Interferences of cannabinoid agonists

and antagonists with synaptic transmission in the cortex may explain the cognitive and memory deficits elicited by these drugs. Neuropsychopharmacology (2012) 37, 1104-1114; doi:10.1038/npp.2011.262; published online 2 November 2011″
“Hepatic stellate cells (HSCs) represent the main fibrogenic cell type accumulating extracellular matrix in the liver. Recent data suggest that hepatitis C virus (HCV) core protein may directly activate HSCs. Therefore, we examined the influence of recombinant HCV core protein on human HSCs. Primary human HSCs and the human HSC line LX-2 were stimulated with recombinant VX-809 in vivo HCV proteins core and envelope 2 protein. Expression of procollagen type I alpha-1, alpha-smooth muscle actin, cysteine-and glycine-rich protein 2, glial fibrillary acidic protein, tissue growth factor beta 1, matrix metalloproteinases 2 (MMP2) and 13, tissue inhibitor of metalloproteinases 1 and 2 was investigated by real-time PCR. Intracellular signaling pathways of ERK1/2, p38 and, jun-amino-terminal

kinase (JNK) were analyzed by western blot analysis. Recombinant HCV core protein induced upregulation of procollagen type I alpha-1, alpha-smooth muscle actin, MMP 2 and 13, tissue inhibitor of metalloproteinases 1 and 2, tissue growth factor beta 1, cysteine-and glycine-rich protein 2, and glial fibrillary acidic protein mRNA expression, whereas HCV envelope 2 protein did not exert any significant effect. Blocking of toll-like Selonsertib datasheet receptor 2 (TLR2) with a neutralizing antibody prevented mRNA upregulation by HCV core protein confirming that the TLR2 pathway was involved. Furthermore, western blot analysis revealed HCV-induced phosphorylation of the TLR2-dependent signaling molecules ERK1/2, p38 and JNK mitogen-activated kinases. Our in vitro results demonstrate a direct effect of HCV core protein on activation of HSCs toward a profibrogenic OSBPL9 state, which is mediated via the TLR2 pathway. Manipulating the TLR2 pathway may thus provide

a new approach for antifibrotic therapies in HCV infection. Laboratory Investigation (2011) 91, 1375-1382; doi: 10.1038/labinvest. 2011.78; published online 2 May 2011″
“The analytical performance of MALDI-MS is highly influenced by sample preparation and the choice of matrix. Here we present an improved MALDI-MS sample preparation method for peptide mass mapping and peptide analysis, based on the use of the 2,5-dihydroxybenzoic acid matrix and prestructured sample supports, termed: matrix layer (ML). This sample preparation is easy to use and results in a rapid automated MALDI-MS and MS/MS with high quality spectra acquisition. The between-spot variation was investigated using standard peptides and statistical treatment of data confirmed the improvement gained with the ML method.

For the compression of an elastic sphere with radius of R, Hertzian theory predicts the

relationship between applied load F and compression depth δ as [26] (2) where E * is the reduced Young’s modulus of the sphere. In this paper, E * is fitted from the load versus compression depth relation in the elastic regime by this website Equation 2. For different twin spacing, the value of E * keeps almost the same as 287.4 GPa. It is seen that the elastic response of nanosphere under compression is determined mainly by the local elastic properties under indenter. Therefore, for a given loading direction, the change of twin spacing does not affect the overall elastic response of nanosphere. And the reduced modulus is much larger than the theoretical prediction 153 GPa of the bulk single crystal material in <111 > direction [27]. In nanowires and nanoparticles, improved

