common childhood disease in developing countries, but the prevalence of hepatitis A antibodies is low in developed regions.1,8 In the Netherlands, an inactivated HAV vaccine is available Montelukast Sodium for risk groups, such as travelers, and is almost 100% effective.9 Typhoid fever is a bacterial systemic infection caused by Salmonella enterica serotype Typhi.3,10 Immunity following infection is limited and can be overcome.9 Two parenteral capsular polysaccharide vaccines are available in the Netherlands, and studies report their efficacy at 35% to 70%.11 Shigellosis is a bacterial enteric disease, caused by one of the four serogroups of Shigella. Immunity following infection is type-specific and probably limited.4 No vaccine is available. To study if the attack rates of fecal-orally transmitted diseases in travelers are influenced by improvements in hygienic standards at travel destinations, we compared trends in vaccine-preventable hepatitis A and typhoid fever with trends in non-vaccine-preventable shigellosis.

, 2009). For this reason, we hypothesized that their facilitation of substance P release was caused by disinhibition, that is, that CB1 receptors inhibit the buy Olaparib release of neurotransmitters that decrease substance P release. Two important inhibitors of substance P release are GABA, acting on GABAB receptors (Malcangio & Bowery, 1993; Marvizon et al., 1999; Riley et al., 2001; Lao et al., 2003), and opioids, acting on μ-opioid receptors (Yaksh et al., 1980; Kondo et al., 2005). CB1 receptors could inhibit GABA or opioid release in the dorsal horn. In this case, and given that endocannabinoids are released during dorsal root stimulation, CB1 antagonists would increase GABA or opioid release, resulting

in an inhibition of substance P release mediated by GABAB or μ-opioid receptors, respectively. This hypothesis predicts that the inhibition produced by AM251 would be reversed by GABAB or μ-opioid receptor antagonists. This prediction was tested in the experiment in Fig. 9, in which we used the selective μ-opioid receptor antagonist CTAP (10 μm) and the GABAB receptor antagonist CGP55845 (100 nm). In previous studies in spinal cord slices we determined that these concentrations of CTAP and CGP55845 produce a complete blockade of μ-opioid receptors (Song & Marvizon, click here 2003) and GABAB receptors (Lao & Marvizon, 2005), respectively. Spinal cord slices were electrically stimulated at the

dorsal root at 100 Hz or 1 Hz, because different frequencies of root stimulation evoke different patterns of neurotransmitter release in the dorsal horn (Marvizon et al., 1999; Lever et al., 2001; Lao & Marvizon, 2005). When the dorsal root was stimulated at 100 Hz (Fig. 9A), the inhibition produced by AM251 (100 nm) was reversed by CTAP but not Chlormezanone by CGP55845. This suggests that during high-frequency stimulation AM251 increases opioid release, leading to inhibition of substance P release mediated by μ-opioid receptors. Two-way anova for the

data in Fig. 9A revealed significant effects of the variables ‘drugs’ (F5 = 21, P < 0.0001) and ‘stimulus’ (F1 = 1352, P < 0.0001), and a significant interaction between them (F5 = 20, P < 0.0001). When the dorsal root was stimulated at 1 Hz (Fig. 9B), the inhibition produced by AM251 (100 nm) was reversed by both CTAP and CGP55845 (100 nm). This suggests that during low-frequency stimulation AM251 increases both opioid and GABA release, leading to inhibition of substance P release mediated by μ-opioid receptors and GABAB receptors. Two-way anova for the data in Fig. 9B revealed significant effects of the variables ‘drugs’ (F5 = 2.5, P = 0.041) and ‘stimulus’ (F1 = 581, P < 0.0001) and a significant interaction between them (F5 = 3.3, P = 0.012). Neither CTAP nor CGP55845 alone affected NK1R internalization evoked with either 100 Hz or 1 Hz stimulation (Fig. 9), indicating that the stimulus elicited little opioid or GABA release in these conditions.

This functional difference between the uvrABbu and the uvrAEco gene products is not surprising given the evolutionary distance between E. coli and B. burgdorferi (Wu et al., 2009). Borrelia burgdorferiΔuvrABbu was significantly more susceptible to H2O2 than the wild-type parental strain (Table 2), with the MIC of H2O2 for the wild-type B. burgdorferi being as much as fivefold higher than that of the ΔuvrABbu mutant. Anti-diabetic Compound Library This increased sensitivity to ROS was partially reversed by complementation with either pAB63 or pMS9 (Table 2). Complementation was not affected by the presence of 3% CO2 (studies using pAB63), or its absence (studies using pMS9), during culture. Because the

inserted kanamycin resistance gene contained its own stop codon, it seems unlikely that polarity effects on the downstream BB0838 gene contributed to the phenotype of the ΔuvrABbu mutant. However, homologues of BB0838 are present in other Borrelia as well as in Treponema, and because the function of this hypothetical protein is unknown, it is not possible to give a definitive answer. In contrast to the sensitivity of the ΔuvrABbu mutant to ROS, its growth and that of its derivatives was not inhibited by exposure to NaNO2, SNAP or SPER/NO (Table 2). The lack of effect of exposure to any of these RNS generators on B. burgdorferi growth even though

exposure to SNAP and SPER/NO was in PBS while exposure to NaNO2 was in BSK-H suggests that this lack of effect was not likely caused Afatinib chemical structure buy Ixazomib by the serum component of BSK-H (Sohaskey & Barbour, 1999). There were also no significant differences in growth of B. burgdorferi and its derivatives in complete BSK-H at pH 6.0, 6.5 or 6.8 (Table 2). None of the strains used in the study (wild-type, uvrA inactivation mutant, complemented mutants) were able to grow at pH 5.5 in complete BSK-H (data not shown).