elastic modulus and yield stress have also been observed [5, 13]. However, the introduction of TBs plays an important role in plastic deformation. The first load-drop, as marked by arrows in Figure 2, indicates the appearance of initial yield. The local peak load corresponding to the first load-drop may be considered as the yield load. It is found that, when the twin spacing decreases from 5.09 to 1.25 nm, the yield load increases from 0.28 to 0.62 μN. In the further development of plasticity, the compression load of the twinned Fulvestrant datasheet nanosphere is significantly larger than that of the twin-free nanosphere for the same compression depth. The highly serrated load-compression response is indicative of dislocation activities inside the deformed nanospheres. 5-FU solubility dmso To estimate the influence of TBs qualitatively, the strain energy stored in nanospheres up to a given compression depth (δ/R = 53.3%) is also shown in Figure 3. It is found that, the strain energy of twinned nanospheres increases clearly as the twin spacing decreases, reaching its maximum at the twin spacing of 1.88 nm, and then declines with further decreasing

twin spacing. Such characteristics are similar to those in nanotwinned polycrystalline materials [4, 9]. Figure 3 Strain energy of the deformed nanosphere as a function of twin spacing up to δ / R  = 53.3%. In order to understand the underlying strengthening mechanisms, we examine the atomistic structures in plastic stage for several samples, as shown in Figure 4. For a twin-free nanosphere, the plastic deformation begins with the nucleation of partial dislocations from the contact edge, and the dislocations then glide on 111 slip planes. Without experiencing obstacles from TBs, most partial dislocations easily glide to the opposite surface and annihilate here, forming surface steps. This process exhausts VX-809 cost nucleated dislocations in nanosphere and reduces dislocation density, corresponding to the dislocation starvation mechanism.

Our data showed that the Epigenetics inhibitor expression of btuB was indeed reduced when E. coli cells were grown to stationary phase in an acidic medium as compared to the same cells grown in neutral medium (Table 4). The reduction in the production of btuB in response to acid stress probably represents a physiological regulatory mechanism of Entospletinib bacteria facing environmental challenges such as low pH. Under stress environment, bacteria need to alter their metabolism to adapt to the environmental change. The transportation of cobalamin by BtuB receptor is driven by proton motive force (PMF)[45]. Since the PMF of bacteria is increased at low pH[46], the cobalamin transportation may be

enhanced by increased PMF. The higher concentration of cobalamin in cytoplasm will initiate riboswitch mechanism to repress

the translation APR-246 datasheet of BtuB receptor, which is in good accord with the repression of btuB transcription by the acid-induced GadX for bacteria to decrease the production of BtuB in response to this acidic stress. Conclusions Through biological and biochemical analysis, we have demonstrated the GadX can directly interact with btuB promoter and affect the expression of btuB. When bacteria were grown to stationary phase in an acidic medium, the increased gadX expression would repress the btuB transcription to help bacteria to adapt to acidic shock. In conclusion, this study provides the first evidence that the expression of btuB gene is transcriptionally repressed by the acid responsive genes gadX and gadY. Methods Plasmid constructions To produce the His6-tagged ColE7/Im7 protein complex for the ColE7 resistance assay, pQE30ColE7-Im7 was constructed. The cea7-cei7 genes encoding the colicin E7 and immunity proteins, that form an active ColE7 complex, were amplified from plasmid K317 [47]

by PCR using primers F/cea7-BamHI and R/cei-PstI (Table 5). The 1,996-bp PCR product thus generated was inserted between BamHI and PstI sites of pQE30 (Qiagen), fusing the His6-tag to the N terminus of ColE7. For searching transcriptional regulators of btuB, a genomic library of E. coli K-12 strain constructed with the pGAD10 vector (Figure 1) was purchased from Clontech (catalog number XL4001AB) and transformed into E. coli strain DH5α. The plasmid pGadXY (Figure Osimertinib 1) was isolated from the library in this study. To investigate the effect of GadX on btuB expression, pGadX was constructed as follows. A 1,077-bp DNA fragment containing gadX was generated by PCR using pGadXY (Figure 1) as the template and the MATCHMAKER 5′ insert screening sequence 5′-TACCACTACAATGGATG-3′ (Clontech) and R/gadX-PstI (Table 5) as primers. This 1.1-kb PCR fragment was inserted into pGEM-TEasy (Promega) by TA cloning, generating pGEMgadX. The 1.1-kb fragment was then isolated from pGEMgadX by EcoRI digestion and inserted into the EcoRI site of pGAD10, resulting in pGadX (Figure 1).

Even after 24 Lorlatinib ic50 h, the viability (Figure 4A) and cell cycle profiles (Figure 4B) were not significantly different for RAW264.7 cells cultured in the absence or presence of FBS. The metabolic activity of RAW264.7 cells

increased after 24 h, but significantly more so in the presence than absence of FBS (Figure 4C), which we speculate was due to greater overall proliferation and number of cells in FBS-enriched medium. These results confirmed that, for at least 4 h, in vitro models of infection can be conducted under entirely non-germinating culture conditions without loss of host cell viability, cell cycle progression, or metabolic function. Figure 4 Effect of non-germinating conditions on RAW264.7 cell viability, cell cycle progression, and metabolic activity. RAW264.7 cells were incubated at 37° in DMEM in the presence (+, black bars) or absence (-, white bars) of FBS, and then evaluated at 4 or 24 h, as indicated, for viability (A), cell cycle progression (B), and metabolic activity (C). (A) The cells were assayed for PI uptake, as described

check details under Materials and Methods. The data are rendered as the relative PI uptake normalized at both 4 and 24 h to cells incubated in the absence of FBS. (B) The cells were analyzed for their cell cycle profiles, as described under Materials and Methods. The data are rendered as the relative numbers of cells in G2/M normalized at both 4 and 24 h to cells incubated in the absence of FBS. (C) The cells were analyzed for conversion of MTT to formazan. The data are rendered as the fold change of formazan production normalized at both 4 and 24

h to cells incubated in the absence of FBS. To generate the bar graphs, data TCL were combined from three independent experiments, each conducted in triplicate. Error bars indicate standard deviations. The P values were calculated to evaluate the statistical significance of the differences in viability (A), cell cycle progression (B), and metabolism (C) between cells cultured in the absence or presence of FBS. Germination state of spores does not alter the uptake by Selleckchem SAHA HDAC mammalian cells The demonstration that cultured RAW264.7 cells remained viable and functional in FBS-free cell culture medium did not directly address the possibility that spore uptake by mammalian cells might be substantially different under germinating and non-germinating cell culture conditions. To evaluate this issue, Alexa Fluor 488-labeled spores were incubated with RAW264.7, MH-S, or JAWSII cells (MOI 10) in the absence or presence of FBS (10%). After 5 or 60 min, intracellular spores were monitored using flow cytometry to measure cell associated fluorescence that was not sensitive to the membrane-impermeable, Alexa Fluor 488 quenching agent, trypan blue [46].

7 fold increase in osmotic stress conditions (data not shown). Therefore, the 16S rRNA gene was again used as the reference to see more determine the change in transcription levels of virulence-associated genes induced by stress relative to bacterial cells in the absence of any stress. As shown in Figure  2, the transcription of dnaJ www.selleckchem.com/products/tpx-0005.html and ciaB was not affected by heat stress and only slightly altered after exposure to the other stresses. A modest up-regulation was observed under oxidative stress (~2.7 and 2 fold

for ciaB and dnaJ, respectively, p < 0.05) while a modest down-regulation (~2.8 to 3.2 fold, p < 0.01) was observed for both genes under low nutrient or osmotic stresses. The transcription of htrA was moderately up-regulated under oxidative stress and slightly down-regulated under low nutrient stress, but the change was not statistically significant (p > 0.05). In contrast, transcription of htrA was up-regulated 2.5 fold under heat stress (p = 0.03) and down-regulated ~10 fold under osmotic stress (p < 0.01). Figure 2 qRT-PCR analysis of the impact of the various stresses on transcription of virulence-associated genes of C. jejuni . Total RNA was isolated, and the expression of ciaB, dnaJ and htrA was measured immediately after exposure to each stress. All data were normalized to the level of expression of the 16S rRNA gene and are presented relatively to the

non-stress control. Therefore, the non-stressed condition has YM155 mouse a fold value of 1. Data are representative of three independent experiments from three RNA extracts. Overall, the qRT-PCR experiments showed that the transcription of the three virulence-associated genes chosen was only slightly up-regulated under heat and oxidative stresses, but tended to be down-regulated