The ability of pathogenic bacteria to repair challenges to their genomes from various DNA-damaging agents produced by host phagocytes is critical to their survival in their hosts (Fang, 2004; Steere et al., 2004). In the absence of DNA-damaging agents, the uvrABbu inactivation mutant grew as well as the wild-type strain but was markedly inhibited by exposure to UV radiation (Fig. 2), MMC (Fig. 3a) and ROS (Table 2). Extrachromosomal complementation with wild-type uvrABbu restored growth. In contrast, growth of the inactivation mutant was identical to that of the wild-type after exposure to RNS or decreased pH, conditions under which uvrA has been shown to be protective in other bacteria (Aertsen et al., 2004; Fang, 2004). The uvrABbu gene product is thus involved in the repair of DNA damage caused by UV radiation, ROS and MMC in B. burgdorferi, but not involved in damage due to RNS or decreased pH. Repair of DNA damage caused by UV irradiation involves UvrA recognition of this damage and nucleotide excision (Sancar, 1996; Savery, 2007).

Cells were pelleted by centrifugation and resuspended in 20 mL of buffer A (20 mM HEPES pH 7.9, 10% glycerol, 100 mM KCl, 5 mM MgCl2, 20 mM imidazole). Cells were lysed by three passages through a French Press at 1000 psi. His-tagged protein was purified by nickel chelate affinity chromatography using Ni-NTA resin (Qiagen)

under batch conditions. A fragment containing the intergenic region between yfeR and yfeH (89 bp) and 221 bp of the yfeH gene, generated by PCR using primers CITXR and OSMTIR, was used as target DNA for band shift assays. To eliminate the T-N11-A binding motif, a crossover PCR deletion was done with oligos MUTUP and MUTDOWN, which contain a 20-bp-long overlapping region. Binding reactions were carried out in 20 μL of DNA-binding buffer (40 mM Tris-HCl, pH 8, 100 mM KCl, 1 mM dithiothreitol, 1 mM EDTA, 2 mM MgCl2, 5% glycerol) with 50 fmol of the corresponding Venetoclax cell line 32P-labeled DNA fragment and various amounts of the purified YfeRHis protein. The mixture was incubated

at 25 °C for 15 min and loaded onto a 5% polyacrylamide gel in Tris-Borate-EDTA buffer. The gels were electrophoresed at 100 V for 1 h and dried. The transcription start points were located with the 5′RACE system for rapid amplification of cDNA ends, version 2.0 (Invitrogen). Five micrograms of total RNA were reverse transcribed with GSP1 primers to copy mRNA into cDNA. After a dC- Navitoclax tailing reaction of cDNA a PCR amplification was carried out using a deoxyinosine-containing anchor primer, provided with the kit, and a GSP2 primer. To reduce the high background of nonspecific amplification, a second PCR was Cobimetinib price performed, using a nested anchor primer of the 5′RACE kit and GSP3 primers. The single DNA bands

for each gene resulting from this second PCR reaction were purified and sequenced. Transcriptomic analyses was performed on a DNA microarray engineered by the Salgenomics consortium of research groups. The Salgenomics microarray contained 6119 probes (including ORFs, RNA genes and intergenic regions) from the genome sequence of S. enterica serovar Typhimurium SL1344 and was developed using sequences from the Welcome Trust Sanger Institute. RNA was isolated from cultures of TT1704 and TT1704Y strains grown in LB 0 M NaCl until mid-exponential phase (OD600 nm=0.5). RNA extraction, retrotranscription, labeling, hybridization, microarray scanning and data analysis were performed as described elsewhere (Mariscotti & García-del Portillo, 2009). To search for osmoregulated genes, we first obtained a collection of 3000 random MudJ insertion mutants of strain TT1704. Clones exhibiting low osmolarity-dependent Lac+ phenotype conditions on indicator LB-X-Gal plates were selected. Evaluation of β-galactosidase activity in cell extracts confirmed lacZ osmoregulation. To show that osmoregulated lacZ expression was linked to the gene where MudJ was inserted, each MudJ insertion was backcrossed into a clean background.

subtilis strains such as strain FT-3 (Morita et al., 1979).