Farnesyltransferase under low nutrient and osmotic stresses, with htrA showing the most down-regulation in response to osmotic stress. Effect of htrA on the uptake of C. jejuni by A. castellanii and its intracellular survival We showed above that the transcription of at least one of the few virulence-associated genes tested (htrA) was affected by osmotic stress at a level that could be biologically significant (10 fold). Transcriptional regulation of virulence-associated genes upon pre-exposure to stress may affect interactions of C. jejuni with host cells, including phagocytosis and the ability of C. jejuni to survive in host cells after internalization. To determine whether this was the case for interactions with amoeba, we tested the biological importance of the stress-related gene for which we had observed the largest transcriptional variations (htrA) using the htrA mutant that was previously described [39]. Both bacterial uptake and intracellular survival were measured after interactions of 2 × 108 bacteria with amoeba at a multiplicity of infection of 100 for 3 h at 25°C (see Methods section for more details).

# Demographic data MDI exposure. year (*PPE) Biomonitoring MDA PRIMA-1MET values (at the time of sampling) Air monitoring. median value 5 ppb Immunological status Reported duration of resp. sympt (year). Lung function SPT MDI-HSA MDI-SIC MDI-HSA-specific antibodies Final clinical diagnosis Sex Age Smo-king status SPT comm. allerg. Total IgE kU/L FVC  % MDV3100 concentration pred FEV1  % pred. NS-BHR MDI-sIgE kU/L MDI-sIgG mg/L Group B: Workplace field controls; workers currently exposed to MDI  1 M 38 Yes 11.3 0.16 μg MDA/g Creatinine Neg. 39.3 –

98 84 n.d. n.d. n.d. <0.02 <3 RCI  2 M 43 Yes 10.1 0.90 μg MDA/g Creatinine. Neg. 42.9 – 102 98 n.d. n.d. n.d. <0.02 <3 RCI  3 M 33 Yes 8.2 (*) 0.30 μg MDA/g Creatinine Neg. 97.3 – 104 click here 84 n.d. n.d. n.d. 0.25 3.5 H  4 M 33 No 7.7 0.32 μg MDA/g Creatinine Neg. 37.7 – 97 88 n.d. n.d. n.d. <0.02 <3 CI  5 M 32 Yes 5.5 0.20 μg MDA/g Creatinine Neg. 13.3 – 109 91 n.d. n.d. n.d. <0.02 <3 CI  6 M 25 No 2.1 0.22 μg MDA/g Creatinine Pos. 28.6 – 96 92 n.d. n.d. n.d. <0.02 <3 RCIDI The six industrial workers involved in the production of MDI cont. coatings reported to have no respiratory symptoms (questioner) before being enrolled for the analysis. 5 showed RC/C symptoms after the work week, only one worker hat no measurable symptoms. Only

one worker was wearing the personal protective mask (PPE) during the whole work shift M, Male; F, Female; comm. allerg., common allergens; MDI exp. duration of work-related exposure to MDI; lag time, lag time since last exposure; resp. sympt, duration of reported respiratory symptoms;

FVC, forced vital capacity; FEV1, forced expiratory volume in 1 s; NSBHR, non-specific bronchial hyper-responsiveness; MDI-SIC, MDI-specific inhalation challenge; sIgE, MDI-specific IgE; sIgG, MDI-specific IgG. OAI, occupational MDI asthma; PI, MDI-induced hypersensitivity pneumonitis; DI, dermatitis, due to MDI; CI, conjunctivitis due Selleckchem Abiraterone to MDI; RCI, rhino-conjunctivities, due to MDI; n.d. not determined; H, healthy There was a linear correlation between both the IgE and IgG values collected with either our fluorescence immunoassay using in-vapor conjugates and the commercially available ImmunoCAPs (Phadia) analysis with r = 1.00 and r = 0.79 (for IgE and IgG, respectively). Because of this high correlation, one can presume that these commercial conjugates were made in-vapor. All positive and negative antibody values in reactive and non-reactive subjects correlated between the two CAP systems within a permissive assay variability of 0.5–20 % for the absolute sIgE values. For the IgG data, however, the values collected with commercial CAPs were up to 35 % higher (resulting in false-positive values in lower range).

4 and 5, respectively. The matrices shown here are representative for optimal PLX3397 mw growth conditions (low to medium light intensity depending on species, nutrient replete growth media and sampled during the exponential growth phase). The F 0 fluorescence matrices show prominent fluorescence emission features in cyanobacteria under orange-red excitation that are characteristic

of PBS (fluorescence emission around 650 nm from allophycocyanin) and Chla (680 nm) pigments. In contrast, the algal strains reflect the absorption buy AC220 of light by chlorophylls and carotenoids in the blue-green spectral region with a sharply defined emission related to Chla fluorescence. Fig. 4 F 0 excitation–emission matrices of a culture of each of the species included in this study. These cultures were sampled under nutrient replete growth conditions and had F v/F m values of 0.6–0.7. The matrices are normalized to the spectral maximum to facilitate PRT062607 manufacturer comparison

of spectral differences between the different species Fig. 5 F v/F m excitation–emission matrices for the cultures shown in Fig. 4 Despite the sharp distinction in F 0 profiles observed between algae and cyanobacteria, F v/F m matrices (Fig. 5) show relatively constant F v/F m in the Chla emission band in both cyanobacteria and algae. For algal fluorescence, the variable component extends along the whole excitation spectrum for emission from ~650 to at least 750 nm (the maximum measured). The excitation–emission patterns for the cyanobacterial cultures show a smoother transition from low to high F v/F m when emission wavelength increases towards the maximum of PSII Chla (680–690 nm), but a sharp drop of F v/F m at longer emission wavelengths (>700 nm). These features

can respectively be explained by a variable component to PBS fluorescence (discussed further below), and the allocation of most Chla molecules to the non-variable PSI in cyanobacteria (Johnsen and Sakshaug 1996, 2007). The feature-rich F v/F m profile of cyanobacteria implies that the spectral location and bandwidth of emission detection can have a major Vitamin B12 influence on readings of F v/F m, when we target Chla emission in cyanobacteria. Optimization of detector slit spectral position and bandwidth for equivalent readings of F v/F m in cyanobacteria and algae are discussed in more detail below. Simulations of community fluorescence F v/F m is used to assess the maximum efficiency of PSII in dark-acclimated cells. F v/F m can be expressed for all waveband pairs in the excitation/emission matrix, and because the fluorescence excitation–emission matrices of algae and cyanobacteria differ prominently (Fig.

23. Di Cristofano C, Minervini A, Menicagli M, Salinitri G, Bertacca G, Pefanis G, Masieri L, Lessi F, Collecchi P, Minervini R, Carini M, Bevilacqua G, Cavazzana A: Nuclear expression of hypoxia-inducible factor-1alpha in clear cell renal cell carcinoma is involved in tumor progression. Am J Surg Pathol 2007, 31: 1875–81.CrossRefPubMed 24. Klatte T, Seligson DB, Riggs SB, Leppert JT, Berkman MK, Kleid MD, Yu H, Kabbinavar FF, Pantuck AJ, Belldegrun AS: Hypoxia-inducible factor 1 alpha in clear cell renal cell carcinoma. Clin