Although specific roles for these polysaccharides have not been proposed, they are known to be comprised of glucose, galactose, fucose, glucuronic acid and O-acetyl groups in an approximate molar ratio of 2 : 2 : 1 : 1 : 1.5 (Morita et al., 1979). Information regarding the genes encoding the proteins that make these exopolysaccharides is also limited. yhxB is a gene related to the synthesis of an uncharacterized exopolysaccharide component of the B. subtilis biofilm matrix and putatively encodes an α-phosphoglucomutase and/or phosphomannomutase (Branda et al., 2004). In B. subtilis 3610, a deletion in yhxB is responsible for the production of a fragile surface pellicle when grown in a liquid culture and flat undifferentiated colonies when grown on CDK activity solid media. On the contrary, the B. subtilis wild-type strain shows a robust pellicle in liquid culture and colonies on

plates with web-like structures (i.e. bundled structures). Other genes important in matrix structure and biofilm architecture include the 16 genes of the eps operon (yveK-yvfF) involved in polysaccharide biosynthesis, modification and export (Branda ERK inhibitor et al., 2001). From sequence comparisons, two genes belonging to the eps operon, named epsG (yveQ) and epsH (yveR), may be involved in the synthesis of exopolysaccharides. epsG encodes a protein that is presumably involved in EPS polymerization, while epsH encodes a glycosyl-transferase (Branda et al., 2001). eps mutants in B. subtilis 3610 show a reduction in the carbohydrate content and complexity of biofilm pellicle (Branda et al., 2006). Blair et al. (2008) have pheromone recently demonstrated that another member of this eps operon,

the EpsE protein, is an inhibitor of cell motility. Despite the extensive study of the eps operon and its role, the structure and function of the polysaccharides resulting from the expression of these genes remain unknown. Characterization of this polysaccharide and its regulation awaits further investigations. The second category of EPS secreted by B. subtilis includes a polymer, which plays a role in the sorption of ions and/or charged molecules. Poly-γ-glutamate (γ-PGA) produced by B. subtilis strain IFO3336 is a well-characterized anionic, nontoxic and biodegradable viscous polymer of d- and l-monomers with a molecular mass of over 10 000 kDa. The γ-PGA of B. subtilis (natto) is composed 50–80% of d- and 20–50% of l-glutamate (Ashiuchi et al., 1999; Morikawa et al., 2006; Inbaraj et al., 2008). γ-PGA is synthesized by several Bacillus species, especially wild-type isolates, including B. subtilis strains IFO3336, IFO3335, TAM-4, F-2-01, B-1 (mucoid-type colonies), ZJU-7, B. subtilis (natto) and Bacillus anthracis (Kubota et al., 1993; Kunioka, 1995; Ito et al., 1996; Shi et al., 2006). The pgsBCA genes are responsible for the synthesis of γ-PGA.

In chloroplasts, this enzyme has been exclusively localized to TMs (Soll et al., 1983; Eckhardt et al., 2004;Fig. 3). If this TM localization of CS also holds for cyanobacteria, TM-synthesized chlide a could be rapidly converted to chl a, whereas chlide Navitoclax manufacturer a synthesized by the minor POR fraction in PDMs would accumulate due to scarce further processing. However, previous CS activity measurements in Synechocystis 6803 suggested the presence of CS in both the – putatively PDM-related – thylakoid centers and TMs (Hinterstoisser et al., 1993). Hence, higher chlide a synthesis rates in PDMs must also be considered. These might be due to the activity of the second, light-independent, POR enzyme (LiPOR) from Synechocystis 6803,

whose localization is still elusive (Armstrong, 1998). Taken together and despite

several open questions, the facts presented draw a picture of PDMs as a subcompartment, in which not only protein complex biogenesis but also the later steps in chlorophyll synthesis and its insertion into polypeptides occur. In conclusion, we propose the following working model for the biogenesis of TMs in the model organism Synechocystis 6803 (Fig. 4): both protein synthesis/assembly and chlorophyll synthesis/insertion are subject to tight spatial organization. These two processes are localized in a specialized membrane region, here termed PDMs, which is marked by the D1-bound form of the PSII biogenesis factor PratA. The fact selleck chemical that non-D1-bound PratA is a soluble periplasmic protein strongly argues for at least temporary contacts of PDMs with the PM. These areas of contact are likely to be identical to the previously described thylakoid centers, which are located at the cell periphery, between PM and TMs (Hinterstoisser et al., 1993; van de Meene et al., 2006). Etomidate Hence, the existence of such structures close to both the PM and the TM could easily explain the involvement of the periplasmic PratA factor in TM biogenesis. Furthermore, the finding that pD1, Pitt and POR are all localized

to a higher amount in PDMs upon inactivation of PratA strongly suggests an essential role of PratA in the functional and/or structural organization of these biogenesis centers and, thus, membrane flow from PDMs to TMs. Although the described model seems to apply to PSII biogenesis, less evidence is available concerning the spatial organization of the PSI assembly process. Nevertheless, the detection of the PSI reaction center proteins PsaA and PsaB in PM or PM-related fractions suggests that also PSI biogenesis is initiated in the PM or even in PDMs similar to PSII (Zak et al., 2001). Future work will be directed toward the visualization of the biogenesis process, for instance by time-resolved studies with green fluorescent protein-tagged proteins. The ultrastructural localization of the various factors involved, especially the PratA protein, will unambiguously answer the question whether, indeed, PDMs and thylakoid centers are directly linked.