Cancer Res 2007, 13: 7388–93.CrossRefPubMed 25. Kubis HP, Hanke Momelotinib price N, MK-4827 Scheibe RJ, Gros G: Accumulation and nuclear import of HIF1 alpha during high and low oxygen concentration in skeletal muscle cells in primary culture. Biochim Biophys Acta 2005, 1745 (2) : 187–195.CrossRefPubMed 26. Minervini A, Di Cristofano C, Serni S, Carini M, Lidgren Anders, Hedberg Ylva, Grankvist Kjell, Rasmuson Torgny, Bergh Anders, Ljungberg Börje: Hypoxia-inducible factor 1 alpha expression in renal cell carcinoma

analyzed by tissue microarray. Eur Urol 2006, 50: 1272–7. Eur Urol 2007, 51 :1451–2CrossRef 27. Bos R, van Diest PJ, de Jong JS, Groep P, Valk P, Wall E: Hypoxia-inducible factor-1alpha is associated with angiogenesis, and expression of bFGF, PDGF-BB, and EGFR in invasive breast cancer. Histopathology GDC-0941 molecular weight 2005, 46: 31–6.CrossRefPubMed 28. Lidgren A, Hedberg Y, Grankvist K, Rasmuson T, Bergh A, Ljungberg B: Hypoxia-inducible factor 1alpha expression in renal cell carcinoma analyzed by tissue microarray. Eur Urol 2006, 50: 1272–7.CrossRefPubMed 29. Moon EJ, Brizel DM, Chi JT, Dewhirst MW: The potential role of intrinsic hypoxia markers as prognostic variables in cancer. Antioxid Redox Signal 2007, 9: 1237–94.CrossRefPubMed Competing interests The authors declare that they have no competing interests. Authors’ contributions GĐ conceived of the study and drafted the manuscript. KMI participated in the design of the study, carried out the immunoassays and performed the statistical analysis. EB carried out the immunoassays, participated in the

sequence alignment and helped to draft the manuscript. IH, MG and BG carried out the molecular studies and participated in the sequence alignment. NJ conceived of the study, and participated in its design and coordination. All authors read and approved the final manuscript.”
“Introduction Aberrations Hydroxychloroquine chemical structure in regulation of a restricted number of key pathways that control cell proliferation and cell survival are mandatory for tumour growth and progression. Deregulated cell proliferation and suppressed apoptosis are both essential for cell transformation and sustained growth. Hematological neoplasia are considered “”special tumors”" for their high sensitivity to the occurrence of spontaneous and pharmacological apoptosis. These cancers origin by tissues that use apoptosis for the regulation of their physiological mechanisms. These considerations explain the high sensitivity of these diseases to chemotherapy.

The importance LY333531 of IL-27 in modulating EMT through the STAT pathways is poorly understood in carcinogenesis. To our knowledge, there have been no studies that have described MET as an anti-tumor mechanism of IL-27. In our study, we hypothesized that IL-27 FHPI inhibits EMT and angiogenesis through STAT dependent pathways. Our results revealed that IL-27-treated lung cancer cells show increased epithelial marker (E-cadherin and γ-catenin), decreased Snail (transcriptional repressor of E-cadherin), and decreased mesenchymal

marker (N-cadherin and vimentin) expression. In addition, IL-27 treatment suppressed in vitro cell migration. The ability of IL-27 to promote MET and inhibit cell migration was abolished by inhibition of the STAT1 pathway, but not the STAT3 pathway, with the exception of N-cadherin expression. The impact of N-cadherin and STAT3 in this process is unclear. Overall, our findings suggest that IL-27 promotes MET and the increased

expression of epithelial marker proteins is STAT1-dependent. The inhibition of EMT through STAT1 dependence is a novel anti-tumor mechanism of IL-27, which has not been previously described. Our results support the body of evidence that STAT1 is associated with tumor suppressive properties, such as inhibition of angiogenesis, tumor growth and metastasis as well as promotion of apoptosis [12, 16]. The role of STAT3 in IL-27 regulation of EMT is not well understood. In present study, the inhibition of STAT3 activation Ro 61-8048 did not reverse the increased expression of epithelial markers (E-cadherin and γ-catenin) and the reduced expression of mesenchymal marker (vimentin) and Snail by IL-27, and STAT3 activation was not required for the inhibition of cell

migration by IL-27. Interestingly, the inhibition of STAT1 activation Exoribonuclease led to increased STAT3 activation in IL-27 treated lung cancer cells whereas inhibition of STAT3 activation alone did not significantly impact STAT1 expression. The current study does not provide a mechanism by which inhibition of STAT1 led to increased STAT3 activation. However, similar to our results, previous studies have demonstrated that STAT1- deficient cells showed increased STAT3 activation [59–61]. Potential mechanisms by which STAT1 may directly inhibit STAT3 include competition for receptor docking sites, promoters of target DNA sequences, and/or binding cofactors. The receptor docking site is a prerequisite for activation by tyrosine phosphorylation and STAT3 can be phosphorylated by receptor bound tyrosine kinases [62, 63]. In fact, it has been shown that STAT1 suppresses STAT3 tyrosine phosphorylation that mediates downstream signaling of other cytokine receptors [60]. Thus it appears likely that STAT1 suppresses IL27-mediated STAT3 activation at least in part by competing for the STAT docking site within the IL-27 receptor cytoplasmic domain.

The resulting tree from the MrBayes analysis revealed several subgroups among the hydrogenase specific proteases, which correlates with respective hydrogenase group according to Vignais et al [25] (Figure 1); Figure 1 Unrooted phylogenetic tree of hydrogenase this website specific proteases. The phylogenetic tree of hydrogenase specific click here proteases from the MrBayes analysis including the different subgroups they may

be divided into. The proposed subgroups for each protease are marked in the figure; 1 (red), 2 (orange), 3a (blue), 3d (purple), 4 (green) and unknown (black). X: The point in the phylogenetic tree when horizontal gene transfer occurred. Y/Z: Suggested positions of root. B. The phylogenetic tree of hydrogenases adapted from Vignais et al 2004 [25]. Type 2a (HupL) and 3d (HoxH) hydrogenases

which can be found in cyanobacteria are marked in bold. The phylogenetic tree was obtained using MrBayes analyses and the claude credibility STAT inhibitor values are given beside each branch. For abbreviations see Table 2. 1. Bacterial proteases (cleaves group 1 hydrogenases) 2. Cyanobacterial proteases, HupW type (cleaves group 2 hydrogenases) 3. Bacterial and Archaean proteases a. Archean proteases (cleaves group 3a hydrogenases) d. Bacterial proteases, HoxW type (cleaves group 3d hydrogenases) 4. Bacterial and Archaean proteases, Hyc type (cleaves group 4 hydrogenases) The phylogenetic groups of the hydrogenase specific protease have been named according to the nomenclature used for [NiFe]-hydrogenase. The result from the

PAUP analysis is less resolved but supports the result from MrBayers analysis with some minor differences within group 3d (HoxW in Synechocysis sp. strain PCC 6803 and HoxW in Synechococcus sp. strain PCC 7002 are shown as more closely related). An extended phylogenetic tree was also constructed containing more strains including hydrogenase specific proteases cleaving Thiamet G type 3b-hydrogenases. This tree was unfortunately less reliable and far from robust with several weak nodes (Additional file 1 and Additional file 2). However the result showed putative group 1 proteases and putative group 3b proteases as less clustered and instead spread around point X (Figure 1 and Additional file 1). Transcriptional studies of hupW in Nostoc punctiforme ATCC 29133 and Nostoc sp strain PCC 7120 Northern hybridisations were performed of hupW in both Nostoc punctiforme and Nostoc PCC 7120 using both N2-fixing and non N2-fixing cultures (Figure 2). The results from Nostoc PCC 7120 revealed two transcripts. The first is shorter (approx. 500 nt) and present under both N2-fixing and non N2-fixing conditions, while the second longer transcript (approx. 1600 nt) is only present under N2-fixing conditions. The size of the longer transcript is comparable with the size of a two-gene operon containing hupW together with the upstream gene alr1422, a gene of unknown function (Figure 3a). RT-PCR confirmed that the two genes are co transcribed (Figure 3a).